To see the other types of publications on this topic, follow the link: Cloning tyrosine kinase genes.
Journal articles on the topic 'Cloning tyrosine kinase genes'
Create a spot-on reference in APA, MLA, Chicago, Harvard, and other styles
Consult the top 50 journal articles for your research on the topic 'Cloning tyrosine kinase genes.'
Next to every source in the list of references, there is an 'Add to bibliography' button. Press on it, and we will generate automatically the bibliographic reference to the chosen work in the citation style you need: APA, MLA, Harvard, Chicago, Vancouver, etc.
You can also download the full text of the academic publication as pdf and read online its abstract whenever available in the metadata.
Browse journal articles on a wide variety of disciplines and organise your bibliography correctly.
1
Beeler, J. F., W. J. LaRochelle, M. Chedid, S. R. Tronick, and S. A. Aaronson. "Prokaryotic expression cloning of a novel human tyrosine kinase." Molecular and Cellular Biology 14, no. 2 (February 1994): 982–88. http://dx.doi.org/10.1128/mcb.14.2.982.
Full textAbstract:
Screening of a human embryonic lung fibroblast cDNA expression library with antiphosphotyrosine antibodies led to isolation of a novel protein kinase. A clone, designated A6, contained a 3-kb cDNA insert with a predicted open reading frame of 350 amino acids. DNA sequence analysis failed to reveal any detectable similarity with previously known genes, and the predicted A6 protein lacked any of the motifs commonly conserved in the catalytic domains of protein kinases. However, the bacterially expressed beta-galactosidase-A6 fusion protein demonstrated both tyrosine and serine phosphorylation in an in vitro kinase assay and phosphorylated exogenous substrates including myelin basic protein specifically on tyrosine residues. The enzyme also displayed biochemical properties analogous to those of other protein tyrosine kinases. The A6 gene was found to be expressed widely at the transcript level in normal tissues and was evolutionarily conserved. Thus, A6 represents a novel tyrosine kinase which is highly divergent from previously described members of this important class of regulatory molecules.
2
Beeler, J. F., W. J. LaRochelle, M. Chedid, S. R. Tronick, and S. A. Aaronson. "Prokaryotic expression cloning of a novel human tyrosine kinase." Molecular and Cellular Biology 14, no. 2 (February 1994): 982–88. http://dx.doi.org/10.1128/mcb.14.2.982-988.1994.
Full textAbstract:
Screening of a human embryonic lung fibroblast cDNA expression library with antiphosphotyrosine antibodies led to isolation of a novel protein kinase. A clone, designated A6, contained a 3-kb cDNA insert with a predicted open reading frame of 350 amino acids. DNA sequence analysis failed to reveal any detectable similarity with previously known genes, and the predicted A6 protein lacked any of the motifs commonly conserved in the catalytic domains of protein kinases. However, the bacterially expressed beta-galactosidase-A6 fusion protein demonstrated both tyrosine and serine phosphorylation in an in vitro kinase assay and phosphorylated exogenous substrates including myelin basic protein specifically on tyrosine residues. The enzyme also displayed biochemical properties analogous to those of other protein tyrosine kinases. The A6 gene was found to be expressed widely at the transcript level in normal tissues and was evolutionarily conserved. Thus, A6 represents a novel tyrosine kinase which is highly divergent from previously described members of this important class of regulatory molecules.
3
Gibson, S., B. Leung, JA Squire, M. Hill, N. Arima, P. Goss, D. Hogg, and GB Mills. "Identification, cloning, and characterization of a novel human T-cell- specific tyrosine kinase located at the hematopoietin complex on chromosome 5q." Blood 82, no. 5 (September 1, 1993): 1561–72. http://dx.doi.org/10.1182/blood.v82.5.1561.1561.
Full textAbstract:
Abstract Signal transduction through the T-cell receptor and cytokine receptors on the surface of T lymphocytes occurs largely via tyrosine phosphorylation of intracellular substrates. Because neither the T-cell receptor nor cytokine receptors contain intrinsic kinase domains, signal transduction is thought to occur via association of these receptors with intracellular protein tyrosine kinases. Although several members of the SRC and SYK families of tyrosine kinases have been implicated in signal transduction in lymphocytes, it seems likely that additional tyrosine kinases involved in signal transduction remain to be identified. To identify unique T-cell tyrosine kinases, we used polymerase chain reaction-based cloning with degenerate oligonucleotides directed at highly conserved motifs of tyrosine kinase domains. We have cloned the complete cDNA for a unique human tyrosine kinase that is expressed mainly in T lymphocytes (EMT) and natural killer (NK) cells. The cDNA of EMT predicts an open reading frame of 1866 bp encoding a protein with a predicted size of 72 Kd, which is in keeping with its size on Western blotting. A single 6.2-kb EMT mRNA and 72-Kd protein were detected in T lymphocytes and NK-like cell lines, but were not detected in other cell lineages. EMT contains both SH2 and SH3 domains, as do many other intracellular kinases. EMT does not contain the N-terminal myristylation site or the negative regulatory tyrosine phosphorylation site in its carboxyterminus that are found in the SRC family of tyrosine kinases. EMT is related to the B-cell progenitor kinase (BPK), which has recently been implicated in X-linked hypogammaglobulinemia, to the TECI mammalian kinase, which has been implicated in liver neoplasia, to the more widely expressed TECII mammalian kinase, and to the Drosophila melanogaster Dsrc28 kinase. Sequence comparison suggests that EMT is likely the human homologue of a recently identified murine interleukin-2 (IL-2)-inducible T cell kinase (ITK). However, unlike ITK, EMT message and protein levels do not vary markedly on stimulation of human IL-2-responsive T cells with IL-2. Taken together, it seems that EMT is a member of a new family of intracellular kinases that includes BPK, TECI, and TECII. EMT was localized to chromosome 5q31–32, a region that contains the genes for several growth factors and receptors as well as early activation genes, particularly those involved in the hematopoietic system. Furthermore, the 5q31–32 region is implicated in the genesis of the 5q- syndrome associated with myelodysplasia and development of leukemia.(ABSTRACT TRUNCATED AT 400 WORDS)
4
Gibson, S., B. Leung, JA Squire, M. Hill, N. Arima, P. Goss, D. Hogg, and GB Mills. "Identification, cloning, and characterization of a novel human T-cell- specific tyrosine kinase located at the hematopoietin complex on chromosome 5q." Blood 82, no. 5 (September 1, 1993): 1561–72. http://dx.doi.org/10.1182/blood.v82.5.1561.bloodjournal8251561.
Full textAbstract:
Signal transduction through the T-cell receptor and cytokine receptors on the surface of T lymphocytes occurs largely via tyrosine phosphorylation of intracellular substrates. Because neither the T-cell receptor nor cytokine receptors contain intrinsic kinase domains, signal transduction is thought to occur via association of these receptors with intracellular protein tyrosine kinases. Although several members of the SRC and SYK families of tyrosine kinases have been implicated in signal transduction in lymphocytes, it seems likely that additional tyrosine kinases involved in signal transduction remain to be identified. To identify unique T-cell tyrosine kinases, we used polymerase chain reaction-based cloning with degenerate oligonucleotides directed at highly conserved motifs of tyrosine kinase domains. We have cloned the complete cDNA for a unique human tyrosine kinase that is expressed mainly in T lymphocytes (EMT) and natural killer (NK) cells. The cDNA of EMT predicts an open reading frame of 1866 bp encoding a protein with a predicted size of 72 Kd, which is in keeping with its size on Western blotting. A single 6.2-kb EMT mRNA and 72-Kd protein were detected in T lymphocytes and NK-like cell lines, but were not detected in other cell lineages. EMT contains both SH2 and SH3 domains, as do many other intracellular kinases. EMT does not contain the N-terminal myristylation site or the negative regulatory tyrosine phosphorylation site in its carboxyterminus that are found in the SRC family of tyrosine kinases. EMT is related to the B-cell progenitor kinase (BPK), which has recently been implicated in X-linked hypogammaglobulinemia, to the TECI mammalian kinase, which has been implicated in liver neoplasia, to the more widely expressed TECII mammalian kinase, and to the Drosophila melanogaster Dsrc28 kinase. Sequence comparison suggests that EMT is likely the human homologue of a recently identified murine interleukin-2 (IL-2)-inducible T cell kinase (ITK). However, unlike ITK, EMT message and protein levels do not vary markedly on stimulation of human IL-2-responsive T cells with IL-2. Taken together, it seems that EMT is a member of a new family of intracellular kinases that includes BPK, TECI, and TECII. EMT was localized to chromosome 5q31–32, a region that contains the genes for several growth factors and receptors as well as early activation genes, particularly those involved in the hematopoietic system. Furthermore, the 5q31–32 region is implicated in the genesis of the 5q- syndrome associated with myelodysplasia and development of leukemia.(ABSTRACT TRUNCATED AT 400 WORDS)
5
Neale, Jennifer C. C., Thomas P. Kenny, and M. Eric Gershwin. "Cloning and Sequencing of Protein Kinase cDNA from Harbor Seal (Phoca vitulina) Lymphocytes." Clinical and Developmental Immunology 11, no. 2 (2004): 157–63. http://dx.doi.org/10.1080/10446670410001722195.
Full textAbstract:
Protein kinases (PKs) play critical roles in signal transduction and activation of lymphocytes. The identification of PK genes provides a tool for understanding mechanisms of immunotoxic xenobiotics. As part of a larger study investigating persistent organic pollutants in the harbor seal and their possible immunomodulatory actions, we sequenced harbor seal cDNA fragments encoding PKs. The procedure, using degenerate primers based on conserved motifs of human protein tyrosine kinases (PTKs), successfully amplified nine phocid PK gene fragments with high homology to human and rodent orthologs. We identified eight PTKs and one dual (serine/threonine and tyrosine) kinase. Among these were several PKs important in early signaling events through the B- and T-cell receptors (FYN, LYN, ITK and SYK) and a MAP kinase involved in downstream signal transduction. V-FGR, RET and DDR2 were also expressed. Sequential activation of protein kinases ultimately induces gene transcription leading to the proliferation and differentiation of lymphocytes critical to adaptive immunity. PKs are potential targets of bioactive xenobiotics, including persistent organic pollutants of the marine environment; characterization of these molecules in the harbor seal provides a foundation for further research illuminating mechanisms of action of contaminants speculated to contribute to large-scale die-offs of marine mammals via immunosuppression.
6
Iwama, A., K. Okano, T. Sudo, Y. Matsuda, and T. Suda. "Molecular cloning of a novel receptor tyrosine kinase gene, STK, derived from enriched hematopoietic stem cells." Blood 83, no. 11 (June 1, 1994): 3160–69. http://dx.doi.org/10.1182/blood.v83.11.3160.3160.
