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Iwaki, Aya, and Shingo Izawa. "Acidic stress induces the formation of P-bodies, but not stress granules, with mild attenuation of bulk translation in Saccharomyces cerevisiae." Biochemical Journal 446, no. 2 (August 14, 2012): 225–33. http://dx.doi.org/10.1042/bj20120583.
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The stress response of eukaryotic cells often causes an attenuation of bulk translation activity and the accumulation of non-translating mRNAs into cytoplasmic mRNP (messenger ribonucleoprotein) granules termed cytoplasmic P-bodies (processing bodies) and SGs (stress granules). We examined effects of acidic stress on the formation of mRNP granules compared with other forms of stress such as glucose deprivation and a high Ca2+ level in Saccharomyces cerevisiae. Treatment with lactic acid clearly caused the formation of P-bodies, but not SGs, and also caused an attenuation of translation initiation, albeit to a lesser extent than glucose depletion. P-body formation was also induced by hydrochloric acid and sulfuric acid. However, lactic acid in SD (synthetic dextrose) medium with a pH greater than 3.0, propionic acid and acetic acid did not induce P-body formation. The results of the present study suggest that the assembly of yeast P-bodies can be induced by external conditions with a low pH and the threshold was around pH 2.5. The P-body formation upon acidic stress required Scd6 (suppressor of clathrin deficiency 6), a component of P-bodies, indicating that P-bodies induced by acidic stress have rules of assembly different from those induced by glucose deprivation or high Ca2+ levels.
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Nelson, K. K., and S. K. Lemmon. "Suppressors of clathrin deficiency: overexpression of ubiquitin rescues lethal strains of clathrin-deficient Saccharomyces cerevisiae." Molecular and Cellular Biology 13, no. 1 (January 1993): 521–32. http://dx.doi.org/10.1128/mcb.13.1.521-532.1993.
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Clathrin-mediated vesicular transport is important for normal growth of the yeast Saccharomyces cerevisiae. Previously, we identified a genetic locus (SCD1) that influences the ability of clathrin heavy-chain-deficient (Chc-) yeast cells to survive. With the scd1-v allele, Chc- yeast cells are viable but grow poorly; with the scd1-i allele, Chc- cells are inviable. To identify the SCD1 locus and other genes that can rescue chc1 delta scd1-i cells to viability, a multicopy suppressor selection strategy was developed. A strain of scd1-i genotype carrying the clathrin heavy-chain gene under GAL1 control (GAL1:CHC1) was transformed with a YEp24 yeast genomic library, and colonies that could grow on glucose were selected. Plasmids from six distinct genetic loci, none of which encoded CHC1, were recovered. One of the suppressor loci was shown to be UBI4, the polyubiquitin gene. UBI4 rescues only in high copy number and is not allelic to SCD1. The conjugation of ubiquitin to intracellular proteins can mediate their selective degradation. Since UBI4 is required for survival of yeast cells under stress and is induced during starvation, ubiquitin expression in GAL1:CHC1 cells was examined. After a shift to growth on glucose to repress synthesis of clathrin heavy chains, UBI4 mRNA levels were elevated > 10-fold, whereas the quantity of free ubiquitin declined severalfold relative to that of Chc+ cells. In addition, novel higher-molecular-weight ubiquitin conjugates appeared in clathrin-deficient cells. We suggest that higher levels of ubiquitin are required for turnover of mislocalized or improperly processed proteins that accumulate in the absence of clathrin and that ubiquitin may play a general role in turnover of proteins in the secretory or endocytic pathway.
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Nelson, K. K., and S. K. Lemmon. "Suppressors of clathrin deficiency: overexpression of ubiquitin rescues lethal strains of clathrin-deficient Saccharomyces cerevisiae." Molecular and Cellular Biology 13, no. 1 (January 1993): 521–32. http://dx.doi.org/10.1128/mcb.13.1.521.
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Clathrin-mediated vesicular transport is important for normal growth of the yeast Saccharomyces cerevisiae. Previously, we identified a genetic locus (SCD1) that influences the ability of clathrin heavy-chain-deficient (Chc-) yeast cells to survive. With the scd1-v allele, Chc- yeast cells are viable but grow poorly; with the scd1-i allele, Chc- cells are inviable. To identify the SCD1 locus and other genes that can rescue chc1 delta scd1-i cells to viability, a multicopy suppressor selection strategy was developed. A strain of scd1-i genotype carrying the clathrin heavy-chain gene under GAL1 control (GAL1:CHC1) was transformed with a YEp24 yeast genomic library, and colonies that could grow on glucose were selected. Plasmids from six distinct genetic loci, none of which encoded CHC1, were recovered. One of the suppressor loci was shown to be UBI4, the polyubiquitin gene. UBI4 rescues only in high copy number and is not allelic to SCD1. The conjugation of ubiquitin to intracellular proteins can mediate their selective degradation. Since UBI4 is required for survival of yeast cells under stress and is induced during starvation, ubiquitin expression in GAL1:CHC1 cells was examined. After a shift to growth on glucose to repress synthesis of clathrin heavy chains, UBI4 mRNA levels were elevated > 10-fold, whereas the quantity of free ubiquitin declined severalfold relative to that of Chc+ cells. In addition, novel higher-molecular-weight ubiquitin conjugates appeared in clathrin-deficient cells. We suggest that higher levels of ubiquitin are required for turnover of mislocalized or improperly processed proteins that accumulate in the absence of clathrin and that ubiquitin may play a general role in turnover of proteins in the secretory or endocytic pathway.
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Roy, Debadrita, and Purusharth I. Rajyaguru. "Suppressor of clathrin deficiency (Scd6)-An emerging RGG-motif translation repressor." Wiley Interdisciplinary Reviews: RNA 9, no. 5 (May 22, 2018): e1479. http://dx.doi.org/10.1002/wrna.1479.
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Munn, A. L., L. Silveira, M. Elgort, and G. S. Payne. "Viability of clathrin heavy-chain-deficient Saccharomyces cerevisiae is compromised by mutations at numerous loci: implications for the suppression hypothesis." Molecular and Cellular Biology 11, no. 8 (August 1991): 3868–78. http://dx.doi.org/10.1128/mcb.11.8.3868-3878.1991.
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The gene encoding clathrin heavy chain in Saccharomyces cerevisiae (CHC1) is not essential for growth in most laboratory strains tested. However, in certain genetic backgrounds, a deletion of CHC1 (chc1) results in cell death. Lethality in these chc1 strains is determined by a locus designated SCD1 (suppressor of clathrin deficiency) which is unlinked to CHC1 (S. K. Lemmon and E. W. Jones, Science 238:504-509, 1987). The lethal allele of SCD1 has no effect on cell growth when the wild-type version of CHC1 is present. This result led to the proposal that most yeast strains are viable in the absence of clathrin heavy chain because they possess the SCD1 suppressor. Discovery of another yeast strain that cannot grow without clathrin heavy chain has allowed us to perform a genetic test of the suppressor hypothesis. Genetic crosses show that clathrin-deficient lethality in the latter strain is conferred by a single genetic locus (termed CDL1, for clathrin-deficient lethality). By constructing strains in which CHC1 expression is regulated by the GAL10 promoter, we demonstrate that the lethal alleles of SCD1 and CDL1 are recessive. In both cases, very low expression of CHC1 can allow cells to escape from lethality. Genetic complementation and segregation analyses indicate that CDL1 and SCD1 are distinct genes. The lethal CDL1 allele does not cause a defect in the secretory pathway of either wild-type or clathrin heavy-chain-deficient yeast. A systematic screen to identify mutants unable to grow in the absence of clathrin heavy chain uncovered numerous genes similar to SCD1 and CDL1. These findings argue against the idea that viability of chc1 cells is due to genetic suppression, since this hypothesis would require the existence of a large number of unlinked genes, all of which are required for suppression. Instead, lethality appears to be a common, nonspecific occurrence when a second-site mutation arises in a strain whose cell growth is already severely compromised by the lack of clathrin heavy chain.
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Munn, A. L., L. Silveira, M. Elgort, and G. S. Payne. "Viability of clathrin heavy-chain-deficient Saccharomyces cerevisiae is compromised by mutations at numerous loci: implications for the suppression hypothesis." Molecular and Cellular Biology 11, no. 8 (August 1991): 3868–78. http://dx.doi.org/10.1128/mcb.11.8.3868.
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The gene encoding clathrin heavy chain in Saccharomyces cerevisiae (CHC1) is not essential for growth in most laboratory strains tested. However, in certain genetic backgrounds, a deletion of CHC1 (chc1) results in cell death. Lethality in these chc1 strains is determined by a locus designated SCD1 (suppressor of clathrin deficiency) which is unlinked to CHC1 (S. K. Lemmon and E. W. Jones, Science 238:504-509, 1987). The lethal allele of SCD1 has no effect on cell growth when the wild-type version of CHC1 is present. This result led to the proposal that most yeast strains are viable in the absence of clathrin heavy chain because they possess the SCD1 suppressor. Discovery of another yeast strain that cannot grow without clathrin heavy chain has allowed us to perform a genetic test of the suppressor hypothesis. Genetic crosses show that clathrin-deficient lethality in the latter strain is conferred by a single genetic locus (termed CDL1, for clathrin-deficient lethality). By constructing strains in which CHC1 expression is regulated by the GAL10 promoter, we demonstrate that the lethal alleles of SCD1 and CDL1 are recessive. In both cases, very low expression of CHC1 can allow cells to escape from lethality. Genetic complementation and segregation analyses indicate that CDL1 and SCD1 are distinct genes. The lethal CDL1 allele does not cause a defect in the secretory pathway of either wild-type or clathrin heavy-chain-deficient yeast. A systematic screen to identify mutants unable to grow in the absence of clathrin heavy chain uncovered numerous genes similar to SCD1 and CDL1. These findings argue against the idea that viability of chc1 cells is due to genetic suppression, since this hypothesis would require the existence of a large number of unlinked genes, all of which are required for suppression. Instead, lethality appears to be a common, nonspecific occurrence when a second-site mutation arises in a strain whose cell growth is already severely compromised by the lack of clathrin heavy chain.
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Nelson, K. K., M. Holmer, and S. K. Lemmon. "SCD5, a suppressor of clathrin deficiency, encodes a novel protein with a late secretory function in yeast." Molecular Biology of the Cell 7, no. 2 (February 1996): 245–60. http://dx.doi.org/10.1091/mbc.7.2.245.
