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Dąbrowska, Katarzyna, i Magdalena Zielińska. "Silencing of Transcription Factor Sp1 Promotes SN1 Transporter Regulation by Ammonia in Mouse Cortical Astrocytes". International Journal of Molecular Sciences 20, nr 2 (9.01.2019): 234. http://dx.doi.org/10.3390/ijms20020234.
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The involvement of the astrocytic SN1 (SNAT3) transporter in ammonia-induced l-glutamine retention was recently documented in mouse-cultured astrocytes. Here we investigated the involvement of specificity protein 1 (Sp1) transcription factor in SN1 regulation in ammonium chloride (“ammonia”)-treated astrocytes. Sp1 expression and its cellular localization were determined using real-time qPCR, Western blot, and confocal microscopy. Sp1 binding to Snat3 promoter was analyzed by chromatin immunoprecipitation. The role of Sp1 in SN1 expression and SN1-mediated [3H]glutamine uptake in ammonia-treated astrocytes was verified using siRNA and mithramycin A. The involvement of protein kinase C (PKC) isoforms in Sp1 level/phosphorylation status was verified using siRNA technology. Sp1 translocation to the nuclei and its enhanced binding to the Snat3 promoter, along with Sp1 dependence of system N-mediated [3H]glutamine uptake, were observed in astrocytes upon ammonia exposure. Ammonia decreased the level of phosphorylated Sp1, and the effect was reinforced by long-term incubation with PKC modulator, phorbol 12-myristate 13-acetate, which is a treatment likely to dephosphorylate Sp1. Furthermore, silencing of the PKCδ isoform appears to enhance the ammonia effect on the Sp1 level. Collectively, the results demonstrate the regulatory role of Sp1 in regulation of SN1 expression and activity in ammonia-treated astrocytes and implicate altered Sp1 phosphorylation status in this capacity.
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Budiarti, Lia Yulia, Isnaini Isnaini, Puteri Dayana, Norma Sari i Nur Almira R. S. "Antimicrobial Activity of Stenochlaena palustris and Sauropus androgynus in Staphylococcus aureus, Escherichia coli and Candidia albicans". Bioinformatics and Biomedical Research Journal 4, nr 1 (29.11.2021): 32–38. http://dx.doi.org/10.11594/bbrj.04.01.05.
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Stenochlaena palustris and Sauropus androgynus are known to contains antimicrobial substances such as flavonoids, saponins and tannins compounds.The purpose of this study was to analyzes the antimicrobial activity of young and old leaf infusions of S. palustris and S. androgynus leaves against Staphylococcus aureus, Escherichia coli and Candida albicans. Analyze the antibacterial activity of a single preparations with a combination preparation of S.palustris (SP) and S.androgynus (SA) leaves infusion against S.aureus, E.coli and C.albicans. Leaves of S.palustris young part (SP1) taken 0-10 cm from shoots and old parts (SP2) 11-20 cm from shoots, while leaves of S.androgynus young part (SA1) leaves number 1 - 10 from the top and the old part (SA2) leaves number 11-20 from the top. The results showed that a single infusion of SP1 75% and SP2 75%, SA1 90% and SA2 90%, and a combination of SP1 75% and SA1 75%, SP2 75% and SA2 75% have the same activity as ampicillin in S.aureus. Single infusion of SP1 90% and SP2 90%, SA1 90% and SA2 90%, combination of SP1 75% and SA1 80% and the combination of SP2 80% and SA2 60% have the same activity as ciprofloxacin in E. coli. Single infusion of SP1 90% and SP2 90%, and a combination of SP1 80% and SA1 80%, SP2 80% and SA2 80% have the same activity as ketoconazole in C.albicans. The difference in activity due to differences in leaf parts used only occurred in E. coli, whereas in S.aureus and C.albicans (p <0.05).
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Dąbrowska, Katarzyna, Katarzyna Skowrońska, Mariusz Popek, Jan Albrecht i Magdalena Zielińska. "The Role of Nrf2 Transcription Factor and Sp1-Nrf2 Protein Complex in Glutamine Transporter SN1 Regulation in Mouse Cortical Astrocytes Exposed to Ammonia". International Journal of Molecular Sciences 22, nr 20 (18.10.2021): 11233. http://dx.doi.org/10.3390/ijms222011233.
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Ammonia toxicity in the brain primarily affects astrocytes via a mechanism in which oxidative stress (OS), is coupled to the imbalance between glutamatergic and GABAergic transmission. Ammonia also downregulates the astrocytic N system transporter SN1 that controls glutamine supply from astrocytes to neurons for the replenishment of both neurotransmitters. Here, we tested the hypothesis that activation of Nrf2 is the process that links ammonia-induced OS formation in astrocytes to downregulation and inactivation of SN1 and that it may involve the formation of a complex between Nrf2 and Sp1. Treatment of cultured cortical mouse astrocytes with ammonia (5 mM NH4Cl for 24 h) evoked Nrf2 nuclear translocation, increased its activity in a p38 MAPK pathway-dependent manner, and enhanced Nrf2 binding to Slc38a3 promoter. Nrf2 silencing increased SN1 mRNA and protein level without influencing astrocytic [3H]glutamine transport. Ammonia decreased SN1 expression in Nrf2 siRNA treated astrocytes and reduced [3H]glutamine uptake. In addition, while Nrf2 formed a complex with Sp1 in ammonia-treated astrocytes less efficiently than in control cells, treatment of astrocytes with hybrid-mode inactivated Sp1-Nrf2 complex (Nrf2 silencing + pharmacological inhibition of Sp1) did not affect SN1 protein level in ammonia-treated astrocytes. In summary, the results document that SN1 transporter dysregulation by ammonia in astrocytes involves activation of Nrf2 but does not require the formation of the Sp1-Nrf2 complex.
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Rui, Xianliang, Jennivine Tsao, Joshua O. Scheys, Gary D. Hammer i Bernard P. Schimmer. "Contributions of Specificity Protein-1 and Steroidogenic Factor 1 to Adcy4 Expression in Y1 Mouse Adrenal Cells". Endocrinology 149, nr 7 (3.04.2008): 3668–78. http://dx.doi.org/10.1210/en.2008-0203.
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The type 4 adenylyl cyclase, Adcy4, is the least abundant of five different adenylyl cyclase isoforms expressed in the Y1 mouse adrenocortical cell line and is deficient in a Y1 mutant with impaired steroidogenic factor 1 (SF1) activity. This study examines the contributions of SF1 and other DNA promoter/regulatory elements to Adcy4 expression in the Y1 cell line and its derivative Adcy4-deficient mutant. Primer extension and in silico analyses indicate that Adcy4 transcription initiates from multiple sites just downstream of a GC-rich sequence. Luciferase reporter gene assays identify a 124-bp sequence, situated 19 bp upstream of the major transcription start site and highly conserved among several mammalian species, as the major determinant of Adcy4 expression in Y1 cells and as a site of compromised activity in the Adcy4-deficient mutant. EMSAs using competitor nucleotides and specific antibodies indicate that this conserved region contains three specificity protein (Sp)-1/Sp3-binding sites and one SF1-binding site. As determined by site-specific mutagenesis, the 5′-most Sp1/Sp3-site enhances promoter activity, whereas the middle Sp1/Sp3 and SF1 sites each repress Adcy4 promoter activity. In the Adcy4-deficient mutant, mutating the SF1 site restores Adcy4 promoter activity and knocking down SF1 with small interfering RNAs increases Adcy4 expression, confirming the contribution of SF1 to the mutant phenotype. These studies demonstrate roles for Sp1/Sp3 and SF1 in Adcy4 expression in Y1 cells and establish a repressor function for SF1 in certain promoter contexts.
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Barnes, John G. P. "SP1". ACM SIGAda Ada Letters XXVII, nr 3 (17.11.2007): 3. http://dx.doi.org/10.1145/1315607.1315584.
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Sward, Ricky E. "SP1". ACM SIGAda Ada Letters 28, nr 3 (grudzień 2008): 3–4. http://dx.doi.org/10.1145/1454497.1454477.
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POPPLETON, Helen M., i Rajendra RAGHOW. "Transcriptional activation of the minimal human Proα1(I) collagen promoter: obligatory requirement for Sp1". Biochemical Journal 323, nr 1 (1.04.1997): 225–31. http://dx.doi.org/10.1042/bj3230225.
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A construct containing human Proα1(I) collagen gene promoter/enhancer-driven chloramphenicol acetyltransferase (CAT), pCOL-KT, failed to be expressed significantly in Sp1-deficient Schneider Drosophila line 2 (SL2) cells. However, CAT expression was induced 200-fold in SL2 cells co-transfected with pCOL-KT and pPACSp1, an Sp1-expression vector driven by the Drosophila actin 5C promoter. Elimination of the four potential Sp1-binding sites from pCOL-KT (pCOL-KTΔI), by removal of the first intron, did not abrogate Sp1-mediated induction of CAT. Even more significantly, a minimal Proα1(I) collagen promoter (-100 to +117 bp), containing a TATA box (-28 to -25 bp) and one putative Sp1-binding site (-87 to -82 bp), elicited strong Sp1-induced transactivation. Furthermore, mutation of the Sp1 motif in the minimal Proα1(I) collagen promoter-CAT construct abolished Sp1-induced expression of the reporter gene. Purified Sp1 protein bound specifically to DNA fragments of the Proα1(I) minimal promoter encompassing the putative Sp1-binding site; Sp1 binding could be competed out by a double-stranded oligonucleotide containing the wild-type Sp1 sequence, while an oligonucleotide containing a mutated Sp1 site failed to compete. Based on these results, we postulate that Sp1 plays an obligatory role in the transcriptional activation of the human Proα1(I) collagen gene. Additionally, we propose that a bona fide Sp1 motif, located most proximal to the TATA box, is necessary and sufficient for Sp1-mediated activation of the minimal Proα1(I) collagen promoter.