Full textAbstract:
Abstract To identify the novel receptor tyrosine kinases (RTKs) critical to the proliferation of hematopoietic stem cells, we performed polymerase chain reaction-based cloning from highly purified murine hematopoietic stem cells. Lineage marker-negative, c-KIT-positive, and Ly6A/E- or Sca- 1-positive (Lin-c-KIT+Sca-1+) cells were sorted by a fluorescence- activated cell sorter. Two sets of degenerate oligonucleotide primers were directed to the conserved sequences of the catalytic domain, and were used to amplify cDNAs that encode protein tyrosine kinases (PTKs). One hundred cDNA clones were sequenced and 8 RTKs were identified, as well as 12 non-RTKs and 2 serine/threonine kinases. Sixteen cDNAs were identical to the known kinase genes (PKC beta, JAK-1, JAK-2, TYK-2, HCK, FGR, FYN, BLK, c-FES, FER, c-ABL, c-KIT, FLK-1, FLK-2, IGF1R, and ECK). Six novel cDNA sequences (stk series) were identified. However, three of them turned out to be BPK, RYK, and TEK. The remaining three showed high homology to S6 kinase II, JAK-2, and v-SEA/c-MET, respectively. Characterization of full-length cDNA sequence of the v- SEA/cMET-related gene showed that this was a novel RTK gene and we named this gene STK (stem cell-derived tyrosine kinase). We identified two distinct forms of STK cDNA; the short one encoded a putative truncated protein that lacked most of the extracellular domain. STK was expressed at various stages of hematopoietic cells, including stem cells, but we could not detect any apparent expression in other adult tissues. The expression of the truncated form of mRNA was more predominant than that of the complete form. STK was assigned by fluorescent in situ hybridization to the R-positive F1 band of chromosome 9, the same region to which hepatic growth factor-like protein has been assigned. Characterization of these PTKs, including STK, will be helpful to elucidate the molecular mechanism of the growth regulation of hematopoietic stem cells.
7
Iwama, A., K. Okano, T. Sudo, Y. Matsuda, and T. Suda. "Molecular cloning of a novel receptor tyrosine kinase gene, STK, derived from enriched hematopoietic stem cells." Blood 83, no. 11 (June 1, 1994): 3160–69. http://dx.doi.org/10.1182/blood.v83.11.3160.bloodjournal83113160.
Full textAbstract:
To identify the novel receptor tyrosine kinases (RTKs) critical to the proliferation of hematopoietic stem cells, we performed polymerase chain reaction-based cloning from highly purified murine hematopoietic stem cells. Lineage marker-negative, c-KIT-positive, and Ly6A/E- or Sca- 1-positive (Lin-c-KIT+Sca-1+) cells were sorted by a fluorescence- activated cell sorter. Two sets of degenerate oligonucleotide primers were directed to the conserved sequences of the catalytic domain, and were used to amplify cDNAs that encode protein tyrosine kinases (PTKs). One hundred cDNA clones were sequenced and 8 RTKs were identified, as well as 12 non-RTKs and 2 serine/threonine kinases. Sixteen cDNAs were identical to the known kinase genes (PKC beta, JAK-1, JAK-2, TYK-2, HCK, FGR, FYN, BLK, c-FES, FER, c-ABL, c-KIT, FLK-1, FLK-2, IGF1R, and ECK). Six novel cDNA sequences (stk series) were identified. However, three of them turned out to be BPK, RYK, and TEK. The remaining three showed high homology to S6 kinase II, JAK-2, and v-SEA/c-MET, respectively. Characterization of full-length cDNA sequence of the v- SEA/cMET-related gene showed that this was a novel RTK gene and we named this gene STK (stem cell-derived tyrosine kinase). We identified two distinct forms of STK cDNA; the short one encoded a putative truncated protein that lacked most of the extracellular domain. STK was expressed at various stages of hematopoietic cells, including stem cells, but we could not detect any apparent expression in other adult tissues. The expression of the truncated form of mRNA was more predominant than that of the complete form. STK was assigned by fluorescent in situ hybridization to the R-positive F1 band of chromosome 9, the same region to which hepatic growth factor-like protein has been assigned. Characterization of these PTKs, including STK, will be helpful to elucidate the molecular mechanism of the growth regulation of hematopoietic stem cells.
8
Guasch, Géraldine, Cornel Popovici, Francine Mugneret, Max Chaffanet, Pierre Pontarotti, Daniel Birnbaum, and Marie-Josèphe Pébusque. "Endogenous retroviral sequence is fused to FGFR1 kinase in the 8p12 stem-cell myeloproliferative disorder with t(8;19)(p12;q13.3)." Blood 101, no. 1 (January 1, 2003): 286–88. http://dx.doi.org/10.1182/blood-2002-02-0577.
Full textAbstract:
Abstract FGFR1, a transmembrane receptor tyrosine kinase for fibroblast growth factors, is constitutively activated by chromosomal translocations in an atypical stem-cell myeloproliferative disorder. The FGFR1 tyrosine domain is fused to dimerization domains encoded by 4 alternative genes: FOP at 6q27, CEP110 at 9q33,FIM/ZNF198 at 13q12, and BCR at 22q11. In this study, we report the molecular cloning of the t(8;19)(p12;q13.3), the fifth translocation associated with this syndrome. Reverse transcriptase–polymerase chain reaction (RT-PCR) analysis and fluorescence in situ hybridization (FISH) demonstrated that the translocation resulted in a long terminal repeat of human endogenous retrovirus gene (HERV-K)/fibroblast growth factor receptor 1 (FGFR1) fusion transcript that incorporated 5′ sequences from HERV-K fused in frame to 3′ FGFR1 sequences encoding the kinase domain. RT-PCR detected only 1 of the 2 possible fusion transcripts,HERV-K/FGFR1.
9
Guasch, Géraldine, Gary J. Mack, Cornel Popovici, Nicole Dastugue, Daniel Birnbaum, Jérome B. Rattner, and Marie-Josèphe Pébusque. "FGFR1 is fused to the centrosome-associated proteinCEP110 in the 8p12 stem cell myeloproliferative disorder with t(8;9)(p12;q33)." Blood 95, no. 5 (March 1, 2000): 1788–96. http://dx.doi.org/10.1182/blood.v95.5.1788.005k15_1788_1796.
Full textAbstract:
The hallmark of the 8p12 stem cell myeloproliferative disorder (MPD) is the disruption of the FGFR1 gene, which encodes a tyrosine kinase receptor for members of the fibroblast growth factor family.FGFR1 can be fused to at least 3 partner genes at chromosomal regions 6q27, 9q33, or 13q12. We report here the cloning of the t(8;9)(p12;q33) and the detection of a novel fusion betweenFGFR1 and the CEP110 gene, which codes for a novel centrosome-associated protein with a unique cell-cycle distribution. CEP110 is widely expressed at various levels in different tissues and is predicted to encode a 994-amino acid coiled-coil protein with 4 consensus leucine zippers [L-X(6)-L-X(6)-L-X(6)-L]. Both reciprocal fusion transcripts are expressed in the patient's cells. The CEP110-FGFR1 fusion protein encodes an aberrant tyrosine kinase of circa 150-kd, which retains most of CEP110 with the leucine zipper motifs and the catalytic domain of FGFR1. Transient expression studies show that the CEP110-FGFR1 protein has a constitutive kinase activity and is located within the cell cytoplasm.
10
Benini, Stefano, Lorenzo Caputi, and Michele Cianci. "Cloning, purification, crystallization and 1.57 Å resolution X-ray data analysis of AmsI, the tyrosine phosphatase controlling amylovoran biosynthesis in the plant pathogenErwinia amylovora." Acta Crystallographica Section F Structural Biology Communications 70, no. 12 (November 28, 2014): 1693–96. http://dx.doi.org/10.1107/s2053230x14024947.
Full textAbstract:
The Gram-negative bacteriumErwinia amylovorais a destructive pathogen of plants belonging to the Rosaceae family. Amongst its pathogenicity factors,E. amylovoraproduces the exopolysaccharide amylovoran, which contributes to the occlusion of plant vessels, causing wilting of shoots and eventually resulting in plant death. Amylovoran biosynthesis requires the presence of 12 genes (fromamsA toamsL) clustered in theamsregion of theE. amylovoragenome. They mostly encode glycosyl transferases (AmsG, AmsB, AmsD, AmsE, AmsJ and AmsK), proteins involved in amylovoran translocation and assembly (AmsH, AmsL and AmsC), and also a tyrosine kinase (AmsA) and a tyrosine phosphatase (AmsI), which are both involved in the regulation of amylovoran biosynthesis. The low-molecular-weight protein tyrosine phosphatase AmsI was overexpressed as a His6-tagged protein inEscherichia coli, purified and crystallized. X-ray diffraction data were collected to a maximum resolution of 1.57 Å in space groupP3121.
11
Shiota, M., J. Fujimoto, M. Takenaga, H. Satoh, R. Ichinohasama, M. Abe, M. Nakano, T. Yamamoto, and S. Mori. "Diagnosis of t(2;5)(p23;q35)-associated Ki-1 lymphoma with immunohistochemistry." Blood 84, no. 11 (December 1, 1994): 3648–52. http://dx.doi.org/10.1182/blood.v84.11.3648.bloodjournal84113648.
Full textAbstract:
Some Ki-1 lymphomas carry a specific chromosomal translocation, t(2;5)(p23;q35). We have recently found a novel hyperphosphorylated 80- kD protein tyrosine kinase, p80, in a human Ki-1 lymphoma with this translocation. Subsequent cDNA cloning showed that p80 is a fusion protein of two different genes on chromosome 2p23 and 5q35, the novel tyrosine kinase gene and nucleophosmin gene, respectively. In this study, we intended to detect p80 on lymphoma tissues with immunologic methods. Thus, we developed rabbit polyclonal antibody using a synthetic peptide corresponding to a part of its kinase domain. The antibody (anti-p80) immunoprecipitated and immunoblotted p80 specifically from AMS3. Then, to examine whether t(2;5)(p23;q35) was present on biopsied lymphomas, reverse transcriptase-polymerase chain reaction (RT-PCR) covering the fusion junction of p80 mRNA was performed. Among 10 Ki-1 lymphomas and 10 additional lymphomas other than the Ki-1 lymphomas, expression of p80 mRNA was detected in three cases exclusively. When these 20 cases and additional 30 lymphomas were immunostained with anti-p80, positive staining was noted exclusively in the three cases found by PCR to have harbored the p80 mRNA. Thus, the present immunostaining, as well as PCR, was shown to be efficient for detecting lymphomas producing this chimeric protein/mRNA.
12
Shiota, M., S. Nakamura, R. Ichinohasama, M. Abe, T. Akagi, M. Takeshita, N. Mori, J. Fujimoto, J. Miyauchi, and A. Mikata. "Anaplastic large cell lymphomas expressing the novel chimeric protein p80NPM/ALK: a distinct clinicopathologic entity." Blood 86, no. 5 (September 1, 1995): 1954–60. http://dx.doi.org/10.1182/blood.v86.5.1954.bloodjournal8651954.
Full textAbstract:
Anaplastic large cell lymphoma (ALCL) is a subtype of non-Hodgkin's lymphoma characterized by the CD30+ large neoplastic cells and sometimes carries a t(2;5)(p23;q35). Recently, we found a novel hyperphosphorylated 80-kD protein tyrosine kinase, p80, in ALCLs with t(2;5). Subsequent cDNA cloning showed p80 to be a fusion protein of two genes, the novel tyrosine kinase gene and the nucleophosmin gene, in accordance with the sequence of the NPM/ALK gene (Morris et al, Science 263:1281, 1994). Meanwhile, the clinicopathologic features of p80-carrying ALCLs have remained unclear. Paraffin sections of 105 cases of ALCL were immunostained using anti-p80 antibody, and 30 of them were shown to express p80. Clinicopathologic comparison between p80-positive and -negative ALCLs showed that p80-positive cases occurred in a far younger patient age group (16.2 +/- 12.9 years; p80- negative cases, 51.0 +/- 22.3 years; P < .0001) and the patients showed a far better 5-year survival rate (79.8%; p80-negative group, 32.9%; P < .01). These data showed that p80-positive ALCL is a distinct entity both clinically and pathogenetically and should be differentiated from p80-negative ALCL.
13
Iwama, A., I. Hamaguchi, M. Hashiyama, Y. Murayama, K. Yasunaga, and T. Suda. "Molecular Cloning and Characterization of Mouse TIE and TEK Receptor Tyrosine Kinase Genes and Their Expression in Hematopoietic Stem Cells." Biochemical and Biophysical Research Communications 195, no. 1 (August 1993): 301–9. http://dx.doi.org/10.1006/bbrc.1993.2045.