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Clathrin and its associated proteins constitute a major class of coat proteins involved in vesicle budding during membrane transport. An interesting characteristic of the yeast clathrin heavy chain gene (CHC1) is that in some strains a CHC1 deletion is lethal, while in others it is not. Recently, our laboratory developed a screen that identified five multicopy suppressors that can rescue lethal strains of clathrin heavy chain-deficient yeast (Chc - scd1-i) to viability. One of these suppressors, SCD5, encodes a novel protein of 872 amino acids containing two regions of repeated motifs of unknown function. Deletion of SCD5 has shown that it is essential for cell growth at 30 degrees C. scd5-delta strains carrying low copy plasmids encoding C-terminal truncations of Scd5p are temperature sensitive for growth at 37 degrees C. At the nonpermissive temperature, cells expressing a 338-amino acid deletion (Scd5P-delta 338) accumulate an internal pool of fully glycosylated invertase and mature alpha-factor, while processing and sorting of the vacuolar hydrolase carboxypeptidase Y is normal. The truncation mutant also accumulates 80- to 100-nm vesicles similar to many late sec mutants. Moreover, at 34 degrees C, overexpression of Scd5p suppresses the temperature sensitivity of a sec2 mutant, which is blocked at a post-Golgi step of the secretory pathway. Biochemical analyses indicate that approximately 50% of Scd5p sediments with a 100,000 x g membrane fraction and is associated as a peripheral membrane protein. Overall, these results indicate that Scd5p is involved in vesicular transport at a late stage of the secretory pathway. Furthermore, this suggests that the lethality of clathrin-deficient yeast can be rescued by modulation of vesicular transport at this late secretory step.
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Lee, Saerom, Ga-Eun Lim, Yong-Nyun Kim, Hyeon-Sook Koo, and Jaegal Shim. "AP2M1 Supports TGF-β Signals to Promote Collagen Expression by Inhibiting Caveolin Expression." International Journal of Molecular Sciences 22, no. 4 (February 6, 2021): 1639. http://dx.doi.org/10.3390/ijms22041639.
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The extracellular matrix (ECM) is important for normal development and disease states, including inflammation and fibrosis. To understand the complex regulation of ECM, we performed a suppressor screening using Caenorhabditis elegans expressing the mutant ROL-6 collagen protein. One cuticle mutant has a mutation in dpy-23 that encodes the μ2 adaptin (AP2M1) of clathrin-associated protein complex II (AP-2). The subsequent suppressor screening for dpy-23 revealed the lon-2 mutation. LON-2 functions to regulate body size through negative regulation of the tumor growth factor-beta (TGF-β) signaling pathway responsible for ECM production. RNA-seq analysis showed a dominant change in the expression of collagen genes and cuticle components. We noted an increase in the cav-1 gene encoding caveolin-1, which functions in clathrin-independent endocytosis. By knockdown of cav-1, the reduced TGF-β signal was significantly restored in the dpy-23 mutant. In conclusion, the dpy-23 mutation upregulated cav-1 expression in the hypodermis, and increased CAV-1 resulted in a decrease of TβRI. Finally, the reduction of collagen expression including rol-6 by the reduced TGF-β signal influenced the cuticle formation of the dpy-23 mutant. These findings could help us to understand the complex process of ECM regulation in organism development and disease conditions.
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Henry, Kenneth R., Kathleen D'Hondt, Ji Suk Chang, David A. Nix, M. Jamie T. V. Cope, Clarence S. M. Chan, David G. Drubin, and Sandra K. Lemmon. "The Actin-Regulating Kinase Prk1p Negatively Regulates Scd5p, a Suppressor of Clathrin Deficiency, in Actin Organization and Endocytosis." Current Biology 13, no. 17 (September 2003): 1564–69. http://dx.doi.org/10.1016/s0960-9822(03)00579-7.
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Robertson, Sarah E., Subba Rao Gangi Setty, Anand Sitaram, Michael S. Marks, Robert E. Lewis, and Margaret M. Chou. "Extracellular Signal-regulated Kinase Regulates Clathrin-independent Endosomal Trafficking." Molecular Biology of the Cell 17, no. 2 (February 2006): 645–57. http://dx.doi.org/10.1091/mbc.e05-07-0662.
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Extracellular signal-regulated kinase (Erk) is widely recognized for its central role in cell proliferation and motility. Although previous work has shown that Erk is localized at endosomal compartments, no role for Erk in regulating endosomal trafficking has been demonstrated. Here, we report that Erk signaling regulates trafficking through the clathrin-independent, ADP-ribosylation factor 6 (Arf6) GTPase-regulated endosomal pathway. Inactivation of Erk induced by a variety of methods leads to a dramatic expansion of the Arf6 endosomal recycling compartment, and intracellular accumulation of cargo, such as class I major histocompatibility complex, within the expanded endosome. Treatment of cells with the mitogen-activated protein kinase kinase (MEK) inhibitor U0126 reduces surface expression of MHCI without affecting its rate of endocytosis, suggesting that inactivation of Erk perturbs recycling. Furthermore, under conditions where Erk activity is inhibited, a large cohort of Erk, MEK, and the Erk scaffold kinase suppressor of Ras 1 accumulates at the Arf6 recycling compartment. The requirement for Erk was highly specific for this endocytic pathway, because its inhibition had no effect on trafficking of cargo of the classical clathrin-dependent pathway. These studies reveal a previously unappreciated link of Erk signaling to organelle dynamics and endosomal trafficking.
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Meyer, Christoph, Eeva-Liisa Eskelinen, Medigeshi Ramarao Guruprasad, Kurt von Figura, and Peter Schu. "μ1A deficiency induces a profound increase in MPR300/IGF-II receptor internalization rate." Journal of Cell Science 114, no. 24 (December 15, 2001): 4469–76. http://dx.doi.org/10.1242/jcs.114.24.4469.
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The mannose-6-phosphate/IGF-II receptor MPR300 mediates sorting of lysosomal enzymes from the trans-Golgi network to endosomes and endocytosis of hormones, for example, of IGF-II. We analyzed transport of MPR300 in μ1A-adaptin-deficient fibroblasts, which lack a functional AP-1 clathrin adaptor complex. In μ1A-adaptin-deficient fibroblasts, the homologous MPR46 accumulates in endosomes due to a block in retrograde transport to the trans-Golgi network. The MPR300-mediated endocytosis is markedly enhanced. We demonstrate that the seven-fold increase in endocytosis is not associated with an increased steady-state concentration of receptors at the plasma membrane, but with an increased internalization rate of MPR300. Internalization of other receptors that are also endocytosed by AP-2 is not affected. More MPR300 receptors are found in clathrin-coated pits of the plasma membrane, whereas outside coated-areas, more MPR300 are concentrated in clusters and all intracellular receptors reside in endosomes, which are in equilibrium with the plasma membrane. Thus AP-1-mediated transport of MPR300 from endosomes to the TGN controls indirectly the recycling rate of the receptor between the plasma membrane and endosomes.
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Awasthi, Sanjay, Joshua Tompkins, Jyotsana Singhal, Arthur D. Riggs, Sushma Yadav, Xiwei Wu, Sharda Singh, et al. "Rlip depletion prevents spontaneous neoplasia in TP53 null mice." Proceedings of the National Academy of Sciences 115, no. 15 (March 23, 2018): 3918–23. http://dx.doi.org/10.1073/pnas.1719586115.
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TP53 (p53) is a tumor suppressor whose functions are lost or altered in most malignancies. p53 homozygous knockout (p53−/−) mice uniformly die of spontaneous malignancy, typically T-cell lymphoma. RALBP1 (RLIP76, Rlip) is a stress-protective, mercapturic acid pathway transporter protein that also functions as a Ral effector involved in clathrin-dependent endocytosis. In stark contrast to p53−/− mice, Rlip−/− mice are highly resistant to carcinogenesis. We report here that partial Rlip deficiency induced by weekly administration of an Rlip-specific phosphorothioate antisense oligonucleotide, R508, strongly inhibited spontaneous as well as benzo(a)pyrene-induced carcinogenesis in p53−/− mice. This treatment effectively prevented large-scale methylomic and transcriptomic abnormalities suggestive of inflammation found in cancer-bearing p53−/− mice. The remarkable efficiency with which Rlip deficiency suppresses spontaneous malignancy in p53−/− mice has not been observed with any previously reported pharmacologic or genetic intervention. These findings are supported by cross-breeding experiments demonstrating that hemizygous Rlip deficiency also reduces the spontaneous malignancy phenotype of p53+/− mice. Rlip is found on the cell surface, and antibodies directed against Rlip were found to inhibit growth and promote apoptosis of cell lines as effectively as Rlip siRNA. The work presented here investigates several features, including oxidative DNA damage of the Rlip–p53 association in malignant transformation, and offers a paradigm for the mechanisms of tumor suppression by p53 and the prospects of suppressing spontaneous malignancy in hereditary cancer syndromes such as Li-Fraumeni.
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Lee, Eric K., Zhaorui Lian, Kurt D'Andrea, Richard Letrero, WeiQi Sheng, Shujing Liu, J. Nathaniel Diehl, et al. "The FBXO4 Tumor Suppressor Functions as a Barrier to BrafV600E-Dependent Metastatic Melanoma." Molecular and Cellular Biology 33, no. 22 (September 9, 2013): 4422–33. http://dx.doi.org/10.1128/mcb.00706-13.
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Cyclin D1–cyclin-dependent kinase 4/6 (CDK4/6) dysregulation is a major contributor to melanomagenesis. Clinical evidence has revealed that p16INK4A, an allosteric inhibitor of CDK4/6, is inactivated in over half of human melanomas, and numerous animal models have demonstrated that p16INK4Adeletion promotes melanoma. FBXO4, a specificity factor for the E3 ligase that directs timely cyclin D1 proteolysis, has not been studied in melanoma. We demonstrate that Fbxo4 deficiency induces Braf-driven melanoma and that this phenotype depends on cyclin D1 accumulation in mice, underscoring the importance of this ubiquitin ligase in tumor suppression. Furthermore, we have identified a substrate-binding mutation,FBXO4I377M, that selectively disrupts cyclin D1 degradation while preserving proteolysis of the other known FBXO4 substrate, TRF1. The I377M mutation and Fbxo4 deficiency result in nuclear accumulation of cyclin D1, a key transforming neoplastic event. Collectively, these data provide evidence that FBXO4 dysfunction, as a mechanism for cyclin D1 overexpression, is a contributor to human malignancy.
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Mou, Pui Kei, Eun Ju Yang, Changxiang Shi, Guowen Ren, Shishi Tao, and Joong Sup Shim. "Aurora kinase A, a synthetic lethal target for precision cancer medicine." Experimental & Molecular Medicine 53, no. 5 (May 2021): 835–47. http://dx.doi.org/10.1038/s12276-021-00635-6.
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AbstractRecent advances in high-throughput sequencing technologies and data science have facilitated the development of precision medicine to treat cancer patients. Synthetic lethality is one of the core methodologies employed in precision cancer medicine. Synthetic lethality describes the phenomenon of the interplay between two genes in which deficiency of a single gene does not abolish cell viability but combined deficiency of two genes leads to cell death. In cancer treatment, synthetic lethality is leveraged to exploit the dependency of cancer cells on a pathway that is essential for cell survival when a tumor suppressor is mutated. This approach enables pharmacological targeting of mutant tumor suppressors that are theoretically undruggable. Successful clinical introduction of BRCA-PARP synthetic lethality in cancer treatment led to additional discoveries of novel synthetic lethal partners of other tumor suppressors, including p53, PTEN, and RB1, using high-throughput screening. Recent work has highlighted aurora kinase A (AURKA) as a synthetic lethal partner of multiple tumor suppressors. AURKA is a serine/threonine kinase involved in a number of central biological processes, such as the G2/M transition, mitotic spindle assembly, and DNA replication. This review introduces synthetic lethal interactions between AURKA and its tumor suppressor partners and discusses the potential of AURKA inhibitors in precision cancer medicine.