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Zhang, Ying, i Maria L. Dufau. "Repression of the Luteinizing Hormone Receptor Gene Promoter by Cross Talk among EAR3/COUP-TFI, Sp1/Sp3, and TFIIB". Molecular and Cellular Biology 23, nr 19 (1.10.2003): 6958–72. http://dx.doi.org/10.1128/mcb.23.19.6958-6972.2003.
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ABSTRACT Transcription of luteinizing hormone receptor (LHR) gene is activated by Sp1/Sp3 at two Sp1 sites and is repressed by nuclear orphan receptors EAR2 and EAR3 through a direct-repeat (DR) motif. To elucidate the mechanism of the orphan receptor-mediated gene repression, we explored the functional connection between the orphan receptors and Sp1/Sp3 complex, and its impact on the basal transcription machinery. The Sp1(I) site was identified as critical for the repression since its mutation reduced the inhibition by EAR2 and abolished the inhibition by EAR3. Cotransfection analyses in SL2 cells showed that both Sp1 and Sp3 were required for this process since EAR3 displayed a complete Sp1/Sp3-dependent inhibitory effect. Functional cooperation between Sp1 and DR domains was further supported by mutual recruitment of EAR3 and Sp1/Sp3 bound to their cognate sites. Deletion of EAR3 N-terminal and DNA-binding domains that reduced its interaction with Sp1 impaired its inhibitory effect on human LHR (hLHR) gene transcription. Furthermore, we demonstrate interaction of TFIIB with Sp1/Sp3 at the Sp1(I) site besides its association with EAR3 and the TATA-less core promoter region. Such interaction relied on Sp1 site-bound Sp1/Sp3 complex and adaptor protein(s) present in the JAR nuclear extracts. We further demonstrated that EAR3 specifically decreased association of TFIIB to the Sp1(I) site without interfering on its interaction with the hLHR core promoter. The C-terminal region of EAR3, which did not participate in its interaction with Sp1, was required for its inhibitory function and may affect the association of TFIIB with Sp1. Moreover, perturbation of the association of TFIIB with Sp1 by EAR3 was reflected in the reduced recruitment of RNA polymerase II to the promoter. Overexpression of TFIIB counteracted the inhibitory effect of EAR3 and activated hLHR gene transcription in an Sp1 site-dependent manner. These findings therefore indicate that TFIIB is a key component in the regulatory control of EAR3 and Sp1/Sp3 on the initiation complex. Such cross talk among EAR3, TFIIB, and Sp1/Sp3 reveals repression of hLHR gene transcription by nuclear orphan receptors is achieved via perturbation of communication between Sp1/Sp3 at the Sp1-1 site and the basal transcription initiator complex.
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NICOLÁS, Marta, Vèronique NOÉ i Carlos J. CIUDAD. "Transcriptional regulation of the human Sp1 gene promoter by the specificity protein (Sp) family members nuclear factor Y (NF-Y) and E2F". Biochemical Journal 371, nr 2 (15.04.2003): 265–75. http://dx.doi.org/10.1042/bj20021166.
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We analysed in detail the minimal promoter of transcription factor Sp1, which extends 217bp from the initiation of transcription. Within this sequence we identified putative binding sites for Sp1, nuclear factor Y (NF-Y), activator protein 2 ('AP-2'), CCAAT/enhancer-binding protein ('C/EBP') and E2F transcription factors. In one case, the boxes for Sp1 and NF-Y are overlapping. Gel-shift and supershift assays demonstrated specific binding of Sp1, Sp3 and NF-Y proteins. Transient transfections and luciferase assays revealed activation of the Sp1 minimal promoter upon overexpression of Sp1 itself, NF-Y and E2F. Whereas overexpression of NF-Y or E2F had an additive effect on Sp1 overexpression, the activation of Sp1 transcription due to Sp1 was counteracted by Sp3 overexpression. Mutagenesis analysis of the NFY/Sp1-overlapping box revealed that both factors compete for this box, and that when the NF-Y site of this overlapping box is specifically mutated there is an increase in Sp1 binding, thus increasing transcriptional activity. These results help to explain the complex regulation of the Sp1 gene, which depends on the relative amounts of Sp1, Sp3, E2F and NF-Y proteins in the cell.
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Chen, L. I., T. Nishinaka, K. Kwan, I. Kitabayashi, K. Yokoyama, Y. H. Fu, S. Grünwald i R. Chiu. "The retinoblastoma gene product RB stimulates Sp1-mediated transcription by liberating Sp1 from a negative regulator". Molecular and Cellular Biology 14, nr 7 (lipiec 1994): 4380–89. http://dx.doi.org/10.1128/mcb.14.7.4380-4389.1994.
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Studies have demonstrated that the retinoblastoma susceptibility gene product, RB, can either positively or negatively regulate expression of several genes through cis-acting elements in a cell-type-dependent manner. The nucleotide sequence of the retinoblastoma control element (RCE) motif, GCCACC or CCACCC, and the Sp1 consensus binding sequence, CCGCCC, can confer equal responsiveness to RB. Here, we report that RB activates transcription of the c-jun gene through the Sp1-binding site within the c-jun promoter. Preincubation of crude nuclear extracts with monoclonal antibodies to RB results in reduction of Sp1 complexes in a mobility shift assay, while addition of recombinant RB in mobility shift assay mixtures with CCL64 cell extracts leads to an enhancement of DNA-binding activity of SP1. These results suggest that RB is directly or indirectly involved in Sp1-DNA binding activity. A mechanism by which RB regulates transactivation is indicated by our detection of a heat-labile and protease-sensitive Sp1 negative regulator(s) (Sp1-I) that specifically inhibits Sp1 binding to a c-jun Sp1 site. This inhibition is reversed by addition of recombinant RB proteins, suggesting that RB stimulates Sp1-mediated transactivation by liberating Sp1 from Sp1-I. Additional evidence for Sp1-I involvement in Sp1-mediated transactivation was demonstrated by cotransfection of RB, GAL4-Sp1, and a GAL4-responsive template into CV-1 cells. Finally, we have identified Sp1-I, a approximately 20-kDa protein(s) that inhibits the Sp1 complexes from binding to DNA and that is also an RB-associated protein. These findings provide evidence for a functional link between two distinct classes of oncoproteins, RB and c-Jun, that are involved in the control of cell growth, and also define a novel mechanism for the regulation of c-jun expression.
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Chen, L. I., T. Nishinaka, K. Kwan, I. Kitabayashi, K. Yokoyama, Y. H. Fu, S. Grünwald i R. Chiu. "The retinoblastoma gene product RB stimulates Sp1-mediated transcription by liberating Sp1 from a negative regulator." Molecular and Cellular Biology 14, nr 7 (lipiec 1994): 4380–89. http://dx.doi.org/10.1128/mcb.14.7.4380.
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Studies have demonstrated that the retinoblastoma susceptibility gene product, RB, can either positively or negatively regulate expression of several genes through cis-acting elements in a cell-type-dependent manner. The nucleotide sequence of the retinoblastoma control element (RCE) motif, GCCACC or CCACCC, and the Sp1 consensus binding sequence, CCGCCC, can confer equal responsiveness to RB. Here, we report that RB activates transcription of the c-jun gene through the Sp1-binding site within the c-jun promoter. Preincubation of crude nuclear extracts with monoclonal antibodies to RB results in reduction of Sp1 complexes in a mobility shift assay, while addition of recombinant RB in mobility shift assay mixtures with CCL64 cell extracts leads to an enhancement of DNA-binding activity of SP1. These results suggest that RB is directly or indirectly involved in Sp1-DNA binding activity. A mechanism by which RB regulates transactivation is indicated by our detection of a heat-labile and protease-sensitive Sp1 negative regulator(s) (Sp1-I) that specifically inhibits Sp1 binding to a c-jun Sp1 site. This inhibition is reversed by addition of recombinant RB proteins, suggesting that RB stimulates Sp1-mediated transactivation by liberating Sp1 from Sp1-I. Additional evidence for Sp1-I involvement in Sp1-mediated transactivation was demonstrated by cotransfection of RB, GAL4-Sp1, and a GAL4-responsive template into CV-1 cells. Finally, we have identified Sp1-I, a approximately 20-kDa protein(s) that inhibits the Sp1 complexes from binding to DNA and that is also an RB-associated protein. These findings provide evidence for a functional link between two distinct classes of oncoproteins, RB and c-Jun, that are involved in the control of cell growth, and also define a novel mechanism for the regulation of c-jun expression.
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Chu, Shijian, i Thomas J. Ferro. "Identification of a hydrogen peroxide-induced PP1-JNK1-Sp1 signaling pathway for gene regulation". American Journal of Physiology-Lung Cellular and Molecular Physiology 291, nr 5 (listopad 2006): L983—L992. http://dx.doi.org/10.1152/ajplung.00454.2005.
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Oxidative stress often results in changes in gene expression through the regulation of transcription factors. In this study, we examine how Sp1 phosphorylation is regulated by H2O2 in a human alveolar epithelial cell line (HAE). Treatment of HAE cells with H2O2 increases phosphorylation of Sp1 and activates JNK. To establish a relationship between JNK and Sp1, we show that JNK activator anisomycin increases Sp1 phosphorylation, and JNK inhibitors as well as dominant-negative JNK1 attenuate H2O2-induced Sp1 phosphorylation. Additionally, JNK1 directly phosphorylates Sp1 in vitro, reducing Sp1 binding to DNA. These results demonstrate the role of JNK in H2O2-induced Sp1 phosphorylation. Because H2O2 inhibits Ser/Thr protein phosphatase-1 (PP1), we examined the role of PP1 in the regulation of JNK. Similar to H2O2, inhibition of PP1 induces phosphorylation of Sp1 and activation of JNK in HAE cells. Inhibition of JNK activity using either inhibitors or dominant-negative mutant JNK1 suppresses PP1 inhibition-induced Sp1 phosphorylation. Furthermore, PP1 directly inactivates JNK1 in vitro. These data suggest that 1) H2O2 increases the phosphorylation level of Sp1, 2) Sp1 is a target of the JNK pathway, 3) PP1 regulates JNK activation, and 4) the “PP1-JNK” pathway plays a role in H2O2-induced Sp1 phosphorylation in lung epithelial cells.