Full text
14
Peeters, Pieter, Sophie D. Raynaud, Jan Cools, Iwona Wlodarska, Josiane Grosgeorge, Patrick Philip, Fabrice Monpoux, et al. "Fusion of TEL, the ETS-Variant Gene 6 (ETV6), to the Receptor-Associated Kinase JAK2 as a Result of t(9; 12) in a Lymphoid and t(9; 15; 12) in a Myeloid Leukemia." Blood 90, no. 7 (October 1, 1997): 2535–40. http://dx.doi.org/10.1182/blood.v90.7.2535.
Full textAbstract:
Abstract Translocations in hematologic disease of myeloid or lymphoid origin with breakpoints at chromosome band 12p13 frequently result in rearrangements of the Ets variant gene 6 (ETV6). As a consequence either the ETS DNA-binding domain or the Helix-Loop-Helix (HLH) oligomerization domain of ETV6 is fused to different partner genes. We show here that a t(9; 12)(p24; p13) in a case of early pre-B acute lymphoid leukemia and a t(9; 15; 12)(p24; q15; p13) in atypical chronic myelogenous leukemia in transformation involve the ETV6 gene at 12p13 and the JAK2 gene at 9p24. In each case different fusion mRNAs were found, with only one resulting in an open reading frame for a chimeric protein consisting of the HLH oligomerization domain of ETV6 and the protein tyrosine kinase (PTK) domain of JAK2. The cloning of the complete human JAK2 coding and genomic sequences and of the genomic junction fragments of the translocations allowed a characterization of the different splice events leading to the various mRNAs. JAK2 plays a central role in non–protein tyrosine kinase receptor signaling pathways, which could explain its involvement in malignancies of different hematologic lineages. Besides hop in Drosophila no member of the JAK family has yet been implicated in tumorigenesis.
15
Peeters, Pieter, Sophie D. Raynaud, Jan Cools, Iwona Wlodarska, Josiane Grosgeorge, Patrick Philip, Fabrice Monpoux, et al. "Fusion of TEL, the ETS-Variant Gene 6 (ETV6), to the Receptor-Associated Kinase JAK2 as a Result of t(9; 12) in a Lymphoid and t(9; 15; 12) in a Myeloid Leukemia." Blood 90, no. 7 (October 1, 1997): 2535–40. http://dx.doi.org/10.1182/blood.v90.7.2535.2535_2535_2540.
Full textAbstract:
Translocations in hematologic disease of myeloid or lymphoid origin with breakpoints at chromosome band 12p13 frequently result in rearrangements of the Ets variant gene 6 (ETV6). As a consequence either the ETS DNA-binding domain or the Helix-Loop-Helix (HLH) oligomerization domain of ETV6 is fused to different partner genes. We show here that a t(9; 12)(p24; p13) in a case of early pre-B acute lymphoid leukemia and a t(9; 15; 12)(p24; q15; p13) in atypical chronic myelogenous leukemia in transformation involve the ETV6 gene at 12p13 and the JAK2 gene at 9p24. In each case different fusion mRNAs were found, with only one resulting in an open reading frame for a chimeric protein consisting of the HLH oligomerization domain of ETV6 and the protein tyrosine kinase (PTK) domain of JAK2. The cloning of the complete human JAK2 coding and genomic sequences and of the genomic junction fragments of the translocations allowed a characterization of the different splice events leading to the various mRNAs. JAK2 plays a central role in non–protein tyrosine kinase receptor signaling pathways, which could explain its involvement in malignancies of different hematologic lineages. Besides hop in Drosophila no member of the JAK family has yet been implicated in tumorigenesis.
16
Yi, T. L., J. B. Bolen, and J. N. Ihle. "Hematopoietic cells express two forms of lyn kinase differing by 21 amino acids in the amino terminus." Molecular and Cellular Biology 11, no. 5 (May 1991): 2391–98. http://dx.doi.org/10.1128/mcb.11.5.2391.
Full textAbstract:
cDNAs for the murine lyn protein tyrosine kinase gene were cloned from mouse bone marrow-derived monocytic cells. Comparison of the human and murine genes demonstrated a 94% homology in peptide sequence. Comparable to the human gene, murine lyn was found to be expressed in myeloid and B-lymphoid lineage cells. During the cloning, two types of cDNAs were obtained that differed by the presence (lynA) or absence (lynB) of 63 bp within the amino-terminal coding region of the gene. The genomic structure of the murine lyn gene demonstrates that the two types of lyn transcripts are derived from alternative splicing utilizing an internal splice donor site. Transcripts for both forms were found to be expressed in myeloid cells. lyn-specific antisera detected comparable levels of proteins of 56 and 53 kDa in hematopoietic cells. these 56- and 53-kDa proteins comigrated with proteins produced by in vitro translation or in vivo expression of the lynA and lynB cDNAs, respectively. The two forms had comparable in vitro kinase activities in immunoprecipitates and showed similar peptide patterns, with partial V8 digestion of the in vitro-phosphorylated proteins. The potential significance of the two lyn proteins is discussed.
17
Yi, T. L., J. B. Bolen, and J. N. Ihle. "Hematopoietic cells express two forms of lyn kinase differing by 21 amino acids in the amino terminus." Molecular and Cellular Biology 11, no. 5 (May 1991): 2391–98. http://dx.doi.org/10.1128/mcb.11.5.2391-2398.1991.
Full textAbstract:
cDNAs for the murine lyn protein tyrosine kinase gene were cloned from mouse bone marrow-derived monocytic cells. Comparison of the human and murine genes demonstrated a 94% homology in peptide sequence. Comparable to the human gene, murine lyn was found to be expressed in myeloid and B-lymphoid lineage cells. During the cloning, two types of cDNAs were obtained that differed by the presence (lynA) or absence (lynB) of 63 bp within the amino-terminal coding region of the gene. The genomic structure of the murine lyn gene demonstrates that the two types of lyn transcripts are derived from alternative splicing utilizing an internal splice donor site. Transcripts for both forms were found to be expressed in myeloid cells. lyn-specific antisera detected comparable levels of proteins of 56 and 53 kDa in hematopoietic cells. these 56- and 53-kDa proteins comigrated with proteins produced by in vitro translation or in vivo expression of the lynA and lynB cDNAs, respectively. The two forms had comparable in vitro kinase activities in immunoprecipitates and showed similar peptide patterns, with partial V8 digestion of the in vitro-phosphorylated proteins. The potential significance of the two lyn proteins is discussed.
18
Jelinek, Jaroslav, Jean-Pierre J. Issa, Rong He, Radek Cmejla, Jana Cmejlova, and Dagmar Pospisilova. "RPS19 and JAK2 Are Not Silenced by DNA Methylation in Diamond-Blackfan Anemia." Blood 108, no. 11 (November 16, 2006): 1312. http://dx.doi.org/10.1182/blood.v108.11.1312.1312.
Full textAbstract:
Abstract Diamond Blackfan Anemia (DBA) is a congenital disorder characterized by decreased red blood cell production accompanied by developmental abnormalities in 30% patients. Twenty-five percent of DBA patients display heterozygous mutations of ribosomal protein S19 (RPS19) on chromosome 19q13.2. No mutations were found in genes for other ribosomal proteins of the translation initiation complex. Although a second DBA locus has been proposed on in the region 8p23.3–8p22, the precise molecular defect is not known in 75% of DBA patients. The exact mechanism of how RPS19 mutations affect erythropoiesis remains unclear. Haploinsufficency of RPS19 may hamper translation machinery important for rapid erythroid differentiation. Reduced gene expression of a cluster of ribosomal proteins including RPS19 in DBA patients was recently reported. No causal therapy for DBA is available, with the exception of bone marrow transplantation. Some DBA patients benefit therapeutically from corticosteroids, cyclosporine A, or metoclopramide. Recently, a long-lasting remission was described in a DBA patient treated with valproic acid, a histone deacetylase inhibitor, suggesting epigenetic suppression of genes critical for erythropoiesis may be involved in the pathogenesis of DBA. DNA methylation of promoter-associated CpG islands is an epigenetic modification resulting in transcriptional silencing functionally equivalent to a loss-of-function mutation. Constitutive activation of JAK2 tyrosine kinase by a somatic V617F mutation leads to excessive erythropoiesis in polycythemia vera, an antonym of DBA. We hypothesized that silencing by DNA methylation of promoter-associated CpG island of the RPS19 or JAK2 genes may play a role in DBA. To test the hypothesis, we analyzed DNA methylation of RPS19 and JAK2 genes in 14 patients from the Czech DBA Registry. Genomic DNA isolated from blood cells of 3 DBA patients carrying heterozygous RPS19 mutations, 11 DBA patients without RPS19 mutation and 4 control samples was treated with bisulfite to convert all unmethylated cytosines to uracils while methylated cytosines were spared from the conversion. A region spanning 13 CpG sites positioned from 1–160 bases downstream from transcription start site (TSS) of RPS19 gene was PCR amplified and cloned in a sequencing vector. Individual bacterial clones were isolated and PCR inserts were sequenced in 8–12 clones per sample. Bisulfite cloning and sequencing revealed that more than 99% of CpG sites were converted to TpG and thus not methylated either in DBA samples (only 4/1466 CpG sites were methylated, methylation frequency was 0.3%) or control samples (2/555 CpG sites methylated, methylation frequency 0.4%). To explore a possibility of epigenetic suppression of erythropoietin signaling in DBA we analyzed DNA methylation of the CpG island of JAK2 tyrosine kinase gene in the same set of samples. Bisulfite-treated DNA was PCR amplified and T/C polymorphisms corresponding to unmethylated or methylated CpG sites were quantified by pyrosequencing. All DBA and control samples showed the absence of DNA methylation at four CpG sites located 12 to 25 bases downstream of TSS. We conclude that epigenetic silencing by DNA methylation is not involved in the expression of ribosomal structural protein RPS19; neither it affects the expression of a transducer of erythropoietin signaling JAK2 tyrosine kinase.
19
Lord, K. A., A. Abdollahi, S. M. Thomas, M. DeMarco, J. S. Brugge, B. Hoffman-Liebermann, and D. A. Liebermann. "Leukemia inhibitory factor and interleukin-6 trigger the same immediate early response, including tyrosine phosphorylation, upon induction of myeloid leukemia differentiation." Molecular and Cellular Biology 11, no. 9 (September 1991): 4371–79. http://dx.doi.org/10.1128/mcb.11.9.4371.
Full textAbstract:
Leukemia inhibitory factor (LIF) and interleukin-6 (IL-6), two multifunctional cytokines lacking structural homology and binding to distinct receptors, share interesting functional similarities, which include induction of hematopoietic differentiation in normal and myeloid leukemia cells, induction of neuronal cell differentiation, and stimulation of acute-phase protein synthesis in hepatocytes. Structural information on the LIF receptor is not yet available, whereas recent cloning of the IL-6 receptor has shown it to be bipartite, with a signal-transducing subunit that lacks sequence homology to known protein kinases and produces second messengers of unknown nature. The molecular nature of the mechanisms which LIF and IL-6 use to induce cell differentiation is not known. To address this issue, we took advantage of a clone of M1 myeloblastic leukemia cells capable of being induced for terminal differentiation by both LIF and IL-6 directly activate the same set of immediate early response genes upon induction of M1 myeloid differentiation. At least two mechanisms of gene activation, one transcriptional and the other posttranscriptional, are shown to be involved. It is also shown that the LIF and IL-6 immediate early response, at suboptimal cytokine concentrations, is additive. Using a variety of protein kinase activators and inhibitors, we have shown that the intracellular signalling pathways for both LIF and IL-6 are distinct from those of known second messengers and involve protein phosphorylation, notably tyrosine phosphorylation of a 160-kDa protein, as an essential step(s) in the immediate early activation of MyD gene expression. These observations indicate that the functional similarities of LIF and IL-6 as inducers of cell differentiation prevail at the level of the complex differentiation immediate early response and implicate common mechanisms of signal transduction for LIF- and IL-6-induced differentiation.