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Poffenberger, M. C., A. Metcalfe-Roach, E. Aguilar, J. Chen, B. E. Hsu, A. H. Wong, R. M. Johnson, et al. "LKB1 deficiency in T cells promotes the development of gastrointestinal polyposis." Science 361, no. 6400 (July 26, 2018): 406–11. http://dx.doi.org/10.1126/science.aan3975.
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Germline mutations in STK11, which encodes the tumor suppressor liver kinase B1 (LKB1), promote Peutz–Jeghers syndrome (PJS), a cancer predisposition syndrome characterized by the development of gastrointestinal (GI) polyps. Here, we report that heterozygous deletion of Stk11 in T cells (LThet mice) is sufficient to promote GI polyposis. Polyps from LThet mice, Stk11+/− mice, and human PJS patients display hallmarks of chronic inflammation, marked by inflammatory immune-cell infiltration, signal transducer and activator of transcription 3 (STAT3) activation, and increased expression of inflammatory factors associated with cancer progression [interleukin 6 (IL-6), IL-11, and CXCL2]. Targeting either T cells, IL-6, or STAT3 signaling reduced polyp growth in Stk11+/− animals. Our results identify LKB1-mediated inflammation as a tissue-extrinsic regulator of intestinal polyposis in PJS, suggesting possible therapeutic approaches by targeting deregulated inflammation in this disease.
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KATNER, H. "Spontaneous suppressor cell activity in patients with the acquired immune deficiency syndrome and associated conditions*1." Journal of Allergy and Clinical Immunology 79, no. 2 (February 1987): 364–70. http://dx.doi.org/10.1016/0091-6749(87)90157-6.
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Chen, Ye, Mark M. W. Chong, Rima Darwiche, Helen E. Thomas, and Thomas W. H. Kay. "Severe Pancreatitis with Exocrine Destruction and Increased Islet Neogenesis in Mice with Suppressor of Cytokine Signaling-1 Deficiency." American Journal of Pathology 165, no. 3 (September 2004): 913–21. http://dx.doi.org/10.1016/s0002-9440(10)63353-6.
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Wang, W. Y., H. H. Chang, and T. Y. Lee. "143 DEFECT OF SUPPRESSOR OF CYTOKINE SIGNALING DURING HEPATIC ISCHEMIA-REPERFUSION INJURY IN MACROSTEATOSIS LIVERS OF LEPTIN DEFICIENCY MICE." Journal of Hepatology 52 (April 2010): S64. http://dx.doi.org/10.1016/s0168-8278(10)60145-6.
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Clark, D. A., K. C. Flanders, D. Banwatt, W. Millar-Book, J. Manuel, J. Stedronska-Clark, and B. Rowley. "Murine pregnancy decidua produces a unique immunosuppressive molecule related to transforming growth factor beta-2." Journal of Immunology 144, no. 8 (April 15, 1990): 3008–14. http://dx.doi.org/10.4049/jimmunol.144.8.3008.
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Abstract Non-T small lymphocytic suppressor cells in murine allopregnancy release a potent immunosuppressive factor in vitro that is neutralized by rabbit anti-transforming growth factor (TGF)-beta. Previous studies have suggested that the decidual suppressor factor (DSF) is smaller than TGF-beta 1, and in this paper, we show that DSF on HPLC-sieving columns also elutes later than TGF-beta 2. Nevertheless, DSF has the ability to promote anchorage-independent growth of NRK fibroblasts similar to TGF-beta s. Using turkey antibodies specific for TGF-beta 1 or beta 2, we show that DSF is related to TGF-beta 2 rather than TGF-beta 1, and this relationship was confirmed by using a panel of murine mAb to TGF-subtypes. PAGE and Western blotting showed that the TGF-beta 2-reactive molecules in HPLC-purified DSF was slightly smaller than TGF-beta 2 and approximately 20 to 23 kDa. The DSF molecule is therefore closely related to TGF-beta 2 but as released from decidua, differs in size. The TGF-beta 2-related decidual suppressor factor was also obtained from the decidua of synpregnant C.B.-17 severe combined immune deficiency (SCID) and pregnant SCID-BG (C57BL/6 background) mice, confirming the lack of T or B cell dependence of DSF production and the generality of production of a TGF-beta-related suppressor factor by decidua associated with successful pregnancy in mice.
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Shields, Benjamin J., Florian Wiede, Esteban N. Gurzov, Kenneth Wee, Christine Hauser, Hong-Jian Zhu, Timothy J. Molloy, et al. "TCPTP Regulates SFK and STAT3 Signaling and Is Lost in Triple-Negative Breast Cancers." Molecular and Cellular Biology 33, no. 3 (November 19, 2012): 557–70. http://dx.doi.org/10.1128/mcb.01016-12.
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ABSTRACTTyrosine phosphorylation-dependent signaling, as mediated by members of the epidermal growth factor receptor (EGFR) family (ErbB1 to -4) of protein tyrosine kinases (PTKs), Src family PTKs (SFKs), and cytokines such as interleukin-6 (IL-6) that signal via signal transducer and activator of transcription 3 (STAT3), is critical to the development and progression of many human breast cancers. EGFR, SFKs, and STAT3 can serve as substrates for the protein tyrosine phosphatase TCPTP (PTPN2). Here we report that TCPTP protein levels are decreased in a subset of breast cancer cell linesin vitroand that TCPTP protein is absent in a large proportion of “triple-negative” primary human breast cancers. Homozygous TCPTP deficiency in murine mammary fat padsin vivois associated with elevated SFK and STAT3 signaling, whereas TCPTP deficiency in human breast cancer cell lines enhances SFK and STAT3 signaling. On the other hand, TCPTP reconstitution in human breast cancer cell lines severely impaired cell proliferation and suppressed anchorage-independent growthin vitroand xenograft growthin vivo. These studies establish TCPTP's potential to serve as a tumor suppressor in human breast cancer.
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Beutler, Ernest, Pauline Lee, Terri Gelbart, Xin Du, and Bruce Beutler. "The Mask Mutation Identifies TMPRSS6 as an Essential Suppressor of Hepcidin Gene Expression, Required for Normal Uptake of Dietary Iron." Blood 110, no. 11 (November 16, 2007): 3. http://dx.doi.org/10.1182/blood.v110.11.3.3.
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Abstract Hepcidin, the central negative regulator of iron absorption and iron release from macrophages, is upregulated by iron. Mutations in hemojuvelin, Hfe, transferrin receptor 2, and SMAD4 are known to prevent upregulation. Additionally, the bone morphogenetic proteins (BMPs), and the inflammatory cytokines IL-1 and IL-6 stimulate hepcidin gene activation. Downregulation of hepcidin is effected by anemia and hypoxia, but nothing is known of the mechanism through which this occurs. Here we describe the recessive ENU-induced phenotype Mask, so called because affected homozygotes developed regional alopecia in which truncal hair was shed while facial hair was retained. The Mask phenotype was found to be a manifestation of iron deficiency, and was eliminated by correcting the iron deficiency. When fed an iron deficient diet, mutant mice absorbed less iron than controls, as measured by total body 59Fe counting. After reaching a plateau total body counts stabilized, indicating that blood loss did not play a role in the iron deficiency. The level of liver hepcidin mRNA of iron deficient mice is normally greatly decreased; in contrast, the Mask mouse had high liver hepcidin mRNA levels. By positional cloning, we were able to ascribe the Mask phenotype to a splicing error in the Tmprss6 gene, which encodes a membrane-bound serine protease of previously unknown function. The mutation truncates the protein, eliminating the serine protease domain. Transfecting HepG2 cells to express the wildtype TMPRSS6 protein decreased baseline hepcidin reporter activity and almost entirely blunted the hepcidin inducing effect of IL-6, IL-1, hemojuvelin, and the BMPs. A construct encoding the Mask truncation mutant had diminished activity. Thus, TMPRSS6 powerfully down-regulates hepcidin gene transcription in the baseline state and prevents its upregulation by all known stimulators. TMPRSS6 is a non-redundant component of a hepcidin suppression pathway that exerts dominant effect over all known hepcidin inducing pathways, and is required for normal absorption of dietary iron.
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Tsai, Yi-Chih, Su-Liang Chen, Shu-Ling Peng, Ya-Li Tsai, Zuong-Ming Chang, Vincent Hung-Shu Chang, and Hui-Ju Ch’ang. "Upregulating sirtuin 6 ameliorates glycolysis, EMT and distant metastasis of pancreatic adenocarcinoma with krüppel-like factor 10 deficiency." Experimental & Molecular Medicine 53, no. 10 (October 2021): 1623–35. http://dx.doi.org/10.1038/s12276-021-00687-8.
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AbstractKrüppel-like factor 10 (KLF10) is a tumor suppressor in multiple cancers. In a murine model of spontaneous pancreatic adenocarcinoma (PDAC), additional KLF10 depletion accelerated distant metastasis. However, Klf10 knockout mice, which suffer from metabolic disorders, do not develop malignancy. The mechanisms of KLF10 in PDAC progression deserve further exploration. KLF10-depleted and KLF10-overexpressing PDAC cells were established to measure epithelial-mesenchymal transition (EMT), glycolysis, and migration ability. A murine model was established to evaluate the benefit of genetic or pharmacological manipulation in KLF10-depleted PDAC cells (PDACshKLF10). Correlations of KLF10 deficiency with rapid metastasis, elevated EMT, and glycolysis were demonstrated in resected PDAC tissues, in vitro assays, and murine models. We identified sirtuin 6 (SIRT6) as an essential mediator of KLF10 that modulates EMT and glucose homeostasis. Overexpressing SIRT6 reversed the migratory and glycolytic phenotypes of PDACshKLF10 cells. Linoleic acid, a polyunsaturated essential fatty acid, upregulated SIRT6 and prolonged the survival of mice injected with PDACshKLF10. Modulating HIF1α and NFκB revealed that EMT and glycolysis in PDAC cells were coordinately regulated upstream by KLF10/SIRT6 signaling. Our study demonstrated a novel KLF10/SIRT6 pathway that modulated EMT and glycolysis coordinately via NFκB and HIF1α. Activation of KLF10/SIRT6 signaling ameliorated the distant progression of PDAC.Clinical Trial Registration: ClinicalTrials.gov. identifier: NCT01666184.
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Serreze, D. V., and E. H. Leiter. "Defective activation of T suppressor cell function in nonobese diabetic mice. Potential relation to cytokine deficiencies." Journal of Immunology 140, no. 11 (June 1, 1988): 3801–7. http://dx.doi.org/10.4049/jimmunol.140.11.3801.