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Peng, Feixia, Ying Zhou, Juan Wang, Baoqiang Guo, Yun Wei, Huan Deng, Zihui Wu i in. "The transcription factor Sp1 modulates RNA polymerase III gene transcription by controlling BRF1 and GTF3C2 expression in human cells". Journal of Biological Chemistry 295, nr 14 (1.03.2020): 4617–30. http://dx.doi.org/10.1074/jbc.ra119.011555.
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Specificity protein 1 (Sp1) is an important transcription factor implicated in numerous cellular processes. However, whether Sp1 is involved in the regulation of RNA polymerase III (Pol III)-directed gene transcription in human cells remains unknown. Here, we first show that filamin A (FLNA) represses Sp1 expression as well as expression of TFIIB-related factor 1 (BRF1) and general transcription factor III C subunit 2 (GTF3C2) in HeLa, 293T, and SaOS2 cell lines stably expressing FLNA-silencing shRNAs. Both BRF1 promoter 4 (BRF1P4) and GTF3C2 promoter 2 (GTF3C2P2) contain putative Sp1-binding sites, suggesting that Sp1 affects Pol III gene transcription by regulating BRF1 and GTF3C2 expression. We demonstrate that Sp1 knockdown inhibits Pol III gene transcription, BRF1 and GTF3C2 expression, and the proliferation of 293T and HeLa cells, whereas Sp1 overexpression enhances these activities. We obtained a comparable result in a cell line in which both FLNA and Sp1 were depleted. These results indicate that Sp1 is involved in the regulation of Pol III gene transcription independently of FLNA expression. Reporter gene assays showed that alteration of Sp1 expression affects BRF1P4 and GTF3C2P2 activation, suggesting that Sp1 modulates Pol III–mediated gene transcription by controlling BRF1 and GTF3C2 gene expression. Further analysis revealed that Sp1 interacts with and thereby promotes the occupancies of TATA box–binding protein, TFIIAα, and p300 at both BRF1P4 and GTF3C2P2. These findings indicate that Sp1 controls Pol III–directed transcription and shed light on how Sp1 regulates cancer cell proliferation.
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Doetzlhofer, Angelika, Hans Rotheneder, Gerda Lagger, Manfred Koranda, Vladislav Kurtev, Gerald Brosch, Erhard Wintersberger i Christian Seiser. "Histone Deacetylase 1 Can Repress Transcription by Binding to Sp1". Molecular and Cellular Biology 19, nr 8 (1.08.1999): 5504–11. http://dx.doi.org/10.1128/mcb.19.8.5504.
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ABSTRACT The members of the Sp1 transcription factor family can act as both negative and positive regulators of gene expression. Here we show that Sp1 can be a target for histone deacetylase 1 (HDAC1)-mediated transcriptional repression. The histone deacetylase inhibitor trichostatin A activates the chromosomally integrated murine thymidine kinase promoter in an Sp1-dependent manner. Coimmunoprecipitation experiments with Swiss 3T3 fibroblasts and 293 cells demonstrate that Sp1 and HDAC1 can be part of the same complex. The interaction between Sp1 and HDAC1 is direct and requires the carboxy-terminal domain of Sp1. Previously we have shown that the C terminus of Sp1 is necessary for the interaction with the transcription factor E2F1 (J. Karlseder, H. Rotheneder, and E. Wintersberger, Mol. Cell. Biol. 16:1659–1667, 1996). Coexpression of E2F1 interferes with HDAC1 binding to Sp1 and abolishes Sp1-mediated transcriptional repression. Our results indicate that one component of Sp1-dependent gene regulation involves competition between the transcriptional repressor HDAC1 and the transactivating factor E2F1.
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Shen, Licong, Xiaxia Hong, Yang Liu, Wenjun Zhou i Yi Zhang. "The miR-25-3p/Sp1 pathway is dysregulated in ovarian endometriosis". Journal of International Medical Research 48, nr 4 (kwiecień 2020): 030006052091843. http://dx.doi.org/10.1177/0300060520918437.
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Objective The transcription factor Specificity protein 1 (Sp1) plays important roles in many critical biological functions; however, its expression and underlying functions in endometriosis remain undefined. Bioinformatics has suggested that Sp1 is potentially regulated by miR-25-3p. This study investigated Sp1 and miR-25-3p expression and their interaction during the pathogenesis of endometriosis. Methods Fifteen women with American Fertility Society stage III/IV ovarian endometriosis and 14 disease-free controls were included. Sp1 expression was detected by qPCR, immunohistochemistry, and western blotting. Using both bioinformatics and genetics, we identified that Sp1 was a potential target of miR-25-3p. Then, the relationship between miR-25-3p and Sp1 was investigated by knockdown and overexpression experiments. Results Sp1 mRNA and protein levels were increased in ectopic and eutopic endometrium compared with normal endometrium samples, with the highest expression in ectopic endometrium samples. In vitro experiments and luciferase reporter assays demonstrated that Sp1 was upregulated when miR-25-3p was depleted and that Sp1 was a direct target of miR-25-3p, respectively. Conclusions Our study revealed increased Sp1 expression in ovarian endometriosis and subsequently demonstrated that miR-25-3p directly targets Sp1. This suggests a novel miRNA/Sp1 pathway in the pathogenesis of endometriosis, which should be further explored for other potential therapeutic targets.
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Chuang, Jian-Ying, Yi-Ting Wang, Shiu-Hwa Yeh, Yi-Wen Liu, Wen-Chang Chang i Jan-Jong Hung. "Phosphorylation by c-Jun NH2-terminal Kinase 1 Regulates the Stability of Transcription Factor Sp1 during Mitosis". Molecular Biology of the Cell 19, nr 3 (marzec 2008): 1139–51. http://dx.doi.org/10.1091/mbc.e07-09-0881.
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The transcription factor Sp1 is ubiquitously expressed in different cells and thereby regulates the expression of genes involved in many cellular processes. This study reveals that Sp1 was phosphorylated during the mitotic stage in three epithelial tumor cell lines and one glioma cell line. By using different kinase inhibitors, we found that during mitosis in HeLa cells, the c-Jun NH2-terminal kinase (JNK) 1 was activated that was then required for the phosphorylation of Sp1. In addition, blockade of the Sp1 phosphorylation via inhibition JNK1 activity in mitosis resulted in the ubiquitination and degradation of Sp1. JNK1 phosphorylated Sp1 at Thr278/739. The Sp1 mutated at Thr278/739 was unstable during mitosis, possessing less transcriptional activity for the 12(S)-lipoxygenase expression and exhibiting a decreased cell growth rate compared with wild-type Sp1 in HeLa cells. In N-methyl-N-nitrosourea–induced mammary tumors, JNK1 activation provided a potential relevance with the accumulation of Sp1. Together, our results indicate that JNK1 activation is necessary to phosphorylate Sp1 and to shield Sp1 from the ubiquitin-dependent degradation pathway during mitosis in tumor cell lines.
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Saffer, J. D., S. P. Jackson i M. B. Annarella. "Developmental expression of Sp1 in the mouse". Molecular and Cellular Biology 11, nr 4 (kwiecień 1991): 2189–99. http://dx.doi.org/10.1128/mcb.11.4.2189-2199.1991.
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The expression of the trans-acting transcription factor Sp1 in mice was defined by a combination of RNA analysis and immunohistochemical localization of the Sp1 protein. Although ubiquitously expressed, there was an unexpected difference of at least 100-fold in the amount of Sp1 message in different cell types. Sp1 protein levels showed corresponding marked differences. Substantial variations in Sp1 expression were also found in some cell types at different stages of development. Sp1 levels appeared to be highest in developing hematopoietic cells, fetal cells, and spermatids, suggesting that an elevated Sp1 level is associated with the differentiation process. These results indicate that Sp1 has a regulatory function in addition to its general role in the transcription of housekeeping genes.
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Saffer, J. D., S. P. Jackson i M. B. Annarella. "Developmental expression of Sp1 in the mouse." Molecular and Cellular Biology 11, nr 4 (kwiecień 1991): 2189–99. http://dx.doi.org/10.1128/mcb.11.4.2189.
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The expression of the trans-acting transcription factor Sp1 in mice was defined by a combination of RNA analysis and immunohistochemical localization of the Sp1 protein. Although ubiquitously expressed, there was an unexpected difference of at least 100-fold in the amount of Sp1 message in different cell types. Sp1 protein levels showed corresponding marked differences. Substantial variations in Sp1 expression were also found in some cell types at different stages of development. Sp1 levels appeared to be highest in developing hematopoietic cells, fetal cells, and spermatids, suggesting that an elevated Sp1 level is associated with the differentiation process. These results indicate that Sp1 has a regulatory function in addition to its general role in the transcription of housekeeping genes.
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Chung, Sun-Ku, Joo-Young Kim, Joong-Yeon Lim, Young Mi Park, Ha-Young Hwang, Jae-Hwan Nam i Sang Ick Park. "Transcription Factor Sp1 Is Involved in Expressional Regulation of Coxsackie and Adenovirus Receptor in Cancer Cells". Journal of Biomedicine and Biotechnology 2011 (2011): 1–9. http://dx.doi.org/10.1155/2011/636497.