20
Lord, K. A., A. Abdollahi, S. M. Thomas, M. DeMarco, J. S. Brugge, B. Hoffman-Liebermann, and D. A. Liebermann. "Leukemia inhibitory factor and interleukin-6 trigger the same immediate early response, including tyrosine phosphorylation, upon induction of myeloid leukemia differentiation." Molecular and Cellular Biology 11, no. 9 (September 1991): 4371–79. http://dx.doi.org/10.1128/mcb.11.9.4371-4379.1991.
Full textAbstract:
Leukemia inhibitory factor (LIF) and interleukin-6 (IL-6), two multifunctional cytokines lacking structural homology and binding to distinct receptors, share interesting functional similarities, which include induction of hematopoietic differentiation in normal and myeloid leukemia cells, induction of neuronal cell differentiation, and stimulation of acute-phase protein synthesis in hepatocytes. Structural information on the LIF receptor is not yet available, whereas recent cloning of the IL-6 receptor has shown it to be bipartite, with a signal-transducing subunit that lacks sequence homology to known protein kinases and produces second messengers of unknown nature. The molecular nature of the mechanisms which LIF and IL-6 use to induce cell differentiation is not known. To address this issue, we took advantage of a clone of M1 myeloblastic leukemia cells capable of being induced for terminal differentiation by both LIF and IL-6 directly activate the same set of immediate early response genes upon induction of M1 myeloid differentiation. At least two mechanisms of gene activation, one transcriptional and the other posttranscriptional, are shown to be involved. It is also shown that the LIF and IL-6 immediate early response, at suboptimal cytokine concentrations, is additive. Using a variety of protein kinase activators and inhibitors, we have shown that the intracellular signalling pathways for both LIF and IL-6 are distinct from those of known second messengers and involve protein phosphorylation, notably tyrosine phosphorylation of a 160-kDa protein, as an essential step(s) in the immediate early activation of MyD gene expression. These observations indicate that the functional similarities of LIF and IL-6 as inducers of cell differentiation prevail at the level of the complex differentiation immediate early response and implicate common mechanisms of signal transduction for LIF- and IL-6-induced differentiation.
21
Ratajczak, MZ, WI Kuczynski, DL Sokol, JS Moore, CH Jr Pletcher, and AM Gewirtz. "Expression and physiologic significance of Kit ligand and stem cell tyrosine kinase-1 receptor ligand in normal human CD34+, c-Kit+ marrow cells." Blood 86, no. 6 (September 15, 1995): 2161–67. http://dx.doi.org/10.1182/blood.v86.6.2161.bloodjournal8662161.
Full textAbstract:
To determine the potential role of autocrine growth factor production in regulating primitive human hematopoietic cell development, we examined highly purified CD34+, c-Kit+ marrow mononuclear cells for expression of c-Kit ligand (KL) and stem cell tyrosine kinase 1 (stk1) ligand (STK1-L). Normal marrow mononuclear cells coexpressing CD34 and c-Kit were isolated by a combination of immunomagnetic bead isolation and fluorescence-activated cell sorting. Purified cells were then screened for expression of KL and stk1-L mRNA using a sensitive reverse transcription-polymerase chain reaction method. Using this approach, expression of both cytokine genes at the mRNA level was found in this highly enriched cell population. We then examined the functional significance of these mRNAs by inhibiting their expression with antisense (AS) oligodeoxynucleotides (ODN). In comparison to untreated or control ODN treated cells, inhibition of KL led to a 70% and 89% inhibition in burst-forming unit-erythroid (BFU-E) and colony-forming unit-Mix (CFU-Mix) colonies but had no significant effect on CFU- granulocyte-macrophage (CFU-GM) cloning efficiency. In contrast, inhibition of STK1-L alone had no effect on colony formation. However, when STK1-L AS ODN was combined with KL AS ODN, additive inhibition of CFU-GM and CFU-MIX but not of BFU-E colonies was observed. These findings, along with those of our previous studies showing inhibition of primitive hematopoietic cell growth with antisense ODN directed towards the stk1 receptor, suggest the possibility that both receptor/ligand axes regulate primitive hematopoietic cell growth via an autocrine growth loop.
22
Nakamura, Fumihiko, Ko Sasaki, Kazuhiro Maki, Yuichi Nakamura, Yuko Sato, and Kinuko Mitani. "Cloning and Characterization of a Novel Leukemogenetic Gene TEL-PTPRR." Blood 104, no. 11 (November 16, 2004): 2892. http://dx.doi.org/10.1182/blood.v104.11.2892.2892.
Full textAbstract:
Abstract The TEL gene mapped on 12p13 is rearranged by chromosomal translocations in a variety of human leukemias and is fused to various partner genes mainly encoding receptor type and non-receptor type tyrosine kinases and transcription factors. Using 3′ RACE method, we have cloned novel chimeric cDNAs TEL/protein tyrosine phosphatase receptor type R (PTPRR) that is generated by inv(12)(p13q13) found in an acute myeloid leukemia case. TEL is a member of ETS family transcription factors and contains the homodimerizing or heterodimerizing helix-loop-helix domain at the N-terminus and the DNA-binding ETS domain at the C-terminus. TEL works as a tumor suppressor. On the other hand, PTPRR is a brain-specific receptor type protein tyrosine phosphatase with a single intracellular catalytic domain. PTPRR is the first tyrosine phosphatase that was identified as a fusion partner for TEL. The chromosomal rearrangement joined exon 4 in the TEL gene and exon 7 in the PTPRR gene and produced ten isoforms of TEL/PTPRR cDNAs through alternative splicing. Among them, only one expresses TEL/PTPRR chimeric protein that consists of the HLH domain from TEL and the catalytic domain from PTPRR and the others express truncated forms of TEL. Because reciprocal PTPRR-TEL mRNA as well as wild-type-PTPRR mRNA was not detected by RT-PCR analysis, TEL-PTPRR is likely to be involved in leukemogenesis. To clarify molecular mechanisms by inv(12), we first molecularly analyzed the TEL/PTPRR chimeric cDNA and the N-terminally truncated TEL cDNA containing exons 1–4 (tTEL). When transiently expressed in NIH3T3 cells, both TEL/PTPRR and tTEL were equally contributed to the cytoplasm and the nucleus. Moreover, both isoforms were found to associate with wild-type-TEL by immunoprecipitation assays. Therefore, we speculated that TEL/PTPRR and tTEL affect wild-type-TEL’s functions in the nucleus through heterodimerizing with it. As expected from their structures, both isoforms did not bind to the ETS binding consensus sequence (EBS). In reporter assays with (ETS)3tkLuc, they did not repress transcription through EBS, while wild-type-TEL did. Interestingly, both TEL/PTPRR and tTEL dominantly interfered with the wild-type-TEL-induced transcriptional repression in a dose-dependent manner. On in vitro phosphatase assays using p-NPP as a substrate, both isoforms were shown to completely lack phosphatase activities. To assess the biological properties of TEL/PTPRR, we employed human megakaryoblastic leukemia cell line UT7/GM that proliferate completely depending on granulocyte macrophage-colony stimulating factor (GM-CSF). Overexpression of TEL/PTPRR resulted in factor-independent proliferation in UT7/GM cells. Interestingly, phosphorylated STAT3 remained after GM-CSF withdrawal in the TEL/PTPRR-overexpressing cells, while it rapidly declined in the mock cells. All these data collectively suggest that appearance of inv(12) causes leukemic transformation in human myeloid cells, at least partly through inactivating tumor-suppressive functions of wild-type-TEL. Considering that STAT3-mediated signals are maintained even after the cytokine depletion when TEL/PTPRR is overexpressed, TEL/PTPRR may regulate JAK/STAT signals through unknown mechanisms.
23
Scalia, Pierluigi, Stephen J. Williams, and Antonio Giordano. "Core Element Cloning, Cis-Element Mapping and Serum Regulation of the Human EphB4 Promoter: A Novel TATA-Less Inr/MTE/DPE-Like Regulated Gene." Genes 10, no. 12 (December 2, 2019): 997. http://dx.doi.org/10.3390/genes10120997.
Full textAbstract:
The EphB4 gene encodes for a transmembrane tyrosine kinase receptor involved in embryonic blood vessel differentiation and cancer development. Although EphB4 is known to be regulated at the post-translational level, little is known about its gene regulation. The present study describes the core promoter elements’ identification and cloning, the cis-regulatory elements’ mapping and the serum regulation of the human EphB4 gene promoter region. Using bioinformatic analysis, Sanger sequencing and recombinant DNA technology, we analyzed the EphB4 gene upstream region spanning +40/−1509 from the actual transcription start site (TSS) and proved it to be a TATA-less gene promoter with dispersed regulatory elements characterized by a novel motif-of-ten element (MTE) at positions +18/+28, and a DPE-like motif and a DPE-like-repeated motif (DRM) spanning nt +27/+30 and +32 +35, respectively. We also mapped both proximal (multiple Sp1) and distal (HoxA9) trans-activating/dispersed cis-acting transcription factor (TF)-binding elements on the region we studied and used a transient transfection reporter assay to characterize its regulation by serum and IGF-II using EphB4 promoter deletion constructs with or without the identified new DNA-binding elements. Altogether, these findings shed new light on the human EphB4 promoter structure and regulation, suggesting mechanistic features conserved among Pol-II TATA-less genes phylogenetically shared from Drosophila to Human genomes.
24
Yamamoto, K., F. W. Quelle, W. E. Thierfelder, B. L. Kreider, D. J. Gilbert, N. A. Jenkins, N. G. Copeland, O. Silvennoinen, and J. N. Ihle. "Stat4, a novel gamma interferon activation site-binding protein expressed in early myeloid differentiation." Molecular and Cellular Biology 14, no. 7 (July 1994): 4342–49. http://dx.doi.org/10.1128/mcb.14.7.4342.
Full textAbstract:
Interferon regulation of gene expression is dependent on the tyrosine phosphorylation and activation of the DNA-binding activity of two related proteins of 91 kDa (STAT1) and/or 113 kDa (STAT2). Recent studies have suggested that these proteins are substrates of Janus kinases and that proteins related in STAT1 are involved in a number of signalling pathways, including those activated in myeloid cells by erythropoietin and interleukin-3 (IL-3). To clone STAT-related proteins from myeloid cells, degenerate oligonucleotides were used in PCRs to identify novel family members expressed in myeloid cells. This approach allowed the identification and cloning of the Stat4 gene, which is 52% identical to STAT1. Unlike STAT1, Stat4 expression is restricted but includes myeloid cells and spermatogonia. In the erythroid lineage, Stat4 expression is differentially regulated during differentiation. Functionally, Stat4 has the properties of other STAT family genes. In particular, cotransfection of expression constructs for Stat4 and Jak1 and Jak2 results in the tyrosine phosphorylation of Stat4 and the acquisition of the ability to bind to the gamma interferon (IFN-gamma)-activated sequence of the interferon regulatory factor 1 (IRF-1) gene. Stat4 is located on mouse chromosome 1 and is tightly linked to the Stat1 gene, suggesting that the genes arose by gene duplication. Unlike Stat1, neither IFN-alpha nor IFN-gamma activates Stat4. Nor is Stat4 activated in myeloid cells by a number of cytokines, including erythropoietin, IL-3, granulocyte colony-stimulating factor, stem cell factor, colon-stimulating factor 1, hepatocyte growth factor, IL-2, IL-4, and IL-6.