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Abstract Nonobese diabetic (NOD) is an inbred mouse strain susceptible to development of T cell-mediated autoimmune diabetes. The strain is characterized by high percentages of T lymphocytes in lymphoid organs. The syngeneic mixed lymphocyte reaction (SMLR), a T cell response to self MHC class II Ag, is reportedly involved in the generation of a number of immunoregulatory cells, including suppressor inducers. A severely depressed SMLR characteristic of certain other autoimmune strains was found in NOD but not in nonautoimmune SWR/Bm mice. Moreover, IL-2 produced by NOD T cells at day 6 in an SMLR was at least one hundredfold reduced compared with SWR, and NOD T cells harvested from an SMLR at day 6 were functionally defective when tested for ability to induce suppression of an allogeneic MLR. However, functionally competent suppressor T cells were generated in NOD splenic leukocyte cultures in response to Con A, and IL-2 release from these was equivalent to that released by Con A-stimulated SWR splenocytes. A deficiency in cytokine release was not limited to IL-2, because peritoneal exudate cells from NOD exhibited a greatly diminished sensitivity to LPS-stimulated IL-1 release in comparison to SWR mice. IL-2 supplementation both in vitro and in vivo restored the ability of NOD T cells to respond in a SMLR, with production of cells capable of inducing suppression. Like SMLR-activated T cells from untreated SWR controls, SMLR blasts from IL-2-treated NOD mice were enriched for the L3T4 phenotype. IL-1 supplementation in vitro resulted in partial restoration of T suppressor activation in a SMLR. The depressed SMLR exhibited by NOD mice was apparently a stimulator cell dysfunction, because NOD stimulator cells failed to activate T cells from (SWR x NOD)F1 mice, whereas stimulators from SWR or F1 mice were capable of doing so. Collectively, these results suggest a defect in suppressor cell activation rather than an absence of this immunoregulatory cell population.
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Ruden, D. M., and H. Jackle. "Mitotic delay dependent survival identifies components of cell cycle control in the Drosophila blastoderm." Development 121, no. 1 (January 1, 1995): 63–73. http://dx.doi.org/10.1242/dev.121.1.63.
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The Drosophila body pattern is laid down by maternal and zygotic factors which act during the early phase of embryonic development. During this period, nascent zygotic transcripts longer than about 6 kilobases are aborted between the rapid mitotic cycles. Resurrector1 (Res1) and Godzilla1 (God1), two newly identified dominant zygotic suppressor mutations, and a heterozygous maternal deficiency of the cyclin B locus, complement the partial loss of function of the segmentation gene knirps (kni) by extending the length of mitotic cycles at blastoderm. The mitotic delay caused by Res1 and God1 zygotically and by the deficiency of the cyclin B locus maternally allows the expression of a much longer transcript of a kni cognate gene normally aborted between the short mitotic cycles and consequently allows survival of kni mutant progeny. In addition to the practical benefits of identifying mutations in Drosophila cell cycle regulatory genes as suppressors of kni, our results have evolutionary implications regarding the flexibility of the genome to meet sudden selective pressures by recruiting cognate genes to function.
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Tuazon Kels, Ma Junaliah, Esther Ng, Zahrah Al Rumaih, Pratikshya Pandey, Sigrid R. Ruuls, Heinrich Korner, Timothy P. Newsome, Geeta Chaudhri, and Gunasegaran Karupiah. "TNF deficiency dysregulates inflammatory cytokine production, leading to lung pathology and death during respiratory poxvirus infection." Proceedings of the National Academy of Sciences 117, no. 27 (June 22, 2020): 15935–46. http://dx.doi.org/10.1073/pnas.2004615117.
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Excessive tumor necrosis factor (TNF) is known to cause significant pathology. Paradoxically, deficiency in TNF (TNF−/−) also caused substantial pathology during respiratory ectromelia virus (ECTV) infection, a surrogate model for smallpox. TNF−/−mice succumbed to fulminant disease whereas wild-type mice, and those engineered to express only transmembrane TNF (mTNF), fully recovered. TNF deficiency did not affect viral load or leukocyte recruitment but caused severe lung pathology and excessive production of the cytokines interleukin (IL)-6, IL-10, transforming growth factor beta (TGF-β), and interferon gamma (IFN-γ). Short-term blockade of these cytokines significantly reduced lung pathology in TNF−/−mice concomitant with induction of protein inhibitor of activated STAT3 (PIAS3) and/or suppressor of cytokine signaling 3 (SOCS3), factors that inhibit STAT3 activation. Consequently, inhibition of STAT3 activation with an inhibitor reduced lung pathology. Long-term neutralization of IL-6 or TGF-β protected TNF−/−mice from an otherwise lethal infection. Thus, mTNF alone is necessary and sufficient to regulate lung inflammation but it has no direct antiviral activity against ECTV. The data indicate that targeting specific cytokines or cytokine-signaling pathways to reduce or ameliorate lung inflammation during respiratory viral infections is possible but that the timing and duration of the interventive measure are critical.
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Riehle, Kimberly J., Jean S. Campbell, Ryan S. McMahan, Melissa M. Johnson, Richard P. Beyer, Theo K. Bammler, and Nelson Fausto. "Regulation of liver regeneration and hepatocarcinogenesis by suppressor of cytokine signaling 3." Journal of Experimental Medicine 205, no. 1 (December 24, 2007): 91–103. http://dx.doi.org/10.1084/jem.20070820.
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Suppressor of cytokine signaling 3 (SOCS3) down-regulates several signaling pathways in multiple cell types, and previous data suggest that SOCS3 may shut off cytokine activation at the early stages of liver regeneration (Campbell, J.S., L. Prichard, F. Schaper, J. Schmitz, A. Stephenson-Famy, M.E. Rosenfeld, G.M. Argast, P.C. Heinrich, and N. Fausto. 2001.J. Clin. Invest. 107:1285–1292). We developed Socs3 hepatocyte-specific knockout (Socs3 h-KO) mice to directly study the role of SOCS3 during liver regeneration after a two-thirds partial hepatectomy (PH). Socs3 h-KO mice demonstrate marked enhancement of DNA replication and liver weight restoration after PH in comparison with littermate controls. Without SOCS3, signal transducer and activator of transcription 3 (STAT3) phosphorylation is prolonged, and activation of the mitogenic extracellular signal-regulated kinase 1/2 (ERK1/2) is enhanced after PH. In vitro, we show that SOCS3 deficiency enhances hepatocyte proliferation in association with enhanced STAT3 and ERK activation after epidermal growth factor or interleukin 6 stimulation. Microarray analyses show that SOCS3 modulates a distinct set of genes, which fall into diverse physiological categories, after PH. Using a model of chemical-induced carcinogenesis, we found that Socs3 h-KO mice develop hepatocellular carcinoma at an accelerated rate. By acting on cytokines and multiple proliferative pathways, SOCS3 modulates both physiological and neoplastic proliferative processes in the liver and may act as a tumor suppressor.
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Westrick, Randal J., Kärt Tomberg, Amy E. Siebert, Guojing Zhu, Mary E. Winn, Sarah L. Dobies, Sara L. Manning, et al. "Sensitized mutagenesis screen in Factor V Leiden mice identifies thrombosis suppressor loci." Proceedings of the National Academy of Sciences 114, no. 36 (August 21, 2017): 9659–64. http://dx.doi.org/10.1073/pnas.1705762114.
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Factor V Leiden (F5L) is a common genetic risk factor for venous thromboembolism in humans. We conducted a sensitized N-ethyl-N-nitrosourea (ENU) mutagenesis screen for dominant thrombosuppressor genes based on perinatal lethal thrombosis in mice homozygous for F5L (F5L/L) and haploinsufficient for tissue factor pathway inhibitor (Tfpi+/−). F8 deficiency enhanced the survival of F5L/LTfpi+/− mice, demonstrating that F5L/LTfpi+/− lethality is genetically suppressible. ENU-mutagenized F5L/L males and F5L/+Tfpi+/− females were crossed to generate 6,729 progeny, with 98 F5L/LTfpi+/− offspring surviving until weaning. Sixteen lines, referred to as “modifier of Factor 5 Leiden (MF5L1–16),” exhibited transmission of a putative thrombosuppressor to subsequent generations. Linkage analysis in MF5L6 identified a chromosome 3 locus containing the tissue factor gene (F3). Although no ENU-induced F3 mutation was identified, haploinsufficiency for F3 (F3+/−) suppressed F5L/LTfpi+/− lethality. Whole-exome sequencing in MF5L12 identified an Actr2 gene point mutation (p.R258G) as the sole candidate. Inheritance of this variant is associated with suppression of F5L/LTfpi+/− lethality (P = 1.7 × 10−6), suggesting that Actr2p.R258G is thrombosuppressive. CRISPR/Cas9 experiments to generate an independent Actr2 knockin/knockout demonstrated that Actr2 haploinsufficiency is lethal, supporting a hypomorphic or gain-of-function mechanism of action for Actr2p.R258G. Our findings identify F8 and the Tfpi/F3 axis as key regulators in determining thrombosis balance in the setting of F5L and also suggest a role for Actr2 in this process.
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Toh, Han Chong, Francis Enane, Marissa Teo, Hideki Makishima, JoAnna Ng, Chit Lai Chee, Soo Fan Ang, Kiat Hon Lim, and Yogen Saunthararajah. "Translational significance of a newly identified hepatocellular carcinoma tumor suppressor gene on chromosome 8p." Journal of Clinical Oncology 31, no. 15_suppl (May 20, 2013): e15097-e15097. http://dx.doi.org/10.1200/jco.2013.31.15_suppl.e15097.
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e15097 Background: After deletion of 17p that removes the tumor suppressor gene (TSG) TP53, deletion of 8p is the next most common chromosome abnormality in hepatocellular carcinoma (HCC). However, 8p TSG are insufficiently defined. Methods: Integrated genomic analysis of HCC and non-malignant liver obtained at therapeutic segmentectomy from the same patients. Results: A minimally deleted region on 8p was identified by SNP array. This incorporated GATA4. Therefore, GATA4 was Sanger sequenced in paired HCC/non-malignant liver: recurrent somatic non-synonymous missense mutations were identified in exon 4 (V267M n=5) or exon 6 (S357T n=6, R362N n=2, T366R n=2). Biallelic abnormalities were deletion and mutation (n=6) or mutation and uniparental disomy (n=4), with mutation or deletion of at least one GATA4 allele in 29/47 (62%) of HCC cases. The other GATA4 exons were mutation free. Although missense mutation is not intrinsically expected to decrease GATA4 expression, GATA4 mRNA was significantly decreased in cases with mutation as well as deletion (p<0.01) compared to non-malignant liver or wild-type GATA4 HCC. GATA4 drives liver differentiation, and the biological significance of GATA4 deficiency was demonstrated by significant enrichment (49%) for liver differentiation genes (p<1.2exp-124, Benjamini corrected) amongst genes with decreased expression in HCC compared to non-malignant liver. From an oncogenesis perspective, the most important of these hepatocyte genes (e.g., HNF4A, CEBPD) antagonize MYC to terminate proliferation: GATA4 introduction (expression vector) into HCC cells containing mutated or deleted GATA4 (HepG2 and PLC respectively) restored HNF4A and CEBPD expression, suppressed MYC protein, upregulated p27/CDKN1B that mediates cell cycle exit by maturation and significantly decreased HCC proliferation without apoptosis. In objectively quantified immunohistochemical analyses (ImageIQ), HCC cases with GATA4 mutation/deletion had significantly increased MYC protein (p<0.05). Conclusions: 8p deletion/GATA4 mutation in HCC suppresses cell cycle exit by maturation, thus complementing 17p deletion that suppresses cell cycle exit by apoptosis.