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Coxsackie and adenovirus receptor (CAR) was first known as a virus receptor. Recently, it is also known to have tumor suppressive activity such as inhibition of cell proliferation, migration, and invasion. It is important to understand how CAR expression can be regulated in cancers. Based on an existence of putative Sp1 binding site within CAR promoter, we investigated whether indeed Sp1 is involved in the regulation of CAR expression. We observed that deletion or mutation of Sp1 binding motif (−503/−498) prominently impaired the Sp1 binding affinity and activity of CAR promoter. Histone deacetylase inhibitor (TSA) treatment enhanced recruitment of Sp1 to the CAR promoter in ChIP assay. Meanwhile, Sp1 binding inhibitor suppressed the recruitment. Exogenous expression of wild-type Sp1 increased CAR expression in CAR-negative cells; meanwhile, dominant negative Sp1 decreased the CAR expression in CAR-positive cells. These results indicate that Sp1 is involved in regulation of CAR expression.
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Gumucio, DL, KL Rood, KL Blanchard-McQuate, TA Gray, A. Saulino i FS Collins. "Interaction of Sp1 with the human gamma globin promoter: binding and transactivation of normal and mutant promoters". Blood 78, nr 7 (1.10.1991): 1853–63. http://dx.doi.org/10.1182/blood.v78.7.1853.1853.
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Abstract We have analyzed the binding of Sp1, a ubiquitously expressed transactivator, to the promoter region of the gamma genes. Low-affinity Sp1 sites were found at -50 and -200. A high-affinity site was detected at -140, over the CACCC sequence. To analyze the function of these sites, Drosophila SL-2 cells, which lack Sp1, were cotransfected with an Sp1 expression plasmid and gamma globin promoter-CAT constructs. In these assays, the gamma promoter was significantly stronger in the presence than in the absence of Sp1. Thus, the three Sp1 sites in the gamma promoter allow binding as well as transactivation of the promoter. The majority of this transactivation was due to the strong binding site at -140 because introduction of a point mutation at -144 (CACCC----AACCC) reduced Sp1-dependent promoter strength by 57%. Analysis of the -200 region suggested that in the wild-type promoter, Sp1 binding at this site contributes little to promoter strength. However, a point mutation (-198 T----C) associated with hereditary persistence of fetal hemoglobin (HPFH) dramatically increased the affinity of this site for Sp1 and significantly increased Sp1 dependent promoter strength in SL-2 cells. Three other point mutations associated with HPFH did not significantly affect the interaction of Sp1 with the - 200 region.
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Gumucio, DL, KL Rood, KL Blanchard-McQuate, TA Gray, A. Saulino i FS Collins. "Interaction of Sp1 with the human gamma globin promoter: binding and transactivation of normal and mutant promoters". Blood 78, nr 7 (1.10.1991): 1853–63. http://dx.doi.org/10.1182/blood.v78.7.1853.bloodjournal7871853.
Pełny tekst źródłaStreszczenie:
We have analyzed the binding of Sp1, a ubiquitously expressed transactivator, to the promoter region of the gamma genes. Low-affinity Sp1 sites were found at -50 and -200. A high-affinity site was detected at -140, over the CACCC sequence. To analyze the function of these sites, Drosophila SL-2 cells, which lack Sp1, were cotransfected with an Sp1 expression plasmid and gamma globin promoter-CAT constructs. In these assays, the gamma promoter was significantly stronger in the presence than in the absence of Sp1. Thus, the three Sp1 sites in the gamma promoter allow binding as well as transactivation of the promoter. The majority of this transactivation was due to the strong binding site at -140 because introduction of a point mutation at -144 (CACCC----AACCC) reduced Sp1-dependent promoter strength by 57%. Analysis of the -200 region suggested that in the wild-type promoter, Sp1 binding at this site contributes little to promoter strength. However, a point mutation (-198 T----C) associated with hereditary persistence of fetal hemoglobin (HPFH) dramatically increased the affinity of this site for Sp1 and significantly increased Sp1 dependent promoter strength in SL-2 cells. Three other point mutations associated with HPFH did not significantly affect the interaction of Sp1 with the - 200 region.
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Han, Deok-Soo, i Eun-Ok Lee. "Sp1 Plays a Key Role in Vasculogenic Mimicry of Human Prostate Cancer Cells". International Journal of Molecular Sciences 23, nr 3 (25.01.2022): 1321. http://dx.doi.org/10.3390/ijms23031321.
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Sp1 transcription factor regulates genes involved in various phenomena of tumor progression. Vasculogenic mimicry (VM) is the alternative neovascularization by aggressive tumor cells. However, there is no evidence of the relationship between Sp1 and VM. This study investigated whether and how Sp1 plays a crucial role in the process of VM in human prostate cancer (PCa) cell lines, PC-3 and DU145. A cell viability assay and three-dimensional culture VM tube formation assay were performed. Protein and mRNA expression levels were detected by Western blot and reverse transcriptase-polymerase chain reaction, respectively. The nuclear twist expression was observed by immunofluorescence assay. A co-immunoprecipitation assay was performed. Mithramycin A (MiA) and Sp1 siRNA significantly decreased serum-induced VM, whereas Sp1 overexpression caused a significant induction of VM. Serum-upregulated vascular endothelial cadherin (VE-cadherin) protein and mRNA expression levels were decreased after MiA treatment or Sp1 silencing. The protein expression and the nuclear localization of twist were increased by serum, which was effectively inhibited after MiA treatment or Sp1 silencing. The interaction between Sp1 and twist was reduced by MiA. On the contrary, Sp1 overexpression enhanced VE-cadherin and twist expressions. Serum phosphorylated AKT and raised matrix metalloproteinase-2 (MMP-2) and laminin subunit 5 gamma-2 (LAMC2) expressions. MiA or Sp1 silencing impaired these effects. However, Sp1 overexpression upregulated phosphor-AKT, MMP-2 and LAMC2 expressions. Serum-upregulated Sp1 was significantly reduced by an AKT inhibitor, wortmannin. These results demonstrate that Sp1 mediates VM formation through interacting with the twist/VE-cadherin/AKT pathway in human PCa cells.
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Samson, SL, i NC Wong. "Role of Sp1 in insulin regulation of gene expression". Journal of Molecular Endocrinology 29, nr 3 (1.12.2002): 265–79. http://dx.doi.org/10.1677/jme.0.0290265.
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Sp1 is a ubiquitous nuclear factor that plays a key role in maintaining basal transcription of 'house-keeping' genes. However, recent evidence points to a more important function for Sp1 in mediating 'cross-talk' between selected signaling cascades to regulate the target genes that respond to these pathways. The role of Sp1 in mediating the actions of the peptide hormone insulin is of specific interest and serves as a model for detailing effects of intracellular signaling on Sp1 activity. This review summarizes studies suggesting that changes in Sp1 phosphorylation provide one potential mechanism for manipulating activity of this protein. A growing body of evidence reveals that the DNA binding and transcription activity of Sp1 may increase or decrease in response to changes in phosphorylation. This enables 'fine-tuning' of Sp1 activity for regulation of gene transcription. Several mechanisms exist by which Sp1 alters gene activity in response to insulin. These include independent Sp1 activity as well as collaboration or competition with others factors. This review points to an ever-increasing role for Sp1 in regulating the transcription of genes in response to extracellular signals such as insulin.
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Hirano, Fuminori, Hirotoshi Tanaka, Yoshiko Hirano, Masaki Hiramoto, Hiroshi Handa, Isao Makino i Claus Scheidereit. "Functional Interference of Sp1 and NF-κB through the Same DNA Binding Site". Molecular and Cellular Biology 18, nr 3 (1.03.1998): 1266–74. http://dx.doi.org/10.1128/mcb.18.3.1266.
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ABSTRACT Gene activation by NF-κB/Rel transcription factors is modulated by synergistic or antagonistic interactions with other promoter-bound transcription factors. For example, Sp1 sites are often found in NF-κB-regulated genes, and Sp1 can activate certain promoters in synergism with NF-κB through nonoverlapping binding sites. Here we report that Sp1 acts directly through a subset of NF-κB binding sites. The DNA binding affinity of Sp1 to these NF-κB sites, as determined by their relative dissociation constants and their relative efficiencies as competitor DNAs or as binding site probes, is in the order of that for a consensus GC box Sp1 site. In contrast, NF-κB does not bind to a GC box Sp1 site. Sp1 can activate transcription through immunoglobulin kappa-chain enhancer or P-selectin promoter NF-κB sites. p50 homodimers replace Sp1 from the P-selectin promoter by binding site competition and thereby either inhibit basal Sp1-driven expression or, in concert with Bcl-3, stimulate expression. The interaction of Sp1 with NF-κB sites thus provides a means to keep an elevated basal expression of NF-κB-dependent genes in the absence of activated nuclear NF-κB/Rel.
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Luo, Jingyan, Xiaoxiao Wang, Zhibo Xia, Lixuan Yang, Zhiming Ding, Shiyuan Chen, Bingquan Lai i Nu Zhang. "Transcriptional factor specificity protein 1 (SP1) promotes the proliferation of glioma cells by up-regulating midkine (MDK)". Molecular Biology of the Cell 26, nr 3 (luty 2015): 430–39. http://dx.doi.org/10.1091/mbc.e14-10-1443.
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Midkine (MDK) expression is associated with the proliferation of many cancers, including glioma. However, the upstream signaling that leads to MDK accumulation remains elusive. This study investigates the molecular mechanism that induces MDK overexpression in human glioma. The Repository for Molecular Brain Neoplasia Data was analyzed to identify potential MDK regulators. Expression of MDK and specificity protein 1 (SP1) was compared in glioma specimens. Chromatin immunoprecipitation assay was used to confirm the transcriptional regulation. MDK-force–expressed, SP1-silenced glioma cells were used to test rescue effects in vitro and in vivo. MDK and SP1 expression in gliomas was significantly higher than in adjacent tissues and was positively correlated in glioma clinical samples and cell lines. The promoter of the human MDK gene has a putative SP1 binding site. SP1 binds to the promoter of the MDK gene and directly regulates MDK expression. MDK or SP1 gene silencing inhibited the proliferation of glioma cells and reduced the tumor volume in nude mice. Overexpression of MDK in SP1-silenced cells could partially rescue the SP1 inhibition effects in vivo and in vitro. SP1 directly up-regulated the expression of MDK, and the SP1-MDK axis cooperated in glioma tumorigenesis.