25
Yamamoto, K., F. W. Quelle, W. E. Thierfelder, B. L. Kreider, D. J. Gilbert, N. A. Jenkins, N. G. Copeland, O. Silvennoinen, and J. N. Ihle. "Stat4, a novel gamma interferon activation site-binding protein expressed in early myeloid differentiation." Molecular and Cellular Biology 14, no. 7 (July 1994): 4342–49. http://dx.doi.org/10.1128/mcb.14.7.4342-4349.1994.
Full textAbstract:
Interferon regulation of gene expression is dependent on the tyrosine phosphorylation and activation of the DNA-binding activity of two related proteins of 91 kDa (STAT1) and/or 113 kDa (STAT2). Recent studies have suggested that these proteins are substrates of Janus kinases and that proteins related in STAT1 are involved in a number of signalling pathways, including those activated in myeloid cells by erythropoietin and interleukin-3 (IL-3). To clone STAT-related proteins from myeloid cells, degenerate oligonucleotides were used in PCRs to identify novel family members expressed in myeloid cells. This approach allowed the identification and cloning of the Stat4 gene, which is 52% identical to STAT1. Unlike STAT1, Stat4 expression is restricted but includes myeloid cells and spermatogonia. In the erythroid lineage, Stat4 expression is differentially regulated during differentiation. Functionally, Stat4 has the properties of other STAT family genes. In particular, cotransfection of expression constructs for Stat4 and Jak1 and Jak2 results in the tyrosine phosphorylation of Stat4 and the acquisition of the ability to bind to the gamma interferon (IFN-gamma)-activated sequence of the interferon regulatory factor 1 (IRF-1) gene. Stat4 is located on mouse chromosome 1 and is tightly linked to the Stat1 gene, suggesting that the genes arose by gene duplication. Unlike Stat1, neither IFN-alpha nor IFN-gamma activates Stat4. Nor is Stat4 activated in myeloid cells by a number of cytokines, including erythropoietin, IL-3, granulocyte colony-stimulating factor, stem cell factor, colon-stimulating factor 1, hepatocyte growth factor, IL-2, IL-4, and IL-6.
26
Dottori, Mirella, Michelle Down, Andreas Hüttmann, David R. Fitzpatrick, and Andrew W. Boyd. "Cloning and Characterization of EphA3 (Hek) Gene Promoter: DNA Methylation Regulates Expression in Hematopoietic Tumor Cells." Blood 94, no. 7 (October 1, 1999): 2477–86. http://dx.doi.org/10.1182/blood.v94.7.2477.419k13_2477_2486.
Full textAbstract:
The Eph family of receptor tyrosine kinases (RTK) has restricted temporal and spatial expression patterns during development, and several members are also found to be upregulated in tumors. Very little is known of the promoter elements or regulatory factors required for expression of Eph RTK genes. In this report we describe the identification and characterization of the EphA3 gene promoter region. A region of 86 bp located at −348 bp to −262 bp upstream from the transcription start site was identified as the basal promoter. This region was shown to be active in both EphA3-expressing and -nonexpressing cell lines, contrasting with the widely different levels of EphA3 expression. We noted a region rich in CpG dinucleotides downstream of the basal promoter. Using Southern blot analyses with methylation-sensitive restriction enzymes and bisulfite sequencing of genomic DNA, sites of DNA methylation were identified in hematopoietic cell lines which correlated with their levels of EphA3 gene expression. We showed that EphA3 was not methylated in normal tissues but that a subset of clinical samples from leukemia patients showed extensive methylation, similar to that observed in cell lines. These results suggest that DNA methylation may be an important mechanism regulating EphA3 transcription in hematopoietic tumors.
27
Brändli, A. W., and M. W. Kirschner. "Molecular cloning of tyrosine kinases in the early Xenopus embryo: Identification of eck-related genes expressed in cranial neural crest cells of the second (Hyoid) Arch." Developmental Dynamics 203, no. 2 (June 1995): 119–40. http://dx.doi.org/10.1002/aja.1002030202.
Full text
28
Halatsch, Marc-Eric, Esther E. Gehrke, Vassilios I. Vougioukas, Ingolf C. Bötefür, Farhad A. Borhani, Thomas Efferth, Erich Gebhart, Sebastian Domhof, Ursula Schmidt, and Michael Buchfelder. "Inverse correlation of epidermal growth factor receptor messenger RNA induction and suppression of anchorage-independent growth by OSI-774, an epidermal growth factor receptor tyrosine kinase inhibitor, in glioblastoma multiforme cell lines." Neurosurgical Focus 16, no. 2 (February 2004): 1–11. http://dx.doi.org/10.3171/foc.2004.16.2.12.
Full textAbstract:
Object Quantitative and qualitative alterations in the epidermal growth factor receptor (EGFR) commonly occur in many cancers in humans, including malignant gliomas. The aim of the current study was to evaluate molecular and cellular effects of OSI-774, a novel EGFR tyrosine kinase inhibitor, on nine glioblastoma multiforme (GBM) cell lines. Methods The effects of OSI-774 on expression of EGFR messenger (m)RNA and protein, proliferation, anchorage-independent growth, and apoptosis were examined using semiquantitative reverse transcription–polymerase chain reaction, immunocytochemical analysis, Coulter counting, soft agar cloning, and terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate nick-end labeling/fluorescence-activated cell sorting, respectively. All p53 genes were completely and bidirectionally sequenced. Suppression of anchorage-independent growth by OSI-774 was inversely correlated to the induction of EGFR mRNA during relative serum starvation (r = −0.74) and was unrelated to p53 status. Overall, suppression of anchorage-independent growth was a considerably stronger effect of OSI-774 than inhibition of proliferation. The extent of OSI-774–induced apoptosis positively correlated with both proliferation and anchorage-independent growth of GBM cell lines (r = 0.75 and 0.79, respectively). In a single cell line derived from a secondary GBM, exposure to concentrations of greater than or equal to 1 μmol/L resulted in a substantial net cell loss during proliferation studies. Conclusions The induction of EGFR mRNA may constitute a cellular mechanism to counteract the inhibitory effect of OSI-774 on the anchorage-independent growth of GBM cells. In contrast, no considerable correlation could be established between baseline expression levels of EGFR (both mRNA and protein) in GBM cell lines and their biological response to OSI-774. The OSI-774 induced greater (p53-independent) apoptosis in more malignant GBM phenotypes and may be a promising therapeutic agent against secondary GBM.
29
Halatsch, Marc-Eric, Esther E. Gehrke, Vassilios I. Vougioukas, Ingolf C. Bötefür, Farhad A.-Borhani, Thomas Efferth, Erich Gebhart, Sebastian Domhof, Ursula Schmidt, and Michael Buchfelder. "Inverse correlation of epidermal growth factor receptor messenger RNA induction and suppression of anchorage-independent growth by OSI-774, an epidermal growth factor receptor tyrosine kinase inhibitor, in glioblastoma multiforme cell lines." Journal of Neurosurgery 100, no. 3 (March 2004): 523–33. http://dx.doi.org/10.3171/jns.2004.100.3.0523.
Full textAbstract:
Object. Quantitative and qualitative alterations in the epidermal growth factor receptor (EGFR) commonly occur in many cancers in humans, including malignant gliomas. The aim of the current study was to evaluate molecular and cellular effects of OSI-774, a novel EGFR tyrosine kinase inhibitor, on nine glioblastoma multiforme (GBM) cell lines. Methods. The effects of OSI-774 on expression of EGFR messenger (m)RNA and protein, proliferation, anchorage-independent growth, and apoptosis were examined using semiquantitative reverse transcription—polymerase chain reaction, immunocytochemical analysis, Coulter counting, soft agar cloning, and terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate nick-end labeling/fluorescence-activated cell sorting, respectively. All p53 genes were completely and bidirectionally sequenced. Suppression of anchorage-independent growth by OSI-774 was inversely correlated to the induction of EGFR mRNA during relative serum starvation (r = −0.74) and was unrelated to p53 status. Overall, suppression of anchorage-independent growth was a considerably stronger effect of OSI-774 than inhibition of proliferation. The extent of OSI-774—induced apoptosis positively correlated with both proliferation and anchorage-independent growth of GBM cell lines (r = 0.75 and 0.79, respectively). In a single cell line derived from a secondary GBM, exposure to concentrations of greater than or equal to 1 Émol/L resulted in a substantial net cell loss during proliferation studies. Conclusions. The induction of EGFR mRNA may constitute a cellular mechanism to counteract the inhibitory effect of OSI-774 on the anchorage-independent growth of GBM cells. In contrast, no considerable correlation could be established between baseline expression levels of EGFR (both mRNA and protein) in GBM cell lines and their biological response to OSI-774. The OSI-774 induced greater (p53-independent) apoptosis in more malignant GBM phenotypes and may be a promising therapeutic agent against secondary GBM.
30
Ku, Matthew, Nisha Narayan, Meaghan Wall, Ruth N. MacKinnon, Lynda J. Campbell, and Harshal Nandurkar. "Identification and Analysis of Oncogenic Pathways in Deletion 20q Acute Myeloid Leukaemia." Blood 120, no. 21 (November 16, 2012): 1324. http://dx.doi.org/10.1182/blood.v120.21.1324.1324.
Full textAbstract:
Abstract Abstract 1324 Deletion of the long arm of chromosome 20 [del(20q)] is a common recurrent chromosomal abnormality in acute myeloid leukaemia (AML). It is a key step in AML development and a better understanding of the associated molecular events is important. The abnormal chromosome 20 in del(20q) AML has been shown to have lost a “Common Deleted Region” (CDR) that contains Protein Tyrosine Phosphatase Receptor T (PTPRT), a tyrosine phosphatase that is mutated in many human cancers such as AML. We have previously reported (MacKinnon et al, Genes, Chromosomes and Cancer 2010) that del(20q) also harbours an amplified “Common Retained Region,” (CRR) which contains Haemopoietic Cell Kinase (HCK). HCK is anoncogenic Src tyrosine kinase and its aberrant activation has been shown to contribute to the pathogenesis of some haematological malignancies. We hypothesize that the amplification of HCK in the CRR cooperates with the loss of PTPRT in the CDR to cause AML. Our model proposes that AML occurs either through direct interaction between HCK and PTPRT, or through aberrant activation of Signal Transducer and Activator of Transcription 3 (STAT3), a cytoplasmic second messenger that is important in cellular signalling. Constitutively activated STAT3 has been shown to be oncogenic in several malignancies, including AML. STAT3 is a direct target of both HCK and PTPRT. It is phosphorylated (hence activated) by HCK, and dephosphorylated (hence inactivated) by PTPRT. This provides a downstream leukaemogenic pathway for our model. The ultimate aim of our experiments is to prove this hypothesis using mouse models. Murine haemopoietic stem cells (HSC) were isolated from the bone marrows of wild type C57BL/6 (WT) and PTPRT-null mice by Fluorescence Activated Cell Sorting for Lineage negative, C-kit and Sca-1 positive (LKS+) cells. Retroviral constructs of HCK were generated by cloning it into the retroviral vector pMSCViresEGFP(MIG), with GFP as reporter. Murine HSC were transduced with either retroviral HCK or MIG vector control and Phoenix cell system was used for retroviral packaging. Experiments using isolated LKS+ HSC were performed to examine for features of AML. Examination of bone marrow cells from del(20q) AML patients by quantitative PCR revealed an increase in HCK mRNA expression and a reduction in PTPRT expression. Wild type (WT) and PTPRT-null murine HSC transduced with either MIG or HCK were cultured in methylcellulose media. Colony forming units (CFU) were enumerated on day7 and day12. We found that both WT and PTPRT-null HSC transduced with HCK showed a significant increase in colony numbers compared to MIG transduced HSC. Furthermore, the fold increment in colony number was higher in the PTPRT-null genotype as shown in figure 1. Moreover, an intracellular anti-phosphoSTAT3 assay was performed to assess STAT3 phosphorylation levels in the transduced HSC. It demonstrated that in both WT and PTPRT-null HSC that have been transduced with HCK, STAT3 hyperphosphorylation, and hence overactivation, occured. This response was again more exaggerated in the PTPRT-null HSC, as seen in figure 2. We are currently transplanting transduced LKS+ HSC (either MIG or HCK) into lethally irradiated murine recipients to assess AML formation in a reconstitution study. The recipient mice will be assessed for evidence of engraftment and subsequent AML. The preliminary data reveals a likely new oncogenic-signalling cascade: that HCK amplification and loss of PTPRT in del(20q) AML may cooperate to cause AML directly, or by aberrant activity of hyperphosphorylated STAT3. Disclosures: No relevant conflicts of interest to declare.