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Zhou, Lu, Yiqiang Zhu, lengshan Mo, Mei Wang, Jie Lin, Yi Zhao, Yuanfa Feng, et al. "TLR7 controls myeloid-derived suppressor cells expansion and function in the lung of C57BL6 mice infected with Schistosoma japonicum." PLOS Neglected Tropical Diseases 16, no. 10 (October 24, 2022): e0010851. http://dx.doi.org/10.1371/journal.pntd.0010851.
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Toll-like receptors (TLRs) play an important role in the induction of innate and adaptive immune responses against Schistosoma japonicum (S. japonicum) infection. However, the role of Toll-like receptor 7 (TLR7) in the mouse lung during S. japonicum infection and the myeloid-derived suppressor cells (MDSCs) affected by the absence of TLR7 are not clearly understood. In this study, the results indicated that the MDSCs were accumulated and the proportion and activation of CD4+ and CD8+ T cells were decreased in the lung of mice at 6–7 weeks after S. japonicum infection. Then, the expression of TLR7 was detected in isolated pulmonary MDSCs and the results showed that the expression of TLR7 in MDSCs was increased after infection. Furthermore, TLR7 agonist R848 could down-regulate the induction effect of the soluble egg antigen (SEA) on pulmonary MDSCs in vitro. Meanwhile, TLR7 deficiency could promote the pulmonary MDSCs expansion and function by up-regulating the expression of PD-L1/2 and secreting of IL-10 in the mice infected with S. japonicum. Mechanistic studies revealed that S. japonicum infection and the antigen effects are mediated by NF-κB signaling. Moreover, TLR7 deficiency aggravates S. japonicum infection-induced damage in the lung, with more inflammatory cells infiltration, interstitial dilatation and granuloma in the tissue. In summary, this study indicated that TLR7 signaling inhibits the accumulation and function of MDSCs in S. japonicum infected mouse lung by down-regulating the expression of PD-L1/2 and secreting of IL-10, via NF-κB signaling.
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Gay, Jérôme, Efi Kokkotou, Michael O’Brien, Charalabos Pothoulakis, and Katia P. Karalis. "Corticotropin-Releasing Hormone Deficiency Is Associated with Reduced Local Inflammation in a Mouse Model of Experimental Colitis." Endocrinology 149, no. 7 (April 10, 2008): 3403–9. http://dx.doi.org/10.1210/en.2007-1703.
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CRH, the hypothalamic component of the hypothalamic-pituitary adrenal axis, attenuates inflammation through stimulation of glucocorticoid release, whereas peripherally expressed CRH acts as a proinflammatory mediator. CRH is expressed in the intestine and up-regulated in patients with ulcerative colitis. However, its pathophysiological significance in intestinal inflammatory diseases has just started to emerge. In a mouse model of acute, trinitrobenzene sulfonic acid-induced experimental colitis, we demonstrate that, despite low glucocorticoid levels, CRH-deficient mice develop substantially reduced local inflammatory responses. These effects were shown by histological scoring of tissue damage and neutrophil infiltration. At the same time, CRH deficiency was found to be associated with higher serum leptin and IL-6 levels along with sustained anorexia and weight loss, although central CRH has been reported to be a strong appetite suppressor. Taken together, our results support an important proinflammatory role for CRH during mouse experimental colitis and possibly in inflammatory bowel disease in humans. Moreover, the results suggest that CRH is involved in homeostatic pathways that link inflammation and metabolism.
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Liao, Nannan, Till Koehne, Jan Tuckermann, Ioanna Triviai, Michael Amling, Jean-Pierre David, Thorsten Schinke, and Julia Luther. "Osteoblast-specific inactivation of p53 results in locally increased bone formation." PLOS ONE 16, no. 11 (November 18, 2021): e0249894. http://dx.doi.org/10.1371/journal.pone.0249894.
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Inactivation of the tumor suppressor p53 (encoded by the Trp53 gene) is relevant for development and growth of different cancers, including osteosarcoma, a primary bone tumor mostly affecting children and young adolescents. We have previously shown that deficiency of the ribosomal S6 kinase 2 (Rsk2) limits osteosarcoma growth in a transgenic mouse model overexpressing the proto-oncogene c-Fos. Our initial aim for the present study was to address the question, if Rsk2 deficiency would also influence osteosarcoma growth in another mouse model. For that purpose, we took advantage of Trp53fl/fl mice, which were crossed with Runx2Cre transgenic mice in order to inactivate p53 specifically in osteoblast lineage cells. However, since we unexpectedly identified Runx2Cre-mediated recombination also in the thymus, the majority of 6-month-old Trp53fl/fl;Runx2-Cre (thereafter termed Trp53Cre) animals displayed thymic lymphomas, similar to what has been described for Trp53-deficient mice. Since we did not detect osteosarcoma formation at that age, we could not follow our initial aim, but we studied the skeletal phenotype of Trp53Cre mice, with or without additional Rsk2 deficiency. Here we unexpectedly observed that Trp53Cre mice display a unique accumulation of trabecular bone in the midshaft region of the femur and the humerus, consistent with its previously established role as a negative regulator of osteoblastogenesis. Since this local bone mass increase in Trp53Cre mice was significantly reduced by Rsk2 deficiency, we isolated bone marrow cells from the different groups of mice and analyzed their behavior ex vivo. Here we observed a remarkable increase of colony formation, osteogenic differentiation and proliferation in Trp53Cre cultures, which was unaffected by Rsk2 deficiency. Our data thereby confirm a critical and tumorigenesis-independent function of p53 as a key regulator of mesenchymal cell differentiation.
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Ko, Hyun-Jeong, Seong-Ryeol Kim, Jae-Hyoung Song, and Bo-Ra Lee. "Interleukin-10 Attenuate Tumor Growth by Inhibiting Interleukin-6/Signal transducer and activator of transcription 3 Axis on Myeloid-Derived Suppressor Cells." Journal of Immunology 196, no. 1_Supplement (May 1, 2016): 73.5. http://dx.doi.org/10.4049/jimmunol.196.supp.73.5.
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Abstract Interleukin-10 (IL-10) is a well-characterized anti-inflammatory cytokine, but its role in anti-cancer immunity is still controversial. When IL-10 knockout (IL-10−/−) mice were subcutaneously injected with TC-1 cancer cells expressing HPV E6 and E7 proteins, tumor growth was accelerated in IL-10−/− mice than wild type (WT) mice. Interestingly, the production of interleukin-6 (IL-6) was significantly increased in IL-10−/− mice than in WT mice. The blockade of IL-6 by systemic administration of anti-IL-6 receptor (IL-6R) monoclonal antibody (mAb) inhibited the tumor growth, and inhibited the generation of myeloid-derived suppressor cells (MDSCs) which were increased in IL-10 deficiency. MDSCs and tumor cells from IL-10−/− mice had increased phosphorylated-signal transducer and activator of transcription 3 (p-STAT3) level and the treatment of mice with STAT3 inhibitor, S3I reduced the tumor growth and inhibited the expansion of MDSCs. Accordingly, S3I also reduced tumor necrosis factor-a (TNF-a) and IL-6 in tumor. Although S3I treatment relieved T cells from suppression by reducing the number of MDSCs, it seems that the proportion of MDSCs having M2 macrophage phenotype was increased among myeloid cells since the transcription of Arg1, Mgl1, Mgl2, and, Fizz1, were significantly increased and the expression of some suppressive functional mediators of MDSCs, including Arg1 and iNOS was up-regulated. Combined treatment of anti-IL-6 mAb and S3I could further suppress the generation of M2-type MDSC and regulate immunosuppressive function of MDSCs, and finally inhibited tumor growth more efficaciously. These results suggested that combined inhibition of IL-6 and STAT3 could be a potential candidate as an anti-cancer strategy.
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Fong, Louise Y., Cristian Taccioli, Alexey Palamarchuk, Guidantonio Malagoli Tagliazucchi, Ruiyan Jing, Karl J. Smalley, Sili Fan, et al. "Abrogation of esophageal carcinoma development in miR-31 knockout rats." Proceedings of the National Academy of Sciences 117, no. 11 (March 2, 2020): 6075–85. http://dx.doi.org/10.1073/pnas.1920333117.
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MicroRNA-31 (miR-31) is overexpressed in esophageal squamous cell carcinoma (ESCC), a deadly disease associated with dietary Zn deficiency and inflammation. In a Zn deficiency-promoted rat ESCC model with miR-31 up-regulation, cancer-associated inflammation, and a high ESCC burden followingN-nitrosomethylbenzylamine (NMBA) exposure, systemic antimiR-31 delivery reduced ESCC incidence from 85 to 45% (P= 0.038) and miR-31 gene knockout abrogated development of ESCC (P= 1 × 10−6). Transcriptomics, genome sequencing, and metabolomics analyses in these Zn-deficient rats revealed the molecular basis of ESCC abrogation by miR-31 knockout. Our identification of EGLN3, a known negative regulator of nuclear factor κB (NF-κB), as a direct target of miR-31 establishes a functional link between oncomiR-31, tumor suppressor target EGLN3, and up-regulated NF-κB–controlled inflammation signaling. Interaction among oncogenic miR-31, EGLN3 down-regulation, and inflammation was also documented in human ESCCs. miR-31 deletion resulted in suppression of miR-31–associated EGLN3/NF-κB–controlled inflammatory pathways. ESCC-free, Zn-deficient miR-31−/−rat esophagus displayed no genome instability and limited metabolic activity changes vs. the pronounced mutational burden and ESCC-associated metabolic changes of Zn-deficient wild-type rats. These results provide conclusive evidence that miR-31 expression is necessary for ESCC development.
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Hornick, Emma L., Laura L. Stunz, Shakoora Sabree, Xiaosheng Wu, Thomas E. Witzig, and Gail A. Bishop. "The Tumor Suppressor Protein TRAF3 Modulates GSK3 Activity and Susceptibility of B Lymphoma Cells to GSK3 Inhibition." Cancers 14, no. 20 (October 14, 2022): 5029. http://dx.doi.org/10.3390/cancers14205029.
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TNF receptor-associated factor 3 (TRAF3) is an adapter protein that inhibits many signals that promote B cell survival and activation. Mice with a B cell-specific TRAF3 deficiency and humans with a rare haploinsufficiency in TRAF3 have enhanced development of BCLs as they age. Loss-of-function mutations in TRAF3 are common in B cell malignancies. Recent studies show that pharmacological inhibition of the enzyme glycogen synthase kinase 3 (GSK3), which regulates cellular growth, survival, and metabolism, inhibits growth and survival of BCL-derived B cells. In this study, we found that TRAF3 and GSK3 associated in B cells. The relative levels of TRAF3 in BCL cell lines correlated positively with the ratio of inactive to total GSK3β, and negatively correlated with susceptibility to GSK3 inhibition by the GSK3 inhibitory drug 9-ING-41, currently in clinical trials. Uniquely in BCLs with low TRAF3, GSK3 inhibition caused increased loss of the TRAF3-regulated, anti-apoptotic protein Mcl-1. GSK3 inhibition also blocked hyperresponsiveness to IL-6 receptor signaling in TRAF3-deficient BCL cells. Together, these results support the utility of 9-ING-41 as a treatment for BCL, and suggest that a decrease or loss of TRAF3 in BCLs could act as a biomarker for increased susceptibility to GSK3 inhibitor treatment.