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Kim, Dool-Bboon, i Neal A. DeLuca. "Phosphorylation of Transcription Factor Sp1 during Herpes Simplex Virus Type 1 Infection". Journal of Virology 76, nr 13 (1.07.2002): 6473–79. http://dx.doi.org/10.1128/jvi.76.13.6473-6479.2002.
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ABSTRACT The expression of most herpes simplex virus type 1 (HSV-1) immediate-early (IE) and early (E) genes decreases late in productive infection. IE and E promoters contain various binding sites for cellular activators, including sites for Sp1, upstream of the TATA box, while late gene promoters generally lack such sites. To address the possibility that Sp1 function may be altered during the course of infection, the modification state and activity of Sp1 were investigated as a function of infection. Sp1 was quantitatively phosphorylated in HSV-1-infected cells without a significant change in abundance. The kinetics of accumulation of phosphorylated Sp1 immediately preceded the decline in E gene (thymidine kinase gene [tk]) mRNA abundance. Phosphorylation of Sp1 required ICP4; however, the proportion of phosphorylated Sp1 was reduced during infection in the presence of phosphonoacetic acid or in the absence of ICP27. While the DNA binding activity of Sp1 was not greatly affected by phosphorylation, the ability of phosphorylated Sp1 isolated from HSV-infected cells to activate transcription in vitro was decreased. These studies suggest that modification of Sp1 may contribute to the decrease of IE and E gene expression late in infection.
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Saha, Arindam, Charles E. Hammond, Monika Gooz i Adam J. Smolka. "The role of Sp1 in IL-1β andH. pylori-mediated regulation of H,K-ATPase gene transcription". American Journal of Physiology-Gastrointestinal and Liver Physiology 295, nr 5 (listopad 2008): G977—G986. http://dx.doi.org/10.1152/ajpgi.90338.2008.
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Helicobacter pylori infection of the gastric body induces transient hypochlorhydria and contributes to mucosal progression toward gastric carcinoma. Acid secretion is mediated by parietal cell H,K-ATPase, in which the catalytic α-subunit (HKα) promoter activity in transfected gastric epithelial [gastric adenocarcinoma (AGS)] cells is repressed by H. pylori through NF-κB p50 homodimer binding to the promoter. IL-1β, an acid secretory inhibitor whose mucosal level is increased by H. pylori, upregulates HKα promoter activity in AGS cells. Because IL-1β also activates NF-κB signaling, we investigated disparate HKα regulation by H. pylori and IL-1β, testing the hypothesis that IL-1β-induced HKα promoter activation is mediated by the transcription factor Sp1. DNase I footprinting revealed Sp1 binding to the HKα promoter at −56 to −39 bp. IL-1β stimulated the activity of three HKα promoter constructs containing NF-κB and Sp1 sites transfected into AGS cells and also stimulated a construct containing only an Sp1 site. This stimulation was abrogated by mutating the HKα promoter Sp1 binding site. Gelshift assays showed that IL-1β increased Sp1 but not p50 binding to cognate HKα probes and that Sp1 also interacts with an HKα NF-κB site when bound to its cognate HKα cis-response element. H. pylori did not augment Sp1 binding to an HKα Sp1 probe, and small interfering RNA-mediated knockdown of Sp1 expression abrogated IL-1β-induced HKα promoter stimulation. We conclude that IL-1β upregulates HKα gene transcription by inducing Sp1 binding to HKα Sp1 and NF-κB sites and that the H. pylori perturbation of HKα gene expression is independent of Sp1-mediated basal HKα transcription.
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Vallian, Sadeq, Khew-Voon Chin i Kun-Sang Chang. "The Promyelocytic Leukemia Protein Interacts with Sp1 and Inhibits Its Transactivation of the Epidermal Growth Factor Receptor Promoter". Molecular and Cellular Biology 18, nr 12 (1.12.1998): 7147–56. http://dx.doi.org/10.1128/mcb.18.12.7147.
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ABSTRACT The promyelocytic leukemia protein (PML) is a nuclear phosphoprotein with growth- and transformation-suppressing ability. Having previously shown it to be a transcriptional repressor of the epidermal growth factor receptor (EGFR) gene promoter, we have now shown that PML’s repression of EGFR transcription is caused by inhibition of EGFR’s Sp1-dependent activity. On functional analysis, the repressive effect of PML was mapped to a 150-bp element (the sequences between −150 and −16, relative to the ATG initiation site) of the promoter. Transient transfection assays with Sp1-negativeDrosophila melanogaster SL2 cells showed that the transcription of this region was regulated by Sp1 and that the Sp1-dependent activity of the promoter was suppressed by PML in a dose-dependent manner. Coimmunoprecipitation and mammalian two-hybrid assays demonstrated that PML and Sp1 were associated in vivo. In vitro binding by means of the glutathione S-transferase (GST) pull-down assay, using the full-length and truncated GST-Sp1 proteins and in vitro-translated PML, showed that PML and Sp1 directly interacted and that the C-terminal (DNA-binding) region of Sp1 and the coiled-coil (dimerization) domain of PML were essential for this interaction. Analysis of the effects of PML on Sp1 DNA binding by electrophoretic mobility shift assay (EMSA) showed that PML could specifically disrupt the binding of Sp1 to DNA. Furthermore, cotransfection of PML specifically repressed Sp1, but not the E2F1-mediated activity of the dihydrofolate reductase promoter. Together, these data suggest that the association of PML and Sp1 represents a novel mechanism for negative regulation of EGFR and other Sp1 target promoters.
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Iwahori, Satoko, Noriko Shirata, Yasushi Kawaguchi, Sandra K. Weller, Yoshitaka Sato, Ayumi Kudoh, Sanae Nakayama, Hiroki Isomura i Tatsuya Tsurumi. "Enhanced Phosphorylation of Transcription Factor Sp1 in Response to Herpes Simplex Virus Type 1 Infection Is Dependent on the Ataxia Telangiectasia-Mutated Protein". Journal of Virology 81, nr 18 (3.07.2007): 9653–64. http://dx.doi.org/10.1128/jvi.00568-07.
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ABSTRACT The ataxia telangiectasia-mutated (ATM) protein, a member of the related phosphatidylinositol 3-like kinase family encoded by a gene responsible for the human genetic disorder ataxia telangiectasia, regulates cellular responses to DNA damage and viral infection. It has been previously reported that herpes simplex virus type 1 (HSV-1) infection induces activation of protein kinase activity of ATM and hyperphosphorylation of transcription factor, Sp1. We show that ATM is intimately involved in Sp1 hyperphosphorylation during HSV-1 infection rather than individual HSV-1-encoded protein kinases. In ATM-deficient cells or cells silenced for ATM expression by short hairpin RNA targeting, hyperphosphorylation of Sp1 was prevented even as HSV-1 infection progressed. Mutational analysis of putative ATM phosphorylation sites on Sp1 and immunoblot analysis with phosphopeptide-specific Sp1 antibodies clarified that at least Ser-56 and Ser-101 residues on Sp1 became phosphorylated upon HSV-1 infection. Serine-to-alanine mutations at both sites on Sp1 considerably abolished hyperphosphorylation of Sp1 upon infection. Although ATM phosphorylated Ser-101 but not Ser-56 on Sp1 in vitro, phosphorylation of Sp1 at both sites was not detected at all upon infection in ATM-deficient cells, suggesting that cellular kinase(s) activated by ATM could be involved in phosphorylation at Ser-56. Upon viral infection, Sp1-dependent transcription in ATM expression-silenced cells was almost the same as that in ATM-intact cells, suggesting that ATM-dependent phosphorylation of Sp1 might hardly affect its transcriptional activity during the HSV-1 infection. ATM-dependent Sp1 phosphorylation appears to be a global response to various DNA damage stress including viral DNA replication.
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Lin, S. Y., A. R. Black, D. Kostic, S. Pajovic, C. N. Hoover i J. C. Azizkhan. "Cell cycle-regulated association of E2F1 and Sp1 is related to their functional interaction." Molecular and Cellular Biology 16, nr 4 (kwiecień 1996): 1668–75. http://dx.doi.org/10.1128/mcb.16.4.1668.
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Because of the large number of growth-regulated genes containing binding sites for the transcription factors Sp1 and E2F and the reported ability of E2F to mediate cell cycle (growth) regulation, we studied interactions between E2F1 and Sp1. In transient transfection assays using Drosophila melanogaster SL2 cells, transfection with both Sp1 and E2F1 expression vectors resulted in greater than 85-fold activation of transcription from a hamster dihydrofolate reductase reporter construct, whereas cotransfection with either the Sp1 or E2F1 expression vector resulted in 30- or <2-fold activation, respectively. Therefore, these transcription factors act synergistically in activation of dihydrofolate reductase transcription. Transient transfection studies demonstrated that E2F1 could superactivate Sp1-dependent transcription in a promoter containing only Sp1 sites and that Sp1 could superactivate transcription of promoters through E2F sites, further demonstrating that these physically associated in Drosophila cells transfected with Sp1 and E2F1 expression vectors and in human cells, with maximal interaction detected in mid- to late G1. Additionally, E2F1 and Sp1 interact in vitro through specific domains of each protein, and the physical interaction and functional synergism appear to require the same regions. Taken together, these data demonstrate that E2F1 and Sp1 both functionally and physically interact; therefore this interaction, Sp1 and E2F1 may regulate transcription of genes containing binding sites for either or both factors.