31
Kwiatkowski, Boguslaw A., and Robert E. Richard. "hDLG Associated Protein 1 (DLGAP1) Supports Mpl-Driven Cell Proliferation and Differentiation Through Its Centrosomal Function,." Blood 118, no. 21 (November 18, 2011): 3402. http://dx.doi.org/10.1182/blood.v118.21.3402.3402.
Full textAbstract:
Abstract Abstract 3402 The Mpl proto-oncogene functions as a basic factor in megakaryocytic development and platelet production as well as contributing to hematopoietic stem cell (HSC) homeostasis and self-renewal. Several mutations in Mpl have been shown to be associated with the myeloproliferative neoplasms. We used retroviral insertional mutagenesis to screen for factors that might give a proliferative and/or survival advantage for cells dependent on Mpl signaling. We developed a retroviral construct expressing a dimerizable form of Mpl that can interrupt genes through insertion, and can activate adjacent genes with its long terminal repeats. We transduced the human leukemia cell line K562 with this construct and blocked the endogenous transforming BCR-ABL kinase using Imatinib. Cells dependent on Mpl signaling were selected by the addition of the dimerizing drug, AP20187. In the absence of Mpl signaling the cells underwent erythroid differentiation and died. Cells that acquired a proliferative advantage and were dependent on Mpl function were expanded. Cloning of retroviral integration sites (RIS) from the selected populations allowed the identification of recurrent RIS that co-localized in the genome. We have identified 668 RIS from 36 independent transductions with a presumed ∼ 1.9 × 106 independent insertions in the initial non-selected, cell population. Among these, 203 RIS represented independent insertion events. Three independent RIS were located in the fourth intron of the Discs large homolog-associated protein 1 (DLGAP1). This protein is a member of the Discs-large/Scribble/Lethal Giant Larvae pathway. DLGAP1, in cooperation with DLG1 and CDC42, has recently been shown to control centrosome positioning and cell polarity in astrocytes (Manneville J-B et al. 2010, JBC, v.191, no 3, pp. 585–598). We investigated the role of DLGAP1 in MPL signaling. Overexpression of full length DLGAP1 had minimal effect on plain K562 cells, but it significantly slowed down the proliferation of K562 cells switched to Mpl signaling. The identified insertion sites would predict the increased expression of an amino-end truncated isoform. When this truncated form was overexpressed in K562 cells and Mpl dependent K562 cells, the cells proliferated at an increased rate. Immunofluorescent studies revealed that the full length DLGAP1 colocalized with major centrosomal markers including gamma-tubulin, PCM1, and APC in K562, HEL, UT7/TPO and Mo7e cell lines. We did not observe colocalization with centrosomes when the truncated isoform of DLGAP1 was overexpressed. Immunoflourescent microscopy revealed that endogenous DLGAP1 in myeloid cell lines was localized in the centriolar satellites, a cellular structure essential for centrosome integrity and function. The immunoflourescent pattern using an anti DLGAP1 antibody showed cell cycle dependent assembly and disassembly. Protein sequence analyses of DLGAP1 indicated consensus sites for presumptive Jak2 phosphorylation as well as consensus sites for SRC tyrosine kinases. To test the possible influence of the two types of kinases on DLGAP1 we treated K562, HEL and UT7/TPO cell lines with an inhibitor of Jak2 (AG490) and independently with an inhibitor of SRC (SU6656). In each case the centrosomal staining of DLGAP1 was diminished with minimal effect on the cytoplasmic fraction of DLGAP1. Interestingly, treatment with SU6656 of the K562 cells switched to Mpl signaling and overexpressing full length GFP-DLGAP1 fusion resulted in a significant increase in the number of polyploid cells. The changes correlated with upregulation of the megakaryocytic marker CD41a and downregulation of the erythroid marker Glycophorin A as assessed by FACS analyses. As centrosomal aberrations are frequent markers of the myeloproliferative neoplasms, we intend to study the role of DLGAP1 in these bone marrow disorders. Disclosures: No relevant conflicts of interest to declare.
32
Brennan, Cameron, and Tatsuya Ozawa. "Novel Tyrosine Kinase Genes Fusions in Glioma." Neurosurgery 62, no. 6 (June 2008): 1425. http://dx.doi.org/10.1227/01.neu.0000333530.85150.19.
Full text
33
Cooper, Jonathan A. "Transforming mutations in protein?tyrosine kinase genes." BioEssays 4, no. 1 (January 1986): 9–15. http://dx.doi.org/10.1002/bies.950040104.
Full text
34
Ku, Matthew, Nisha Narayan, Ruth N. MacKinnon, Meaghan Wall, Carl Walkley, Louise E. Purton, Heung-Chin Cheng, Lynda J. Campbell, and Harshal Nandurkar. "Identification and Analysis of Oncogenic Pathways in Deletion 20q Acute Myeloid Leukaemia." Blood 124, no. 21 (December 6, 2014): 5195. http://dx.doi.org/10.1182/blood.v124.21.5195.5195.
Full textAbstract:
Abstract Deletion of the long arm of chromosome 20 [del(20q)] is a common recurrent chromosomal abnormality in acute myeloid leukaemia (AML) and myeloproliferative neoplasms (MPN). The abnormal chromosome 20 has a “Common Deleted Region” (CDR) that contains Protein Tyrosine Phosphatase Receptor T (PTPRT), a tyrosine phosphatase that is mutated in many human cancers. We have previously reported (MacKinnon et al, Genes, Chromosomes and Cancer 2010) that del(20q) also harbours an amplified “Common Retained Region,” (CRR) which contains multiple copies of Haemopoietic Cell Kinase (HCK). HCK is an oncogenic Src tyrosine kinase and its aberrant activation has been shown to contribute to the pathogenesis of some haematological malignancies. Our hypothesis is that the amplification of HCK in the CRR cooperated with the loss of PTPRT in the CDR. Indeed, our results strongly suggested that HCK amplification with PTPRT loss led to a myeloproliferative phenotype seen in MPN. Our model proposed that MPN occurred either through the consequence of direct interaction between HCK and PTPRT, or through aberrant activation of Signal Transducer and Activator of Transcription 3 (STAT3). Constitutively activated STAT3 has been shown to be oncogenic in several haematological malignancies. STAT3 is a direct target of both HCK and PTPRT. It is phosphorylated (hence activated) by HCK, and dephosphorylated (hence inactivated) by PTPRT. This provides a downstream leukaemogenic pathway for our model.The ultimate aim of our experiments is to prove this hypothesis using mouse models. Murine haemopoietic stem cells (HSC) were isolated from the bone marrows of wild type C57BL/6 (WT) and PTPRT-null mice by Fluorescence Activated Cell Sorting for Lineage negative, C-kit and Sca-1 positive (LKS+) cells. Retroviral constructs of HCK were generated by cloning it into the retroviral vector pMSCViresEGFP(MIG), with GFP as reporter. Murine HSC were transduced with either retroviral HCK or MIG vector control. Hence wild type (WT) and PTPRT-null murine HSC transduced with either MIG or HCK were used in our experiments (WTMIG, WTHCK, PTPRTMIG, PTPRTHCK). We have previously presented that PTPRTHCK showed a significant increase in methylcelluose colony numbers and colony sizes compared to PTPRTMIG, while there was no difference between WTMIG and WTHCK. In addition, an intracellular anti-phosphoSTAT3 antibody assay demonstrated that in both WTHCK and PTPRTHCK, significant STAT3 hyperphosphorylation, and hence overactivation, occurred. This response was more exaggerated in the PTPRTHCK. Finally, direct interaction between HCK and PTPRT was shown with an interaction assay using recombinant proteins. We now present the in vivo data in the murine recipients. In Vivo Reconstitution LKS+ HSC transduced with either MIG or HCK were transplanted into lethally irradiated SJL/PTPRCa murine recipients to assess AML or MPN formation in a reconstitution study over 12 months. The recipients from the four cohorts (WTMIG, WTHCK, PTPRTMIG and PTPRTHCK) were regularly analysed by full blood counts, peripheral blood GFP%, and blood films. At 12 months, they were culled and the organs harvested and analysed by flow cytometry, cytospin, and paraffin-embedded histology (H + E and immunohistochemistry). Recipents of PTPRTHCK HSC had a significantly higher proportion of nucleated erythroid populations in the bone marrow by flow and larger splenic size by weight. They also showed a higher haemoglobin (Hb) at 12 months, although this did not reach statistical significance. Fig 1. Haemoglobin level of the recipient mice. PTPRTHCK recipients had a higher average Hb compared to the others, although only reaching statistical significance when compared to PTPRTMIG recipients. Fig 1. Haemoglobin level of the recipient mice. PTPRTHCK recipients had a higher average Hb compared to the others, although only reaching statistical significance when compared to PTPRTMIG recipients. Fig 2. %Nucleated erythroid population in bone marrow by flow. PTPRTHCK recipients had a higher % of nucleated erythroid cells compared to the others Fig 2. %Nucleated erythroid population in bone marrow by flow. PTPRTHCK recipients had a higher % of nucleated erythroid cells compared to the others Fig 3. Splenic size of recipient mice at 12 months. PTPRTHCK recipients had a larger splenic size compared to others, although this was just outside statistical significance when compared to WTHCK. Fig 3. Splenic size of recipient mice at 12 months. PTPRTHCK recipients had a larger splenic size compared to others, although this was just outside statistical significance when compared to WTHCK. Conclusion The data reveal a likely new oncogenic-signalling cascade: that HCK amplification and loss of PTPRT in del(20q) may cooperate to cause MPN directly, or by aberrant activity of hyperphosphorylated STAT3. Disclosures No relevant conflicts of interest to declare.
35
Kohmura, N., T. Yagi, Y. Tomooka, M. Oyanagi, R. Kominami, N. Takeda, J. Chiba, Y. Ikawa, and S. Aizawa. "A novel nonreceptor tyrosine kinase, Srm: cloning and targeted disruption." Molecular and Cellular Biology 14, no. 10 (October 1994): 6915–25. http://dx.doi.org/10.1128/mcb.14.10.6915.