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Pilot, Thomas, Aurélie Fratti, Chloé Thinselin, Anaïs Perrichet, Lucie Demontoux, Emeric Limagne, Valentin Derangère, et al. "Heat shock and HSP70 regulate 5-FU-mediated caspase-1 activation in myeloid-derived suppressor cells and tumor growth in mice." Journal for ImmunoTherapy of Cancer 8, no. 1 (May 2020): e000478. http://dx.doi.org/10.1136/jitc-2019-000478.
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BackgroundWe have previously shown that 5-fluorouracil (5-FU) selectively kills myeloid-derived suppressor cells (MDSCs) and activates NLRP3 (NOD-leucine rich repeat and pyrin containing protein 3) inflammasome. NLRP3 activation leads to caspase-1 activation and production of IL-1β, which in turn favors secondary tumor growth. We decided to explore the effects of either a heat shock (HS) or the deficiency in heat shock protein (HSP) 70, previously shown to respectively inhibit or increase NLRP3 inflammasome activation in macrophages.MethodsCaspase-1 activation was detected in vitro in MSC-2 cells by western blot and in vivo or ex vivo in tumor and/or splenic MDSCs by flow cytometry. The effects of HS, HSP70 deficiency and anakinra (an IL-1 inhibitor) on tumor growth and mice survival were studied in C57BL/6 WT orHsp70−/−tumor-bearing mice. Finally, Th17 polarization was evaluated by qPCR (Il17a, Rorc) and angiogenic markers by qPCR (Pecam1, Eng) and immunohistochemistry (ERG).ResultsHS inhibits 5-FU-mediated caspase-1 activation in vitro and in vivo without affecting its cytotoxicity on MDSCs. Moreover, it enhances the antitumor effect of 5-FU treatment and favors mice survival. Interestingly, it is associated to a decreased Th17 and angiogenesis markers in tumors. IL-1β injection is able to bypass HS+5-FU antitumor effects. In contrast, inHsp70−/−MDSCs, 5-FU-mediated caspase-1 activation is increased in vivo and in vitro without effect on 5-FU cytotoxicity. InHsp70−/−mice, the antitumor effect of 5-FU was impeded, with an increased Th17 and angiogenesis markers in tumors. Finally, the effects of 5-FU on tumor growth can be restored by inhibiting IL-1β, using anakinra.ConclusionThis study provides evidence on the role of HSP70 in tuning 5-FU antitumor effect and suggests that HS can be used to improve 5-FU anticancer effect.
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McRae, Helen M., Samantha Eccles, Lachlan Whitehead, Warren S. Alexander, Jozef Gécz, Tim Thomas, and Anne K. Voss. "Downregulation of the GHRH/GH/IGF1 axis in a mouse model of Börjeson-Forssman-Lehman syndrome." Development 147, no. 21 (September 29, 2020): dev187021. http://dx.doi.org/10.1242/dev.187021.
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ABSTRACTBörjeson-Forssman-Lehmann syndrome (BFLS) is an intellectual disability and endocrine disorder caused by plant homeodomain finger 6 (PHF6) mutations. Individuals with BFLS present with short stature. We report a mouse model of BFLS, in which deletion of Phf6 causes a proportional reduction in body size compared with control mice. Growth hormone (GH) levels were reduced in the absence of PHF6. Phf6−/Y animals displayed a reduction in the expression of the genes encoding GH-releasing hormone (GHRH) in the brain, GH in the pituitary gland and insulin-like growth factor 1 (IGF1) in the liver. Phf6 deletion specifically in the nervous system caused a proportional growth defect, indicating a neuroendocrine contribution to the phenotype. Loss of suppressor of cytokine signaling 2 (SOCS2), a negative regulator of growth hormone signaling partially rescued body size, supporting a reversible deficiency in GH signaling. These results demonstrate that PHF6 regulates the GHRH/GH/IGF1 axis.
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Kim, Byung-Gyu, Sung Hee Choi, Janet Robinson, and John Letterio. "Loss of p27 enhances gastrointestinal eptithelial malignancy in mice with Smad4 deficiency in T cells. (120.17)." Journal of Immunology 188, no. 1_Supplement (May 1, 2012): 120.17. http://dx.doi.org/10.4049/jimmunol.188.supp.120.17.
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Abstract We previously reported that selective loss of Smad4 signaling in T cells (Smad4KO) leads to spontaneous gastrointestinal inflammation and cancer. The expression of p27Kip1 is significantly decreased in the colon of Smad4KO mice, thus we investigated the role of p27Kip1 in the maintenance of intestinal mucosal immune and epithelial homeostasis. Smad4KO and p27Kip1 germ line mutation (Smad4/p27DKO) mice develop spontaneous gastrointestinal inflammation and carcinoma at 1-2 months of age and 3 months of age, respectively. Smad4/p27DKO had little in vitro TGF-β-induced Foxp3+CD4+ T cells (iTregs) and decreased natural Foxp3+CD4+ T cells (nTregs) in vivo, whereas Smad4KO had little iTregs and p27KO had decreased nTregs. Furthermore, Smad4/p27DKO have an increased number of mucosal CD4+ T cells and an increased percentage of memory T cells. The production of pro-inflammatory cytokines including IL-6, IFN-γ and TNF-α was greatly increased in the colon mucosa of Smad4/p27DKO mice. The colon epithelium of Smad4/p27DKO mice exhibit an oncogenic signature with a significant elevation in expression of iNOS, p-stat1 and p-stat3 and a repression of the tumor suppressor gene, 15-PGDH when compared with WT, p27KO and Smad4KO mice. Thus, our data indicate that p27Kip1 deficiency promotes gastrointestinal epithelial malignancy through both enhancement of the activation of pathogenic T cells and through promotion of the expression of pro-inflammatory mediators by mucosal epithelial cells.
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Krömer, G., K. Schauenstein, N. Neu, K. Stricker, and G. Wick. "In vitro T cell hyperreactivity in Obese strain (OS) chickens is due to a defect in nonspecific suppressor mechanism(s)." Journal of Immunology 135, no. 4 (October 1, 1985): 2458–63. http://dx.doi.org/10.4049/jimmunol.135.4.2458.
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Abstract Spontaneous autoimmune thyroiditis of OS chickens is associated with a marked hyperreactivity of the T cell system. The purpose of the present study was to investigate the underlying regulatory mechanisms. Co-cultivation experiments between Con A-stimulated OS and NWL lymphocytes in communicating cultures revealed soluble regulatory factors to be responsible for the observed functional differences: the high proliferative response to Con A and hyperproduction of IL 2 of OS cells was found to be due to a deficiency in the conditioned medium of dialyzable inhibitory factor(s) that regulate IL 2 secretion of NWL lymphocytes. Furthermore, sera of young NWL chickens were found to profoundly inhibit the IL 2-promoted lymphoblast proliferation. This IL 2 antagonizing activity is lost with age (3 to 6 yr) and was found to be significantly diminished in OS birds throughout ontogeny, thus pointing to possible parallels between immune regulatory dysfunction in autoimmunity and in physiologic aging. Both enhanced T cell response and the defect in serum suppressor were inherited by (OS X CB)F1 animals, indicating that these two aberrations may be related to each other.
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Hoffmann, Dirk, Johanna Sens, Sebastian Brennig, Daniel Brand, Friederike Philipp, Philippe Vollmer Barbosa, Johannes Kuehle, et al. "Genetic Correction of IL-10RB Deficiency Reconstitutes Anti-Inflammatory Regulation in iPSC-Derived Macrophages." Journal of Personalized Medicine 11, no. 3 (March 20, 2021): 221. http://dx.doi.org/10.3390/jpm11030221.
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Patient material from rare diseases such as very early-onset inflammatory bowel disease (VEO-IBD) is often limited. The use of patient-derived induced pluripotent stem cells (iPSCs) for disease modeling is a promising approach to investigate disease pathomechanisms and therapeutic strategies. We successfully developed VEO-IBD patient-derived iPSC lines harboring a mutation in the IL-10 receptor β-chain (IL-10RB) associated with defective IL-10 signaling. To characterize the disease phenotype, healthy control and VEO-IBD iPSCs were differentiated into macrophages. IL-10 stimulation induced characteristic signal transducer and activator of transcription 3 (STAT3) and suppressor of cytokine signaling 3 (SOCS3) downstream signaling and anti-inflammatory regulation of lipopolysaccharide (LPS)-mediated cytokine secretion in healthy control iPSC-derived macrophages. In contrast, IL-10 stimulation of macrophages derived from patient iPSCs did not result in STAT3 phosphorylation and subsequent SOCS3 expression, recapitulating the phenotype of cells from patients with IL-10RB deficiency. In line with this, LPS-induced cytokine secretion (e.g., IL-6 and tumor necrosis factor-α (TNF-α)) could not be downregulated by exogenous IL-10 stimulation in VEO-IBD iPSC-derived macrophages. Correction of the IL-10RB defect via lentiviral gene therapy or genome editing in the adeno-associated virus integration site 1 (AAVS1) safe harbor locus led to reconstitution of the anti-inflammatory response. Corrected cells showed IL-10RB expression, IL-10-inducible phosphorylation of STAT3, and subsequent SOCS3 expression. Furthermore, LPS-mediated TNF-α secretion could be modulated by IL-10 stimulation in gene-edited VEO-IBD iPSC-derived macrophages. Our established disease models provide the opportunity to identify and validate new curative molecular therapies and to investigate phenotypes and consequences of additional individual IL-10 signaling pathway-dependent VEO-IBD mutations.
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Lee, Young-Kwan, Ji-Min Ju, Myung-Soo Kang, Chang-Ki Min, Woo-Jeong Shon, Dong-Mi Shin, and Eun Young Choi. "Impairment of myeloid-derived suppressor cell expansion and enhancement of dendritic cell differentiation of MyD88-deficient bone marrow cells in graft-versus-host disease." Journal of Immunology 198, no. 1_Supplement (May 1, 2017): 140.2. http://dx.doi.org/10.4049/jimmunol.198.supp.140.2.
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Abstract Myeloid differentiation 88 (MyD88) is an adaptor molecule in toll-like receptor-associated signaling. To examine the effect of MyD88-deficiency in myeloid cell differentiation under inflammatory condition, we transplanted bone marrow (BM) cells isolated from MyD88-deficient C57BL/6 (B6) mice together with B6 T cells into MHC-matched allogeneic BALB.B strain mice. This transplantation induced more serious graft-versus-host disease in the BALB.B BM recipients, compared with the GVHD induced in the control BALB.B mice which received wild type B6 BM and T cells. The aggravation of GVHD was associated with impaired expansion of CD11b+Gr1+ myeloid derived suppressor cells (MDSCs) from the MyD88-deficient BM cells during the GVHD development. Instead of MDSC expansion, myeloid cells from MyD88-deficient BM cells showed enhanced apoptosis and dendritic cell (DC) differentiation, which resulted in enhancement of activation of allo-reactive B6 T cells. These results provide information helpful for the understanding of the role of MyD88 in myeloid cell differentiation and development of strategy to reduce the GVHD severity.