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Chupreta, Sergey, Ming Du, Andrea Todisco i Juanita L. Merchant. "EGF stimulates gastrin promoter through activation of Sp1 kinase activity". American Journal of Physiology-Cell Physiology 278, nr 4 (1.04.2000): C697—C708. http://dx.doi.org/10.1152/ajpcell.2000.278.4.c697.
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Epidermal growth factor (EGF) receptor activation stimulates gastrin gene expression through a GC-rich element called gastrin EGF response element (gERE). This element is bound by Sp1 family members and is a target of the ras-extracellular signal-regulated kinase (Erk) signal transduction cascade. This raised the possibility that Sp1 may be phosphorylated by kinases of this signaling pathway. Erk is capable of phosphorylating other mitogen-inducible transcription factors, e.g., Elk and Sap, suggesting that Erk may also mediate EGF-dependent phosphorylation of Sp1. This possibility was tested by studying Sp1-dependent kinase activity in extracts prepared from EGF-activated AGS cells by use of solid-phase kinase assays and immunoprecipitation of metabolically labeled Sp1. The results revealed that Sp1 kinase activity (like gastrin promoter activation) is inhibited by PD-98059 and, therefore, is dependent on mitogen-activated protein kinase kinase 1 (Mek 1). However, EGF-dependent activation of endogenous Erk did not account for most of the Sp1 kinase activity, since Erk and additional Sp1 kinase activity analyzed in a solid-phase kinase assay eluted from an ion-exchange column in different fractions. Phosphoamino acid analysis of in vivo radiolabeled Sp1 demonstrated that the kinase phosphorylates Sp1 on Ser and Thr in response to EGF. Therefore, most EGF-stimulated Sp1 kinase activity is Mek 1 dependent and distinct from Erk.
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Bai, Liqun, James F. Collins, Hua Xu i Fayez K. Ghishan. "Transcriptional regulation of rat Na+/H+exchanger isoform-2 (NHE-2) gene by Sp1 transcription factor". American Journal of Physiology-Cell Physiology 280, nr 5 (1.05.2001): C1168—C1175. http://dx.doi.org/10.1152/ajpcell.2001.280.5.c1168.
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The rat Na+/H+ exchanger isoform-2 ( NHE-2) gene promoter lacks a TATA box and is very GC rich. A minimal promoter extending from bp −36 to +116 directs high-level expression of NHE-2 in mouse inner medullary collecting duct (mIMCD-3) cells. Four Sp1 consensus elements were found in this region. The introduction of mutations within these Sp1 consensus elements and DNA footprinting revealed that only two of them were utilized and are critical for basal transcriptional activation in mIMCD-3 cells. The use of Sp1, Sp3, and Sp4 antisera in electrophoretic mobility shift assays demonstrated that Sp1, Sp3, and Sp4 bound to this minimal promoter. We further analyzed the transcriptional regulation of NHE-2 by members of the Sp1 multigene family. In Drosophila SL2 cells, which lack endogenous Sp1, the minimal promoter cannot drive transcription. Introduction of Sp1 activated transcription over 100-fold, suggesting that Sp1 is critical for transcriptional regulation. However, neither Sp3 nor Sp4 was able to activate transcription in these cells. Furthermore, in mIMCD-3 cells, Sp1-mediated transcriptional activation was repressed by expression of Sp3 and Sp4. These data suggest that Sp1 is critical for the basal promoter function of rat NHE-2 and that Sp3 and Sp4 may repress transcriptional activation by competing with Sp1 for binding to core cis-elements.
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Sugawara, Teruo, Masaki Saito i Seiichiro Fujimoto. "Sp1 and SF-1 Interact and Cooperate in the Regulation of Human Steroidogenic Acute Regulatory Protein Gene Expression*". Endocrinology 141, nr 8 (1.08.2000): 2895–903. http://dx.doi.org/10.1210/endo.141.8.7602.
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Steroidogenic acute regulatory (StAR) protein plays a critical role in the movement of cholesterol from the outer to the inner mitochondrial membrane. Steroidogenic factor 1 (SF-1) controls basal and cAMP-stimulated transcription of the StAR gene. The 1.3-kb StAR promoter has three SF-1 binding sites, and two consensus transcription factor Sp1 binding sequences near the two most distal SF-1 binding sites. Sp1 mediates cAMP-dependent transcription of steroidogenic P450 enzyme genes, raising the possibility of Sp1 involvement in cAMP regulation of the StAR gene. However, the mechanism of Sp1-mediated, cAMP-stimulated responsiveness is not known. In this study, we elucidated the roles of Sp1 and SF-1 in the regulation of the human StAR gene promoter. We found that there was negligible promoter activity in a pGL2 StAR construct (−235 to +39) in which Sp1 and SF-1 binding sites were mutated in Y-1 adrenal tumor cells. An Sp1 binding site mutation (pGL2Sp1M) did not support promoter activity, suggesting that Sp1 cooperates with SF-1 in regulating StAR promoter function. In gel shift assays, the SF-1 binding site formed a complex with an SF-1-GST fusion protein and Sp1. Coimmunoprecipitation cross-linking experiments indicated that SF-1 physically interacts with Sp1 in vitro. Finally, a mammalian two-hybrid system was employed to demonstrate that Sp1 and SF-1 associate in vivo. In conclusion, our data indicate that Sp1 and SF-1 physically interact and cooperate in the regulation of human StAR promoter activity.
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Zhang, Ying, Mingjuan Liao i Maria L. Dufau. "Phosphatidylinositol 3-Kinase/Protein Kinase Cζ-Induced Phosphorylation of Sp1 and p107 Repressor Release Have a Critical Role in Histone Deacetylase Inhibitor-Mediated Depression of Transcription of the Luteinizing Hormone Receptor Gene". Molecular and Cellular Biology 26, nr 18 (15.09.2006): 6748–61. http://dx.doi.org/10.1128/mcb.00560-06.
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ABSTRACT We have demonstrated that silencing of luteinizing hormone receptor (LHR) gene transcription is mediated via a proximal Sp1 site at its promoter. Trichostatin A (TSA) induced histone acetylation and gene activation in JAR cells that prevailed in the absence of changes in Sp1/Sp3 expression, their binding activity, disassociation of the histone deacetylase/mSin3A complex from the Sp1 site, or demethylation of the promoter. This indicated a different mechanism involved in TSA-induced derepression. The present studies have revealed that phosphatidylinositol 3-kinase/protein kinase Cζ (PI3K/PKCζ)-mediated Sp1 phosphorylation accounts for Sp1 site-dependent LHR gene activation. TSA caused marked phosphorylation of Sp1 at serine 641 in JAR and MCF-7 cells. Blockade of PI3K or PKCζ activity by specific inhibitors, kinase-deficient mutants, or small interfering RNA abolished the effect of TSA on the LHR gene and Sp1 phosphorylation. PKCζ was shown to associate with Sp1, and this association was enhanced by TSA. Sp1 phosphorylation at serine 641 was required for the release of the pRb homologue p107 from the LHR gene promoter, while p107 acted as a repressor of the LHR gene. Inhibition of PKCζ activity blocked the dissociation of p107 from the LHR gene promoter and markedly reduced Sp1 phosphorylation and transcription. These results have demonstrated that phosphorylation of Sp1 by PI3K/PKCζ is critical for TSA-activated LHR gene expression. These studies have revealed a novel mechanism of TSA action through derecruitment of a repressor from the LHR gene promoter in a PI3K/PKCζ-induced Sp1 phosphorylation-dependent manner.
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Chen, Shuqin, Huating Li, Jing Zhang, Shan Jiang, Mingliang Zhang, Yilan Xu, Kun Dong, Ying Yang, Qichen Fang i Weiping Jia. "Identification of Sp1 as a Transcription Activator to Regulate Fibroblast Growth Factor 21 Gene Expression". BioMed Research International 2017 (2017): 1–10. http://dx.doi.org/10.1155/2017/8402035.
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Fibroblast growth factor 21 (FGF21) is a metabolic hormone with multiple beneficial effects on lipid and glucose homeostasis. Previous study demonstrated that FGF21 might be one of the Sp1 target genes. However, the transcriptional role of Sp1 on FGF21 in adipose tissue and liver has not been reported. In this study, we found that the proximal promoter of mouse FGF21 is located between −63 and −20 containing two putative Sp1-binding sites. Sp1 is a mammalian transcription factor involved in the regulation of many genes during physiological and pathological processes. Our study showed that overexpression of Sp1 or suppressing Sp1 expression resulted in increased or reduced FGF21 promoter activity, respectively. Mutation analysis demonstrated that the Sp1-binding site located between −46 and −38 plays a primary role in transcription of FGF21. Electrophoretic mobility shift assay and chromatin immunoprecipitation analysis indicated that Sp1 specifically bound to this region. Furthermore, the binding activity of Sp1 was significantly increased in adipose tissues of HFD-induced obese mouse and liver of DEN-treated mouse. Thus, our results demonstrate that Sp1 positively regulates the basal transcription of FGF21 in the liver and adipose tissue and contributes to the obesity-induced FGF21 upregulation in mouse adipose tissue and hepatic FGF21 upregulation in hepatocarcinogenesis.
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He, Shihua, Jian-Min Sun, Lin Li i James R. Davie. "Differential Intranuclear Organization of Transcription Factors Sp1 and Sp3". Molecular Biology of the Cell 16, nr 9 (wrzesień 2005): 4073–83. http://dx.doi.org/10.1091/mbc.e05-05-0388.