Full textAbstract:
We have isolated a novel nonreceptor tyrosine kinase, Srm, that maps to the distal end of chromosome 2. It has SH2, SH2', and SH3 domains and a tyrosine residue for autophosphorylation in the kinase domain but lacks an N-terminal glycine for myristylation and a C-terminal tyrosine which, when phosphorylated, suppresses kinase activity. These are structural features of the recently identified Tec family of nonreceptor tyrosine kinases. The Srm N-terminal unique domain, however, lacks the structural characteristics of the Tec family kinases, and the sequence similarity is highest to Src in the SH region. The expression of two transcripts is rather ubiquitous and changes according to tissue and developmental stage. Mutant mice were generated by gene targeting in embryonic stem cells but displayed no apparent phenotype as in mutant mice expressing Src family kinases. These results suggest that Srm constitutes a new family of nonreceptor tyrosine kinases that may be redundant in function.
36
Kohmura, N., T. Yagi, Y. Tomooka, M. Oyanagi, R. Kominami, N. Takeda, J. Chiba, Y. Ikawa, and S. Aizawa. "A novel nonreceptor tyrosine kinase, Srm: cloning and targeted disruption." Molecular and Cellular Biology 14, no. 10 (October 1994): 6915–25. http://dx.doi.org/10.1128/mcb.14.10.6915-6925.1994.
Full textAbstract:
We have isolated a novel nonreceptor tyrosine kinase, Srm, that maps to the distal end of chromosome 2. It has SH2, SH2', and SH3 domains and a tyrosine residue for autophosphorylation in the kinase domain but lacks an N-terminal glycine for myristylation and a C-terminal tyrosine which, when phosphorylated, suppresses kinase activity. These are structural features of the recently identified Tec family of nonreceptor tyrosine kinases. The Srm N-terminal unique domain, however, lacks the structural characteristics of the Tec family kinases, and the sequence similarity is highest to Src in the SH region. The expression of two transcripts is rather ubiquitous and changes according to tissue and developmental stage. Mutant mice were generated by gene targeting in embryonic stem cells but displayed no apparent phenotype as in mutant mice expressing Src family kinases. These results suggest that Srm constitutes a new family of nonreceptor tyrosine kinases that may be redundant in function.
37
Cross, NCP, and A. Reiter. "Tyrosine kinase fusion genes in chronic myeloproliferative diseases." Leukemia 16, no. 7 (July 2002): 1207–12. http://dx.doi.org/10.1038/sj.leu.2402556.
Full text
38
Grant, Seth G. N., and Thomas J. O'Dell. "Targeting tyrosine kinase genes and long-term potentiation." Seminars in Neuroscience 6, no. 1 (February 1994): 45–52. http://dx.doi.org/10.1006/smns.1994.1006.
Full text
39
Tan, J. L., and J. A. Spudich. "Developmentally regulated protein-tyrosine kinase genes in Dictyostelium discoideum." Molecular and Cellular Biology 10, no. 7 (July 1990): 3578–83. http://dx.doi.org/10.1128/mcb.10.7.3578.
Full textAbstract:
Dictyostelium discoideum, an organism that undergoes development and that is amenable to biochemical and molecular genetic approaches, is an attractive model organism with which to study the role of tyrosine phosphorylation in cell-cell communication. We report the presence of protein-tyrosine kinase genes in D. discoideum. Screening of a Dictyostelium cDNA expression library with an anti-phosphotyrosine antibody identifies fusion proteins that exhibit protein-tyrosine kinase activity. Two distinct cDNAs were identified and isolated. Though highly homologous to protein kinases in general, these kinases do not exhibit many of the hallmarks of protein-tyrosine kinases of higher eucaryotes. In addition, these genes are developmentally regulated, which suggests a role for tyrosine phosphorylation in controlling Dictyostelium development.
40
Tan, J. L., and J. A. Spudich. "Developmentally regulated protein-tyrosine kinase genes in Dictyostelium discoideum." Molecular and Cellular Biology 10, no. 7 (July 1990): 3578–83. http://dx.doi.org/10.1128/mcb.10.7.3578-3583.1990.
Full textAbstract:
Dictyostelium discoideum, an organism that undergoes development and that is amenable to biochemical and molecular genetic approaches, is an attractive model organism with which to study the role of tyrosine phosphorylation in cell-cell communication. We report the presence of protein-tyrosine kinase genes in D. discoideum. Screening of a Dictyostelium cDNA expression library with an anti-phosphotyrosine antibody identifies fusion proteins that exhibit protein-tyrosine kinase activity. Two distinct cDNAs were identified and isolated. Though highly homologous to protein kinases in general, these kinases do not exhibit many of the hallmarks of protein-tyrosine kinases of higher eucaryotes. In addition, these genes are developmentally regulated, which suggests a role for tyrosine phosphorylation in controlling Dictyostelium development.
41
Ryotokuji, Takeshi, Hiroki Yamaguchi, Ikuko Omori, Tuneaki Hirakawa, Satoshi Wakita, Tomoaki Kitano, Yoshio Mitamura, et al. "The Clinical Features and Prognostic Impact of DNMT3A Gene Mutation in Japanese Patients with De Novo AML." Blood 118, no. 21 (November 18, 2011): 2537. http://dx.doi.org/10.1182/blood.v118.21.2537.2537.
Full textAbstract:
Abstract Abstract 2537 Background and Aims: Several gene mutations were found in acute myeloid leukemia (AML) and were expected to be used as prognostic factors. The nucleophosmin 1 (NPM1) and CCAAT/enhancer-binding protein alpha (CEBPA) gene mutations are associated with a favorable outcome and lack of a transplant benefit, but FMS-like tyrosine kinase 3 (FLT3) mutations leading to an internal tandem duplication (ITD) are associated with adverse outcome. Recently the DNA methyltransferase 3A (DNMT3A) gene mutation was identified by whole-genome sequencing in patients with AML. DNMT3A encodes for the enzyme DNA (cytosine-5)-methyltransferase 3A and belongs to the family of other methyltransferases like DNMT1 and DNMT3B. These enzymes are involved in adding methyl groups to the cytosine residue of CpG dinucleotides and thus play an important role in epigenetic regulation of genes, but the mechanism of DNMT3A mutation -associated leukemogenesis was still unknown. About clinical impact of DNMT3A mutation, several groups reported that DNMT3A mutation have been associated with adverse outcome. We analyzed clinical features and prognostic impact of DNMT3A mutation in patients with Japanese patients with AML. Methods: We retrospectively analyzed 188 cases of de novo AML treated at Nippon Medical School Hospital and its affiliated facilities from 2000 to 2010. We analyzed 188 samples at initial presentation and 70 samples at relapse. Mutation analyses were performed for FLT3ITD by PCR amplification, and NPM1, CEBPA, and DNMT3A mutations by direct sequence. To validate sequencing results, PCR products were inserted into the pCR2.1-TOPO vector using a TOPO TA cloning kit (Invitrogen, Carlsbad, CA). Recombinant plasmids isolated from 8 to 12 white colonies were sequenced. Results: The median age was 53.0 years (range, 17–87 years) with 62.1% being males. Patients were followed-up for 0.03–126.1 months after initial presentation, with a median of 19.9 months. DNMT3A mutations were detected in 35 cases (18.6%) at initial presentation, and 12 cases (17.1%) at relapse. Most frequently, codon R882 located in exon 23 was mutated, and 32 cases (91.4%) of these mutations were located within methyltrasferase domain. We evaluated the correlation of clinical and genetic characteristics with DNMT3A mutations. Age (p=0.2884), sex (p=0.6964), and platelet count (p=0.9940) were not significantly different in AML patients with and without DNMT3A mutations. However the frequency of higher white blood cell count (p=0.0001), M5 in FAB classification (p=0.0018), and intermediate risk karyotype (p=0.0032) in patients with DNMT3A mutations were significantly higher than those in patients without them. Also, patients with DNMT3A mutations had a mutation in NPM1 (p<0.0001) and FLT3ITD (p=0.009) more frequently. We next assessed the prognostic impact of DNMT3A mutations. For total cohort of patients with AML, complete remission rate and rates of relapse free survival (RFS) at 5 years were not statistically different between AML patients with and without DNMT3A mutations. However the overall survival (OS) at 5 years in patients with DNMT3A mutation (11.0%) was significantly lower than those in patients without them (28.9%) (p=0.0005). Among the intermediate risk karyotype or FLT3ITD negative AML patients, RFS at 5 years were not statistically different between AML patients with and without DNMT3A mutations, but OS at 5 years in patients with DNMT3A mutation (intermediate risk karyotype: 12.6%, Flt3ITD negative: 11.6%) was significantly lower than those in patients without them (intermediate risk karyotype: 23.9%, p=0.0231, FLT3ITD negative: 30.9%, p=0.0046). Conclusions: This study shows that DNMT3A mutation is an important adverse prognostic factor among intermediate risk karyotype or FLT3ITD negative AML patients. Recently TET2 and Isocitrate Dehydrogenase (IDH) 1/2 gene mutations in de novo AML were reported. These genes including DNMT3A play an important role in epigenetic regulation of genes such as methylation etc, and mutations of these genes may be associated with leukemogenesis of AML. Now we are analyzing TET2 and IDH1/2 mutations among same our AML cohort, and we will show the prognostic impact of TET2, IDH1/2 and DNMT3A mutations in patients with de novo AML. Disclosures: No relevant conflicts of interest to declare.
42
Uckun, Fatih M., Lei Sun, Hong Ma, Sanjive Qazi, Ilker Dibirdik, and Zahide Ozer. "Targeting Human B-Precursor Acute Lymphoblastic Leukemia Cells with Recombinant Human CD19 Ligand." Blood 116, no. 21 (November 19, 2010): 599. http://dx.doi.org/10.1182/blood.v116.21.599.599.
Full textAbstract:
Abstract Abstract 599 CD19 is a 95-kDa B-lineage restricted receptor molecule that functions as a key regulator of transmembrane signals in both B-cells and B-cell precursors. Here we report the cloning and characterization of a novel high-mobility group (HMG)-box protein as the membrane-associated natural ligand of human CD19 receptor (CD19-L) on human immature as well as mature lymphoid cells. We cloned the gene encoding CD19-L from a human thymus cDNA library by yeast two-hybrid screening using the cDNA encoding human CD19 extracellular domain (AA 1 to 273) (CD19ECD) fused to the GAL4 DNA binding domain as the bait plasmid. The cDNA for the surface membrane-associated CD19-L protein is 2290-bp in length encoding a 487-aa protein with a predicted molecular mass of 54-kDa. The comparison of the amino acid sequence of CD19-L protein with reported sequences in Genebank Database revealed that CD19-L is a new member of the HMG-box protein family. CD19-L contains two leucine-rich hydrophobic nuclear export signal (NES) motifs associated with unconventional ER- and Golgi-independent transport of nuclear/cytoplasmic secretory proteins to the surface membrane. Expression of the CD19-L gene expression is limited to the lymphocyte compartment within the human lymphohematopoietic system and particularly abundant in T-lineage cells. CD19-L displays abundant expression on immature double-positive (DP) thymocytes as well as leukemic T-cell precursors from T-lineage ALL patients corresponding to immature double-negative (DN) pro-thymocyte and DP cortico-thymocyte stages of human T-cell ontogeny. CD19-L is also expressed on B-lineage lymphoid cells at all stages of human B-cell ontogeny, including fetal liver derived biphenotypic CD2+CD19+ pro-B/T cells, pro-B cells, pre-B cells and mature B-cells. Soluble recombinant human CD19-L protein produced in a baculovirus expression system exhibited exquisite specificity for the extracellular domain of CD19 and had profound effects on apoptosis-related signaling and gene expression in CD19+ human leukemia cells. Engagement of CD19 co-receptor on B-lineage ALL cells with soluble CD19-L protein perturbed CD19-associated signaling network and triggered tyrosine phosphorylation of CD19 in a time-dependent fashion with peak phosphorylation occurring within 1–5 min. CD19-phosphorylation was associated with rapid and transient activation of SYK tyrosine kinase. Treatment of B-lineage ALL cells with 100 ng/mL CD19-L for 24 h corrupted the regulation of gene expression and altered the expression levels of 13 genes directly involved in regulation of apoptosis. We next examined the ability of CD19-L to induce apoptosis in leukemic B-cell precursors from chemotherapy-resistant CD19-positive human B-lineage ALL cell lines NALM-6 (pre-B ALL), RS4;11 (MLL-AF4+ Pro-B ALL), and ALL-1 (BCR-ABL+ Pre-pre-B ALL). CD19-L (but not control proteins CD19ECD or CD19ICD) caused apoptosis in each of these 3 ALL cell lines. As CD19-L specifically targets CD19ECD, excess CD19ECD protein was able to compete with surface CD19 receptor for CD19-L binding and thereby prevent CD19-L induced apoptosis. Excess CD19 intracellular domain protein (CD19ICD) that was included as a negative control did not affect CD19-L induced apoptosis. CD19-L was also capable of causing apoptosis in chemotherapy-resistant primary leukemic cells from relapsed CD19+ B-lineage ALL patients. This collection of experimental data provides compelling evidence that CD19-L is a potent biotherapeutic new agent candidate against CD19+ lymphoid malignancies. The identification of human CD19-L may lead to therapeutic innovation for B-lineage ALL as well as other B-lineage lymphoid malignancies by providing an effective alternative to CD19-directed monoclonal antibody-based biotherapeutic agents that have encountered several limitations in clinical settings. Disclosures: No relevant conflicts of interest to declare.