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Cho, Kyung-Ah, Jee Won Suh, and So-Youn Woo. "Immune regulatory mechanism in mouse IBD model (168.26)." Journal of Immunology 186, no. 1_Supplement (April 1, 2011): 168.26. http://dx.doi.org/10.4049/jimmunol.186.supp.168.26.
Full textAbstract:
Abstract Inflammatory bowel disease (IBD) in humans, such as Crohn’s disease and ulcerative colitis, is characterized by chronic inflammation with dysregulated mucosal immune tolerance. Some reported that immune responses to intestinal microbiota are critical to develop IBD and NOD-like receptor-mediated inflammasome is related to epithelial integrity. In order to investigate dysregulated mechanism focusing on the relationship between inflammasome and regulatory cells in IBD, we used a 2, 4, 6-trinitrobenzene sulfonic acid (TNBS)-induced colitis model in WT-C57BL/6 mice and caspase-1-/- mice. In addition, to observe Th2-dependent immune responses in IBD mouse, we also used WT-Balb/c mice and eosinophil-deficient ΔdblGATA mice. As a result, mice treated TNBS had significant changes in clinical parameters, weight loss, and colonic weights compared with those of sham control mice. Histologic analysis also showed severe inflammation in intestines of TNBS treated mice. We found that deficiency of eosinophil or IL-1β had resistance in initial stage of disease progression, represented by survival score and body weight loss and we also noted that proportion of Foxp3+ cells and myeloid-derived suppressor cells (MDSCs) increased in above TNBS-treated ΔdblGATA and caspase-1-/- mice. These results provide us another mechanism of immune modulation in the pathogenesis of intestinal inflammation.
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Balomenos, Dimitrios, Cristina Vazquez-Mateo, Kathrin Weber, and Carlos Martínez-A. "p21 suppresses autoimmunity by regulating ERK1/2 phosphorylation, NF-κB activation and IFNγ production in hyperactivated T cells (BA4P.141)." Journal of Immunology 194, no. 1_Supplement (May 1, 2015): 47.21. http://dx.doi.org/10.4049/jimmunol.194.supp.47.21.
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Abstract Peripheral tolerance is essential for suppressing autoimmunity and depends on the regulation of T cell activation, proliferation and apoptosis. p21, known originally as a cell cycle inhibitor, is implicated in effector/memory T cell responses and is considered a suppressor of lupus-like autoimmunity. However, the mechanism by which p21 regulates T cell responses and autoimmunity remains unknown. We investigated these questions by studying T cells of C57BL/6 p21-/- and C57BL/6-lpr p21-/- mice. p21 regulated activation of repeatedly stimulated effector/memory T cells in a cell cycle-independent manner, but had no effect on naïve T cells. We identified a previously non-described role for p21, in controlling T cell activation via downstream MAPK and NF-κB pathways in the cytoplasm. Lack of p21 caused hyperactivation of ERK1/2 and NF-κB in reactivated T cells. Importantly, p21-deficiency led to increased IFNγ production due to an effect of p21 on the IL-12- and IL-18-dependent IFNγ induction. Stimulation of TCR-transgenic T cells with specific antigens in vivo confirmed that p21 is involved in effector/memory T cell activation. Lack of p21 dramatically increased the mild lupus symptoms of C57BL/6-lpr mice to full-blown disease and death in C57BL/6-lpr p21-/- mice. The mechanism by which p21 regulates disease severity was based on its control of T cell activation and IFNγ production. Analysis of these p21 functions might be of therapeutic value for Lupus.
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Kidder, Koby S., Zhen Bian, Lei Shi, and Yuan Liu. "Arginase-1 is neither constitutively expressed in nor require for myeloid-derived suppressor cell (MDSC)-mediated inhibition of T cell proliferation." Journal of Immunology 200, no. 1_Supplement (May 1, 2018): 177.7. http://dx.doi.org/10.4049/jimmunol.200.supp.177.7.
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Abstract Although previous reports suggest that tumor-induced myeloid-derived suppressor cells (MDSC) inhibit T cell proliferation by L-arginine depletion through arginase-1 activity, we herein show that arginase-1 is neither inherently expressed in MDSC nor required for MDSC-mediated inhibition of T cell proliferation. Employing Percoll density gradient centrifugation, large expansions of MDSC in the bone marrow of tumor-bearing mice were isolated and exhibit potent inhibition on T cell proliferation induced via TCR-ligation, Concanavalin A, PMA plus ionomycin, or IL-2. These MDSC, though suppressive toward T cell proliferation, do not hinder TCR-induced IFNγ secretion or granzyme B expression. Despite effectively inhibiting T cell proliferation, both G- and M-MDSC exhibit no arginase-1 expression; however, arginase-1 can be induced by exposure to TCR-activated T cells or their culture medium, but not by T cells activated by other means or growing tumor cells. Western blot analysis revealed that TCR-activated T cells confer arginase-1 expression in MDSC by secreting cytokines that manifest as two distinct signaling-relay axes, IL-6-to-IL-4 or GM-CSF/IL-4-to-IL-10. Specifically, IL-6 signaling increases IL-4R on MDSC, enabling IL-4 to induce arginase-1 expression; similarly, GM-CSF in concert with IL-4 induces IL-10R on MDSC, allowing IL-10-mediated induction. Surprisingly, our data demonstrate that the induction of arginase-1 is not conducive, per se, for MDSC-mediated inhibition of T cell proliferation, as neither supplementation of L-arginine nor arginase-1 inhibitor rescue T cell proliferation. Instead, direct cell contact is required for the inhibition and is not dampened by PD-L1 blockade or SIRPα deficiency.
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Roberts, Andrew W., Ben A. Croker, Warren Alexander, and Donald Metcalf. "Aberrant G-CSF- and IL-6-Induced Proliferation and Differentiation in Suppressor of Cytokine Signaling-3 (SOCS3)-Deficient Progenitor Cells." Blood 104, no. 11 (November 16, 2004): 821. http://dx.doi.org/10.1182/blood.v104.11.821.821.
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Abstract Studies using mice with germline or tissue-specific deletion of SOCS3 indicate that SOCS3 is a negative regulator of IL-6, LIF, Leptin and G-CSF-induced STAT3 phosphorylation. We have investigated the physiological importance of SOCS3 in blood cells by creating conditionally-targeted mice with SOCS3-deficient hematopoiesis (Immunity2004; 20:153–65). These mice develop a fatal inflammatory disease in adulthood characterized by tissue infiltration with neutrophils and macrophages, and display hyper-responsiveness to G-CSF in vitro and in vivo, with the development of pathological myeloid cell-mediated tissue damage. In hepatocytes and macrophages stimulated with IL-6, we and others (Nature Immunol2003; 4:540–5, and 4:546–60) have found that SOCS3 is not only important for modulating the intensity of signalling from cytokine receptors, but also for the maintenance of specificity of the signal. Whether these qualitative changes revealed by microarray experiments have physiological significance remains to be proven. To investigate this issue in a more plastic system, we analyzed the consequences of SOCS3-deficiency in hematopoietic progenitor cells. We hypothesised that if SOCS3 was required to maintain the specificity, as well as intensity of signals arising from cytokine receptors, then changes in differentiation as well as proliferation would be observed. To exclude confounding effects from cytokine production by mature cells, and to minimize potential selection bias within the starting populations, purified SOCS3-deficient lin- kit+ progenitor cells from healthy young mice with SOCS3-deficent hematopoiesis, were compared with both littermate control and C57BL/6 lin- kit+ progenitor cells. Proliferation was monitored in liquid and agar cultures stimulated with SCF, IL-3, GM-CSF, G-CSF and IL-6 alone, or in combination with SCF. At the completion of the experiments ( 1–7 days), proportions of neutrophil, macrophage and precursors were determined by microscopy. The number of divisions progenitors underwent was monitored by clone-mapping experiments in agar and CFSE-labelling in liquid cultures. No differences between SOCS3-deficient and wild-type (WT) cells were observed after stimulation with SCF, IL-3, GM-CSF or combinations of these, suggesting that progenitor cells of each genotype were similar in their developmental potential. However, marked differences were observed for G-CSF and IL-6. G-CSF induced a 2-3-fold increase in cellular output in both liquid and agar cultures, and the distribution of CFSE-intensity was consistent with an additional division occurring over a 4 day timespan in SOCS3-deficient cells. With IL-6 stimulation, while SOCS3-deficient progenitor cells initiated 1.7-fold more colonies, the overall cellular output was no greater than that of WT progenitors. With WT progenitors for both stimuli, the vast majority of clones were neutrophil colonies and >90% of emergent cells at 7 days were neutrophils or their precursors. In contrast, SOCS3-deficient progenitors stimulated with IL-6 generated 43% granulocyte-macrophage colonies and 13% pure macrophage colonies with >70% of emergent cells being macrophages. For G-CSF, a similar, but less pronounced shift towards macrophage development was observed. We conclude that SOCS3 is required to maintain normal cellular proliferative and differentiative responses to G-CSF and IL-6. The precise perturbations in signalling responsible for these aberrations are being defined through microarray and biochemical experiments.
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Salazar-Gonzalez, J. F., D. J. Moody, J. V. Giorgi, O. Martinez-Maza, R. T. Mitsuyasu, and J. L. Fahey. "Reduced ecto-5'-nucleotidase activity and enhanced OKT10 and HLA-DR expression on CD8 (T suppressor/cytotoxic) lymphocytes in the acquired immune deficiency syndrome: evidence of CD8 cell immaturity." Journal of Immunology 135, no. 3 (September 1, 1985): 1778–85. http://dx.doi.org/10.4049/jimmunol.135.3.1778.