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Sp1 and Sp3 are ubiquitously expressed mammalian transcription factors that activate or repress the expression of a variety of genes and are thought to compete for the same DNA binding site. We used indirect immunofluorescence microscopy and image deconvolution to show that Sp1 and Sp3 are organized into distinct nonoverlapping domains in human breast and ovarian cells. Domains of Sp1 and Sp3 infrequently associate with sites of transcription. Sp3 partitions with the tightly bound nuclear protein fraction of hormone responsive MCF-7 breast cancer cells, whereas only a subpopulation of Sp1 is found in that fraction. Both Sp1 and Sp3 are bound to the nuclear matrix, and the nuclear matrix-associated sites of Sp1 and Sp3 are different. Indirect immunofluorescence studies demonstrate that Sp1 and Sp3 associate with histone deacetylases 1 and 2 and with the estrogen receptor α, albeit at low frequencies in MCF-7 cells. Chromatin immunoprecipitation (ChIP) and re-ChIP assays revealed that although both Sp1 and Sp3 bind to the estrogen-responsive trefoil factor 1 promoter in MCF-7 cells, they do not occupy the same promoter. Our results demonstrate the different features of Sp1 and Sp3, providing further evidence that Sp3 is not a functional equivalent of Sp1.
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Fulciniti, Mariateresa, Samir B. Amin, Puru Nanjappa, Scott J. Rodig, Teru Hideshima, Jagannath Pal, Varuna Mohan i in. "Biology and Therapeutic Targeting of Sp1 Transactivation In Myeloma". Blood 116, nr 21 (19.11.2010): 134. http://dx.doi.org/10.1182/blood.v116.21.134.134.
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Abstract Abstract 134 Alteration in expression and function of transcription factors has been frequently associated with neoplastic transformation. We here provide both experimental and clinical evidence that Sp1, a transcription factor that controls number of cellular processes, plays an important regulatory role in MM cell growth and survival. Although Sp1 is ubiquitously expressed, its nuclear localization observed in MM is functionally both relevant and important. We have confirmed high Sp1 activity in MM cells both by demonstrating increased DNA binding as well as increased Sp1-responsive promoter activity measured by luciferase reporter assay. MM cell-BMSC interaction led to Sp1 activation which was completely abrogated by the ERK pathway inhibitor U0126 but not by the AKT inhibitor LY29004. Furthermore, using both SiRNA and ShRNA mediated Sp1 knock-down, we have confirmed the growth and survival effects of Sp1 on MM cell growth. Using gene expression profile of MM cells from 172 uniformly treated patients, we have further confirmed these in vitro results by observing that overexpression of Sp1-related genes, including HIF-1a, HDAC1 and MAPK genes, correlate with poor prognosis in MM. This clinical correlation further suggests the role of Sp1 in MM biology, providing the rationale to preclinically target Sp1 in MM. We have investigated TMP, a derivative of the plant lignan nordihydroguaiaretic acid (NDGA) which disrupts the interaction between Sp1 and GC-rich motifs inhibiting Sp1 activity. We have previously confirmed specific inhibition of both Sp1 binding and transcriptional activity in MM cells by TMP, including in the context of MM-BMSC interaction. Along with inhibition of Sp1 activity, we observed both in vitro and in vivo in murine models of human myeloma, anti-myeloma effect of TMP. Importantly, there was no significant synergistic effect when MM cells transfected with Sp1 siRNA were treated with TMP, confirming specificity of TMP's mechanism of action. We have further observed activation of the mitochondrial apoptotic pathway by TMP via activation of caspase-3, -9 and -7 and PARP cleavage while caspase-8 was not activated suggesting possible synergism with activators of alternate apoptotic pathways. We have also observed that lenalidomide and dexamethasone upregulate Sp1 activity, suggesting that Sp1 may be a common resistance mechanism. We have confirmed that the increased Sp1 activity by lenalidomide or dexamethasone is abrogated by TMP and the combination provides synergistic cytotoxicity, in MM cell lines as well as primary MM cells. In conclusion, we report significant role of Sp1 in myeloma cell growth, survival and drug resistance with its influence on clinical outcome in MM. Our results suggest that specific inhibition of Sp1 activity may be an important therapeutic target in MM. Disclosures: Avet-Loiseau: Celgene: Membership on an entity's Board of Directors or advisory committees; Janssen-Cilag: Membership on an entity's Board of Directors or advisory committees. Anderson:Millennium Pharmaceuticals: Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding, Speakers Bureau. Munshi:Millennium Pharmaceuticals: Honoraria, Speakers Bureau.
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Yan, Yutao, Guillaume Dalmasso, Shanthi Sitaraman i Didier Merlin. "Characterization of the human intestinal CD98 promoter and its regulation by interferon-γ". American Journal of Physiology-Gastrointestinal and Liver Physiology 292, nr 2 (luty 2007): G535—G545. http://dx.doi.org/10.1152/ajpgi.00385.2006.
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Growing evidence that epithelial CD98 plays an important role in intestinal inflammation focused our interest to investigate the transcriptional regulation of CD98. Our mouse-based in vivo and in vitro experiments revealed that epithelial colonic CD98 mRNA expression was transcriptionally increased in intestinal inflammation. We then isolated and characterized a 5′-flanking fragment containing the promoter region required for CD98 gene transcription. Primer extension and rapid amplification of 5′-cDNA ends were used to map a transcriptional initiation site 129 bp upstream from the translational start codon (ATG). Direct sequencing of the 5′-flanking region revealed the presence of four GC-rich stimulating protein (Sp)1 binding domains, one NF-κB binding domain, and no TATA box. Binding of Sp1 [Sp1(−874), SP1(−386), Sp1(−187), and Sp1(−177)] and NF-κB [NF-κB(−213)] to the promoter was confirmed by EMSA and supershift assays. Furthermore, chromatin immunoprecipitation experiments showed the in vivo DNA-Sp1 and DNA-NF-κB interactions under basal and IFN-γ-stimulated conditions. Reporter genes driven by serially truncated and site-mutated CD98 promoters were used to examine basal and IFN-γ-responsive transcription in transiently transfected Caco2-BBE cells. Our results revealed that Sp1(−187), Sp1(−177), and the NF-κB binding site were essential for basal and IFN-γ-stimulated CD98 promoter activities, whereas Sp1(−874) and Sp1(−386) were not. The results from additional site-mutated CD98 promoters suggested that Sp1(−187), Sp1(−177), and the NF-κB site may cooperate in mediating basal and IFN-γ-stimulated CD98 promoter activities. Finally, we demonstrated that a reduction of Sp1 or NF-κB expression reduced CD98 protein expression in unstimulated and IFN-γ-stimulated Caco2-BBE cells. Collectively, these findings indicate that the Sp1 and NF-κB transcription factors are likely to play a significant role in IFN-γ-mediated transcriptional regulation of CD98 in the intestinal epithelium, providing new insights into the regulation of CD98 expression in intestinal inflammation.
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Majumdar, Gipsy, Ashley Harmon, Rosalind Candelaria, Antonio Martinez-Hernandez, Rajendra Raghow i Solomon S. Solomon. "O-glycosylation of Sp1 and transcriptional regulation of the calmodulin gene by insulin and glucagon". American Journal of Physiology-Endocrinology and Metabolism 285, nr 3 (wrzesień 2003): E584—E591. http://dx.doi.org/10.1152/ajpendo.00140.2003.
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Both insulin and glucagon stimulate steady-state levels of Sp1 transcription factor, but only insulin stimulates transcription of the calmodulin (CaM) gene in liver. Because O-glycosylation of Sp1 by O-linked N-acetylglucosamine ( O-GlcNAc) is thought to regulate its ability to activate transcription, we assayed the levels of Sp1 with anti-Sp1 and anti- O-GlcNAc antibodies in Western blots by use of extracts of H-411E liver cells treated with insulin (10,000 μU/ml) or glucagon (1.5 × 10-5 M). We also assessed subcellular localization of the native and glycosylated Sp1 in H411E cells treated with either hormone in the presence of deoxynorleucine (DON, an indirect inhibitor of O-glycosylation) or streptozotocin (STZ, an indirect stimulator of O-glycosylation). Insulin stimulated both total and O-GlcNAc-modified Sp1 primarily in the nucleus and induced CaM gene transcription ( P < 0.0001). In contrast, glucagon promoted accumulation of Sp1 in the cytoplasm but not the nucleus, without significantly stimulating ( P = not significant) either its O-glycosylation or transcription of the CaM gene. DON inhibited O-glycosylation of Sp1 and its ability to migrate to the nucleus and transactivate CaM gene transcription. In contrast, cotreatment of cells with STZ and glucagon enhanced O-glycosylation of Sp1, promoting its migration to the nucleus and resulting in increased CaM gene transcription. Thus O-glycosylation of Sp1 by insulin, but not glucagon, apparently enhances its (Sp1) nuclear recruitment and results in activation of CaM gene transcription.
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Datta, P. K., P. Raychaudhuri i S. Bagchi. "Association of p107 with Sp1: genetically separable regions of p107 are involved in regulation of E2F- and Sp1-dependent transcription." Molecular and Cellular Biology 15, nr 10 (październik 1995): 5444–52. http://dx.doi.org/10.1128/mcb.15.10.5444.
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The retinoblastoma-related protein p107 has been shown to be a regulator of the transcription factor E2F. p107 associates with E2F via its pocket region and represses E2F-dependent transcription. In this study, we provide evidence for a novel interaction between p107 and the transcription factor Sp1. We show that p107 can be found endogenously associated with Sp1 in the extracts of several different cell lines. Moreover, in transient transfection assays, expression of p107 represses Sp1-dependent transcription. This repression of Sp1-dependent transcription does not require the DNA-binding domain of Sp1. Transcription driven by a chimeric protein containing the Ga14 DNA-binding domain and the Sp1 activation domains is inhibited by p107. Interestingly, unlike the repression of E2F-dependent transcription, the repression of Sp1-dependent transcription does not depend on an intact pocket region. We show that distinct regions of p107 are involved in the control of Sp1 and E2F.