43
Duhé, Roy J., Hallgeir Rui, John D. Greenwood, Kevin Garvey, and William L. Farrar. "Cloning of the gene encoding rat JAK2, a protein tyrosine kinase." Gene 158, no. 2 (June 1995): 281–85. http://dx.doi.org/10.1016/0378-1119(95)00041-4.
Full text
44
Charest, D. L., G. Mordret, K. W. Harder, F. Jirik, and S. L. Pelech. "Molecular cloning, expression, and characterization of the human mitogen-activated protein kinase p44erk1." Molecular and Cellular Biology 13, no. 8 (August 1993): 4679–90. http://dx.doi.org/10.1128/mcb.13.8.4679.
Full textAbstract:
p44erk1 is a member of a family of tyrosyl-phosphorylated and mitogen-activated protein (MAP) kinases that participate in cell cycle control. A full-length erk1 cDNA was isolated from a human hepatoma cell line (Hep G2) library. The erk1 cDNA clone shared approximately 96% predicted amino acid identity with partial sequences of rodent erk1 cognates, and the erk1 gene was assigned to human chromosome 16 by hybrid panel analysis. Human erk1 expressed in Escherichia coli as a glutathione S-transferase fusion (GST-Erk1) protein was substantially phosphorylated on tyrosine in vivo. It underwent further autophosphorylation in vitro (up to 0.01 mol of P per mol) at the regulatory Tyr-204 site and at additional tyrosine and serine residues. Threonine autophosphorylation, presumably at the regulatory Thr-202 site, was also detected weakly when the recombinant kinase was incubated in the presence of manganese, but not in the presence of magnesium. Before and after cleavage of the GST-Erk1 protein with thrombin, it exhibited a relatively high level of myelin basic protein phosphotransferase activity, which could be reduced eightfold by treatment of the kinase with the protein-tyrosine phosphatase CD45, but not by treatment with the protein-serine/threonine phosphatase 2A. The protein-tyrosine kinase p56lck catalyzed phosphorylation of GST-Erk1 at two autophosphorylations sites, including Tyr-204, and at a novel site. A further fivefold stimulation of the myelin basic protein phosphotransferase activity of the GST-Erk1 was achieved in the presence of a partially purified MAP kinase kinase from sheep platelets. Under these circumstances, there was primarily an enhancement of the tyrosine phosphorylation of GST-Erk1. This MAP kinase kinase also similarly phosphorylated a catalytically compromised version of GST-Erk1 in which Lys-71 was converted to Ala by site-directed mutagenesis.
45
Charest, D. L., G. Mordret, K. W. Harder, F. Jirik, and S. L. Pelech. "Molecular cloning, expression, and characterization of the human mitogen-activated protein kinase p44erk1." Molecular and Cellular Biology 13, no. 8 (August 1993): 4679–90. http://dx.doi.org/10.1128/mcb.13.8.4679-4690.1993.
Full textAbstract:
p44erk1 is a member of a family of tyrosyl-phosphorylated and mitogen-activated protein (MAP) kinases that participate in cell cycle control. A full-length erk1 cDNA was isolated from a human hepatoma cell line (Hep G2) library. The erk1 cDNA clone shared approximately 96% predicted amino acid identity with partial sequences of rodent erk1 cognates, and the erk1 gene was assigned to human chromosome 16 by hybrid panel analysis. Human erk1 expressed in Escherichia coli as a glutathione S-transferase fusion (GST-Erk1) protein was substantially phosphorylated on tyrosine in vivo. It underwent further autophosphorylation in vitro (up to 0.01 mol of P per mol) at the regulatory Tyr-204 site and at additional tyrosine and serine residues. Threonine autophosphorylation, presumably at the regulatory Thr-202 site, was also detected weakly when the recombinant kinase was incubated in the presence of manganese, but not in the presence of magnesium. Before and after cleavage of the GST-Erk1 protein with thrombin, it exhibited a relatively high level of myelin basic protein phosphotransferase activity, which could be reduced eightfold by treatment of the kinase with the protein-tyrosine phosphatase CD45, but not by treatment with the protein-serine/threonine phosphatase 2A. The protein-tyrosine kinase p56lck catalyzed phosphorylation of GST-Erk1 at two autophosphorylations sites, including Tyr-204, and at a novel site. A further fivefold stimulation of the myelin basic protein phosphotransferase activity of the GST-Erk1 was achieved in the presence of a partially purified MAP kinase kinase from sheep platelets. Under these circumstances, there was primarily an enhancement of the tyrosine phosphorylation of GST-Erk1. This MAP kinase kinase also similarly phosphorylated a catalytically compromised version of GST-Erk1 in which Lys-71 was converted to Ala by site-directed mutagenesis.
46
Lindberg, R. A., and T. Hunter. "cDNA cloning and characterization of eck, an epithelial cell receptor protein-tyrosine kinase in the eph/elk family of protein kinases." Molecular and Cellular Biology 10, no. 12 (December 1990): 6316–24. http://dx.doi.org/10.1128/mcb.10.12.6316.
Full textAbstract:
A human epithelial (HeLa) cDNA library was screened with degenerate oligonucleotides designed to hybridize to highly conserved regions of protein-tyrosine kinases. One cDNA from this screen was shown to contain a putative protein-tyrosine kinase catalytic domain and subsequently used to isolate another cDNA from a human keratinocyte library that encompasses the entire coding region of a 976-amino-acid polypeptide. The predicted protein has an external domain of 534 amino acids with a presumptive N-terminal signal peptide, a transmembrane domain, and a cytoplasmic domain of 418 amino acids that includes a canonical protein-tyrosine kinase catalytic domain. Molecular phylogeny indicates that this protein kinase is closely related to eph and elk and that this receptor family is more closely related to the non-receptor protein-tyrosine kinase families than to other receptor protein-tyrosine kinases. Antibodies raised against a TrpE fusion protein immunoprecipitated a 130-kDa protein that became phosphorylated on tyrosine in immune complex kinase assays, indicating that this protein is a bona fide protein-tyrosine kinase. Analysis of RNA from 13 adult rat organs showed that the eck gene is expressed most highly in tissues that contain a high proportion of epithelial cells, e.g., skin, intestine, lung, and ovary. Several cell lines of epithelial origin were found to express the eck protein kinase at the protein and RNA levels. Immunohistochemical analysis of several rat organs also showed staining in epithelial cells. These observations prompted us to name this protein kinase eck, for epithelial cell kinase.
47
Lindberg, R. A., and T. Hunter. "cDNA cloning and characterization of eck, an epithelial cell receptor protein-tyrosine kinase in the eph/elk family of protein kinases." Molecular and Cellular Biology 10, no. 12 (December 1990): 6316–24. http://dx.doi.org/10.1128/mcb.10.12.6316-6324.1990.
Full textAbstract:
A human epithelial (HeLa) cDNA library was screened with degenerate oligonucleotides designed to hybridize to highly conserved regions of protein-tyrosine kinases. One cDNA from this screen was shown to contain a putative protein-tyrosine kinase catalytic domain and subsequently used to isolate another cDNA from a human keratinocyte library that encompasses the entire coding region of a 976-amino-acid polypeptide. The predicted protein has an external domain of 534 amino acids with a presumptive N-terminal signal peptide, a transmembrane domain, and a cytoplasmic domain of 418 amino acids that includes a canonical protein-tyrosine kinase catalytic domain. Molecular phylogeny indicates that this protein kinase is closely related to eph and elk and that this receptor family is more closely related to the non-receptor protein-tyrosine kinase families than to other receptor protein-tyrosine kinases. Antibodies raised against a TrpE fusion protein immunoprecipitated a 130-kDa protein that became phosphorylated on tyrosine in immune complex kinase assays, indicating that this protein is a bona fide protein-tyrosine kinase. Analysis of RNA from 13 adult rat organs showed that the eck gene is expressed most highly in tissues that contain a high proportion of epithelial cells, e.g., skin, intestine, lung, and ovary. Several cell lines of epithelial origin were found to express the eck protein kinase at the protein and RNA levels. Immunohistochemical analysis of several rat organs also showed staining in epithelial cells. These observations prompted us to name this protein kinase eck, for epithelial cell kinase.
48
Hunter, Tony. "Discovering the first tyrosine kinase." Proceedings of the National Academy of Sciences 112, no. 26 (June 30, 2015): 7877–82. http://dx.doi.org/10.1073/pnas.1508223112.
Full textAbstract:
In the middle of the 20th century, animal tumor viruses were heralded as possible models for understanding human cancer. By the mid-1970s, the molecular basis by which tumor viruses transform cells into a malignant state was beginning to emerge as the first viral genomic sequences were reported and the proteins encoded by their transforming genes were identified and characterized. This was a time of great excitement and rapid progress. In 1978, prompted by the discovery from Ray Erikson’s group that the Rous sarcoma virus (RSV) v-Src–transforming protein had an associated protein kinase activity specific for threonine, my group at the Salk Institute set out to determine whether the polyomavirus middle T-transforming protein had a similar kinase activity. Here, I describe the experiments that led to the identification of a kinase activity associated with middle T antigen and our serendipitous discovery that this activity was specific for tyrosine in vitro, and how this in turn led to the fortuitous observation that the v-Src–associated kinase activity was also specific for tyrosine. Our finding that v-Src increased the level of phosphotyrosine in cellular proteins in RSV-transformed cells confirmed that v-Src is a tyrosine kinase and transforms cells by phosphorylating proteins on tyrosine. My colleague Bart Sefton and I reported these findings in the March issue of PNAS in 1980. Remarkably, all of the experiments in this paper were accomplished in less than one month.
49
Uhm, J. "Mosaic Amplification of Multiple Receptor Tyrosine Kinase Genes in Glioblastoma." Yearbook of Neurology and Neurosurgery 2012 (January 2012): 100–101. http://dx.doi.org/10.1016/j.yneu.2012.05.036.
Full text
50
Krady, J. Kyle, Anirban Basu, Steven W. Levison, and Robert J. Milner. "Differential expression of protein tyrosine kinase genes during microglial activation." Glia 40, no. 1 (September 16, 2002): 11–24. http://dx.doi.org/10.1002/glia.10101.
Full text