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Abstract Markedly reduced ecto-5'-nucleotidase activity was found in peripheral blood lymphocytes from 27 out of 30 homosexual men with the acquired immune deficiency syndrome (AIDS) in association with Kaposi's sarcoma (AIDS-KS; 2.67 +/- 1.70 U/10(6) cells; n = 13), opportunistic infections (AIDS-OI; 9.29 +/- 7.32; n = 7), or the AIDS-related complex (ARC; 9.82 +/- 6.12; n = 10). These values were significantly different from healthy controls (22.70 +/- 4.58; p less than 0.001). In AIDS-KS patients, both T cells and non-T cells exhibited significantly reduced ecto-5'-NT activity (p less than 0.001). AIDS-KS CD8 cells contained 20% of the mean ecto-5'-NT activity (7.04 +/- 3.53) displayed by control CD8 cells (34.07 +/- 4.86; p less than 0.001). No significant difference in enzyme level was observed between control and AIDS-KS CD4 cells (11.93 +/- 4.98 vs 7.98 +/- 3.28, respectively). In AIDS patients, lymphocyte ecto-5'-NT activity was inversely related (r = -0.518; p less than 0.01) to the absolute number of OKT10+ cells, but no correlation was found with the number of HLA-DR+ cells (r =-0.224). Two-color analysis of lymphocytes from AIDS-KS patients revealed that 75 +/- 12% of circulating CD8 cells expressed the OKT10 antigen, whereas only 10 +/- 6% of control CD8 cells did. HLA-DR antigens, which are not normally found on circulating resting T cells, were expressed in AIDS-KS CD8 cells, although to a lesser extent than OKT10. These data demonstrate that most AIDS CD8 cells differ from control CD8 cells. Although it has been suggested that these cells are activated cytotoxic or suppressor cells, the data presented here support the hypothesis they are immature. Reduced T cell ecto-5'-NT activity and enhanced expression of OKT10 and HLA-DR antigens on circulating CD8 cells, in conjunction with lack of transferrin receptor-(OKT9) and IL 2 receptor-(Tac) bearing lymphocytes, sustain this latter hypothesis. The correlation of the numerical reduction of CD4 cells with the reduced levels of ecto-5'-NT (r = 0.606; p less than 0.01) suggests that the abnormal maturation of CD8 cells seen in AIDS might be a consequence of the CD4 deficiency characteristic of this syndrome.
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Rasmussen, Kasper D., Guangshuai Jia, Jens V. Johansen, Olivier A. Bernard, and Kristian Helin. "Loss Of Tet2 Collaborates With AML1-ETO In Inducing Leukemogenesis." Blood 122, no. 21 (November 15, 2013): 1251. http://dx.doi.org/10.1182/blood.v122.21.1251.1251.
Full textAbstract:
Abstract Disruption of the epigenome can contribute to both cancer initiation and tumor progression. The methylcytosine dioxygenase TET2, which is involved in the regulation DNA methylation patterns, is frequently mutated in Acute Myeloid Leukemia (AML) patients and plays a central role in normal hematopoiesis. However, Tet2–deficient mouse models do not readily develop acute leukemia suggesting that additional oncogenic events are necessary for full-blown transformation. The translocation t(8;21)(q22;q22) results in expression of the AML1-ETO (AE) fusion protein and is present in ∼5-10% of AML patients. Interestingly, the combination of t(8;21)(q22;q22) and TET2 mutations, although rare, have been observed in several studies in patients groups covering both pediatric and adult de novo AML. To confirm a direct role of TET2 mutations in the development of t(8;21)(q22;q22)-rearranged leukemias we expressed the AE fusion protein in hematopoietic stem and progenitor cells (HSPC) isolated from WT or Tet2-deficient animals. Serial replating assays in semisolid media as well as growth in liquid culture suggested a cooperative effect of AE and Tet2 deficiency as indicated by a significant increase in proliferation and colony formation efficiency. We then used a bone marrow transplantation model to assess the in vivo leukemic potential of AE;Tet2-/- HSPCs. The majority of recipient mice transplanted with AE;Tet2-/- (n=5/6) HSPCs succumbed to an aggressive and transplantable AML-like condition with a median latency of 5-6 months, whereas mice receiving AE HSPCs remained healthy over a 12 month observation period (n=6/6). The morbid mice showed many signs of disease including loss of weight, anemia, splenomegaly and large number of leukemic blasts (CD45neg) in circulation. Further analysis of the bone marrow and spleen revealed a massive expansion of a linneg (CD3e, Gr-1, Mac-1, B220, Ter119), c-Kitlow-high, CD16/32pos cell population indicating a block of differentiation and expansion of an immature myeloid progenitor population from AE:Tet2-/- HSPCs. To investigate how Tet2-deficiency contributes to transformation of t(8;21)(q22;q22)-rearranged cells we generated and cultured pre-leukemic AE-transduced cells in vitro from Tet2fl/fl HPSCs harboring the inducible Rosa26-CreERT2 allele. Genome-wide analysis of methylation patterns and gene expression showed that induction of Tet2-deficiency in pre-leukemic cells leads to progressive hypermethylation of gene-regulatory elements and altered expression of several genes implicated in tumorigenesis. Thus, this study illustrates how aberrant DNA methylation patterns can contribute to disease and confirm the role of Tet2 as a tumor suppressor preventing leukemogenesis. Disclosures: Helin: EpiTherapeutics : Consultancy, Equity Ownership.
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Kubo, Takashi, Toshiko Aiso, and Reiko Ohki. "Eight UCA codons differentially affect the expression of the lacZ gene in the divE42 mutant of Escherichia coli." Canadian Journal of Microbiology 46, no. 6 (June 1, 2000): 577–83. http://dx.doi.org/10.1139/w00-019.
Full textAbstract:
In the divE mutant, which has a temperature-sensitive mutation in the tRNA1Ser gene, the synthesis of beta-galactosidase is dramatically decreased at the non-permissive temperature. In Escherichia coli, the UCA codon is only recognized by tRNA1Ser. Several genes containing UCA codons are normally expressed at 42°C in the divE mutant. Therefore, it is unlikely that the defect is due to the general translational deficiency of the mutant tRNA1Ser. In this study, we constructed mutant lacZ genes, in which one or several UCA codons at eight positions were replaced with other serine codons such as UCU or UCC, and we examined the expression of these mutant genes in the divE mutant. We found that a single UCA codon at position 6 or 462 was sufficient to cause the same level of reduced beta-galactosidase synthesis as that of the wild-type lacZ gene, and that the defect in beta-galactosidase synthesis was accompanied by a low level of lacZ mRNA. It was also found that introduction of an rne-1 pnp-7 double mutation restored the expression of mutant lacZ genes with only UCA codons at position 6 or 462. A polarity suppressor mutation in the rho gene had no effect on the defect in lacZ gene expression in the divE mutant. We propose a model to explain these results.Key words: divE gene, tRNA1Ser, lacZ gene expression, UCA codon.
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Liu, Yanling, Tong Meng, Tong Wang, Jing Li, Wei Zhang, Biaosheng Lin, Jihui Chen, Zhanxi Lin, and Biaosheng Lin. "Hot water extract from spent mushroom substrate of Ganoderma lucidu m improves immune function in immune deficient mice." Tropical Journal of Pharmaceutical Research 22, no. 1 (February 18, 2023): 143–50. http://dx.doi.org/10.4314/tjpr.v22i1.20.
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Purpose: To investigate the effects of hot water extract of the spent mushroom substrate from Ganoderma lucidum (HWE) on the immune functions of immune-deficient mice in the presence or absence of cyclophosphamide.
Methods: The C57BL/6 mice were treated with varying doses of HWE and cyclophosphamide, and the spleen transcriptome was evaluated by high-throughput sequencing. Pathway enrichment analysis was conducted to generate an overview of differentially-expressed genes (DEGs) functions and interactions.
Results: Significant DEGs were observed among the control (CK), normal control (HWE), model (CY), and high-dose HWE (CY + HWEH) groups (p < 0.05). Compared with CK group, HWE and CY + HWEH groups showed upregulation of genes (Alas2, CCNE1, and CCNA2), whereas genes encoding major histocompatibility complex components, costimulatory factors, proinflammatory chemokines, and inflammatory chemokines were significantly downregulated (p < 0.05). Compared with CY group, multiple tumor suppressor and tumorigenesis genes, such as CDKN1A, CDKN1B, MAPK10, and TN-C, were downregulated in CK, HWE, and CY + HWEH groups.
Conclusion: This study highlights changes in the spleen transcriptomic profiles of C57BL/6 mice treated with HWE and CY, indicating that HWE improves immunomodulation in a mouse model with immune deficiency. Hot water extract of the spent mushroom substrate from Ganoderma lucidum (HWE) has potentials as an immune enhancer in immunocompromised patients.
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Lai, Mingqiang, Wenchong Zou, Zelong Han, Ling Zhou, Zeyou Qiu, Juan Chen, Sheng Zhang, et al. "Tsc1 regulates tight junction independent of mTORC1." Proceedings of the National Academy of Sciences 118, no. 30 (July 23, 2021): e2020891118. http://dx.doi.org/10.1073/pnas.2020891118.
Full textAbstract:
Tuberous sclerosis complex 1 (Tsc1) is a tumor suppressor that functions together with Tsc2 to negatively regulate the mechanistic target of rapamycin complex 1 (mTORC1) activity. Here, we show that Tsc1 has a critical role in the tight junction (TJ) formation of epithelium, independent of its role in Tsc2 and mTORC1 regulation. When an epithelial cell establishes contact with neighboring cells, Tsc1, but not Tsc2, migrates from the cytoplasm to junctional membranes, in which it binds myosin 6 to anchor the perijunctional actin cytoskeleton to β-catenin and ZO-1. In its absence, perijunctional actin cytoskeleton fails to form. In mice, intestine-specific or inducible, whole-body Tsc1 ablation disrupts adherens junction/TJ structures in intestine or skin epithelia, respectively, causing Crohn’s disease–like symptoms in the intestine or psoriasis-like phenotypes on the skin. In patients with Crohn’s disease or psoriasis, junctional Tsc1 levels in epithelial tissues are markedly reduced, concomitant with the TJ structure impairment, suggesting that Tsc1 deficiency may underlie TJ-related diseases. These findings establish an essential role of Tsc1 in the formation of cell junctions and underpin its association with TJ-related human diseases.
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Zhang, Xingli, Yan Wang, Jia Yuan, Ni Li, Siyu Pei, Jing Xu, Xuan Luo, et al. "Macrophage/microglial Ezh2 facilitates autoimmune inflammation through inhibition of Socs3." Journal of Experimental Medicine 215, no. 5 (April 6, 2018): 1365–82. http://dx.doi.org/10.1084/jem.20171417.
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Histone 3 Lys27 (H3K27) trimethyltransferase Ezh2 is implicated in the pathogenesis of autoimmune inflammation. Nevertheless, the role of Ezh2 in macrophage/microglial activation remains to be defined. In this study, we identified that macrophage/microglial H3K27me3 or Ezh2, rather than functioning as a repressor, mediates toll-like receptor (TLR)-induced proinflammatory gene expression, and therefore Ezh2 depletion diminishes macrophage/microglial activation and attenuates the autoimmune inflammation in dextran sulfate sodium–induced colitis and experimental autoimmune encephalomyelitis. Mechanistic characterizations indicated that Ezh2 deficiency directly stimulates suppressor of cytokine signaling 3 (Socs3) expression and therefore enhances the Lys48-linked ubiquitination and degradation of tumor necrosis factor receptor–associated factor 6. As a consequence, TLR-induced MyD88-dependent nuclear factor κB activation and the expression of proinflammatory genes in macrophages/microglia are compromised in the absence of Ezh2. The functional dependence of Ezh2 for Socs3 is further illustrated by the rescue experiments in which silencing of Socs3 restores macrophage activation and rescues autoimmune inflammation in macrophage/microglial Ezh2-deficient mice. Together, these findings establish Ezh2 as a macrophage lineage-specific mediator of autoimmune inflammation and highlight a previously unknown mechanism of Ezh2 function.