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Isnawati, Isnawati. "Aktivitas Sellulolitik Fungi Indigenus pada Fermetoge: Pakan Fermentasi Hewan Ruminansia Terbuat dari Eceng Gondok (Eichhornia crassipes) dan Tongkol Jagung (Zea mays)". Jurnal Riset Biologi dan Aplikasinya 1, nr 1 (28.01.2019): 26. http://dx.doi.org/10.26740/jrba.v1n1.p26-31.
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Eceng gondok dan tongkol jagung tergolong bahan bersellulosa. Pada campuran kedua bahan itu terdapat mikroba indigenus. Tujuan pertama riset ini untuk mengetahui aktivitas sellulolitik fungi indigenus yang terdapat pada fermetoge, pakan fermentasi dari campuran eceng gondok dan tongkol jagung. Eceng gondok dipotong dan tongkol jagung dihancurkan sampai berukuran sekitar 1-2 cm, dikukus, dan difermentasi secara alamiah menggunakan mikroorganisme indigenus. Mikroorganisme tersebut diisolasi dari pakan tersebut setiap hari selama 15 hari selama fermentasi berlangsung. Selanjutnya,isolate yang diperoleh dimurnikan, dikarakterisasi, dan diidentifikasi. Terdapat 10 fungi indigenus dalam pakan. Berdasarkan observasi karakteristik mikroskopik dan makroskopik fungi-fungi tersebut meliputi Aspergillus sp1, Rhizopus sp1, Aspergillus terreus, Mucor sp1, Aspergillus sp2, Aspergillus niger, Trichoderma sp1, Aspergillus flavus, Aspergillus sp3, dan Penicillium sp1. Uji aktivitas sellulolitik pada medium spesifik CMC memaparkan bahwa Mucor Sp1, Rhizopus sp1 dan Trichoderma sp1 adalah tiga fungi dengan aktivitas sellulolitik tinggi, karena membentuk zona halo yang luas pada permukaan media setelah diwarnai dengan Congo red 2%.
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SU, Kaihong, Xiaoyong YANG, Mark D. ROOS, Andrew J. PATERSON i Jeffrey E. KUDLOW. "Human Sug1/p45 is involved in the proteasome-dependent degradation of Sp1". Biochemical Journal 348, nr 2 (23.05.2000): 281–89. http://dx.doi.org/10.1042/bj3480281.
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The transcription factor Sp1 was previously shown to undergo proteasome-dependent degradation when cells were glucose-starved and stimulated with the adenylate cyclase inducer, forskolin. However, the control of the Sp1 degradation process is largely unknown. Using in vitro and in vivo interaction studies, we show in the present study that Sp1 interacts with human Sug1 [hSug1, also known as p45 or thyroid-hormone-receptor interacting protein (‘TRIP1’)], an ATPase subunit of the 26 S proteasome and a putative transcriptional modulator. This interaction with Sp1 occurs through the C-terminus of hSug1, the region that contains the conserved ATPase domain in this protein. Both in vitro studies, in reconstituted degradation assays, and in vivo experiments, in which hSug1 is overexpressed in normal rat kidney cells, show that full-length hSug1 is able to stimulate the proteasome-dependent degradation of Sp1. However, hSug1 truncations that lack either the N- or C-terminal domain of hSug1 act as dominant negatives, inhibiting Sp1 degradation in vitro. Also, an ATPase mutant of hSug1, while still able to bind Sp1, acts as a dominant negative, blocking Sp1 degradation both in vitro and in vivo. These results demonstrate that hSug1 is involved in the degradation of Sp1 and that ATP hydrolysis by hSug1 is necessary for this process. Our findings indicate that hSug1 is an exchangeable proteasomal component that plays a critical regulatory role in the proteasome-dependent degradation of Sp1. However, hSug1 is not the factor limiting Sp1 degradation in the cells treated with glucosamine. This and other considerations suggest that hSug1 co-operation with other molecules is necessary to target Sp1 for proteasome degradation.
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Noti, J. D., B. C. Reinemann i M. N. Petrus. "Sp1 binds two sites in the CD11c promoter in vivo specifically in myeloid cells and cooperates with AP1 to activate transcription." Molecular and Cellular Biology 16, nr 6 (czerwiec 1996): 2940–50. http://dx.doi.org/10.1128/mcb.16.6.2940.
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The leukocyte integrin gene, CD11c, is transcriptionally regulated and is expressed predominantly on differentiated cells of the myelomonocytic lineage. In this study we have demonstrated that the regions -72 to -63 and -132 to -104 of the CD11c promoter contain elements responsible for phorbol ester-induced differentiation of the myeloid cell line HL60. DNase I footprinting analysis revealed that these regions can bind purified Sp1, and supershift analysis with Sp1 antibody confirmed that Sp1 in HL60 nuclear extracts could bind these regions. Transfection analysis of CD11c promoter-chloramphenicol acetyltransferase constructs containing deletions of these Sp1-binding sites revealed that these sites are essential for expression of the CD11c gene in HL60 cells but not in the T-cell line Molt4 or the cervical carcinoma cell line HeLa. Moreover, cotransfection of pPacSp1 along with these CD11c promoter-chloramphenicol acetyltransferase constructs into Sp1-deficient Drosophila Schneider 2 cells verified that these sites are essential for Sp1-dependent expression of the CD11c promoter. In vivo genomic footprinting revealed that Sp1 contacts the CD11c promoter within the regions -69 to -63 and -116 to -105 in phorbol 12-myristate 13-acetate-differentiated HL60 cells but not in undifferentiated HL60 cells or in Molt4 or HeLa cells. Cotransfection assays in HL60 cells revealed that Sp1 acts synergistically with Ap1 to activate CD11c. Further, both Sp1 sites are capable of cooperating with AP1. In vitro DNase I footprinting analysis with purified Sp1 and c-jun proteins showed that Sp1 binding could facilitate binding of c-jun. We propose that myeloid-specific expression of the CD11c promoter and is facilitated by cooperative interaction between the Sp1- and Ap1-binding sites.
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Li, Jie, i Jing-hsiung Ou. "Differential Regulation of Hepatitis B Virus Gene Expression by the Sp1 Transcription Factor". Journal of Virology 75, nr 18 (15.09.2001): 8400–8406. http://dx.doi.org/10.1128/jvi.75.18.8400-8406.2001.
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ABSTRACT The expression of hepatitis B virus (HBV) genes is regulated by a number of transcription factors. One such factor, Sp1, has two binding sites in the core promoter and one in its upstream regulatory element, which is also known as the ENII enhancer. In this study, we have analyzed the effects of these three Sp1 binding sites on the expression of HBV genes. Our results indicate that both Sp1 binding sites in the core promoter are important for the transcription of the core RNA and the precore RNA. Moreover, while the downstream Sp1 site (the Sp1-1 site) in the core promoter did not affect the transcription of the S gene and the X gene, the upstream Sp1 site (the Sp1-2 site) in the core promoter was found to negatively regulate the transcription of the S gene and the X gene, as removal of the latter led to enhancement of transcription of these two genes. The Sp1 binding site in the ENII enhancer (the Sp1-3 site) positively regulates the expression of all of the HBV genes, as its removal by mutation suppressed the expression of all of the HBV genes. However, the suppressive effect of the Sp1-3 site mutation on the expression of the S gene and the X gene was abolished if the two Sp1 sites in the core promoter were also mutated. These results indicate that Sp1 can serve both as a positive regulator and as a negative regulator for the expression of HBV genes. This dual activity may be important for the differential regulation of HBV gene expression.
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Liu, Shujun, Zhongfa Liu, Zhiliang Xie, Jiuxia Pang, Jianhua Yu, Esther Lehmann, Lenguyen Huynh i in. "Bortezomib induces DNA hypomethylation and silenced gene transcription by interfering with Sp1/NF-κB–dependent DNA methyltransferase activity in acute myeloid leukemia". Blood 111, nr 4 (15.02.2008): 2364–73. http://dx.doi.org/10.1182/blood-2007-08-110171.
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Bortezomib reversibly inhibits 26S proteasomal degradation, interferes with NF-κB, and exhibits antitumor activity in human malignancies. Zinc finger protein Sp1 transactivates DNMT1 gene in mice and is functionally regulated through protein abundance, posttranslational modifications (ie, ubiquitination), or interaction with other transcription factors (ie, NF-κB). We hypothesize that inhibition of proteasomal degradation and Sp1/NF-κB–mediated transactivation may impair aberrant DNA methyltransferase activity. We show here that, in addition to inducing accumulation of polyubiquitinated proteins and abolishment of NF-κB activities, bortezomib decreases Sp1 protein levels, disrupts the physical interaction of Sp1/NF-κB, and prevents binding of the Sp1/NF-κB complex to the DNMT1 gene promoter. Abrogation of Sp1/NF-κB complex by bortezomib causes transcriptional repression of DNMT1 gene and down-regulation of DNMT1 protein, which in turn induces global DNA hypomethylation in vitro and in vivo and re-expression of epigenetically silenced genes in human cancer cells. The involvement of Sp1/NF-κB in DNMT1 regulation is further demonstrated by the observation that Sp1 knockdown using mithramycin A or shRNA decreases DNMT1 protein levels, which instead are increased by Sp1 or NF-κB overexpression. Our results unveil the Sp1/NF-κB pathway as a modulator of DNA methyltransferase activity in human cancer and identify bortezomib as a novel epigenetic-targeting drug.
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Vellingiri, Iyer, Devi Subramaniam, Jayaramayya, Siama, Giridharan, Narayanasamy, Abdal Dayem i Cho. "Understanding the Role of the Transcription Factor Sp1 in Ovarian Cancer: from Theory to Practice". International Journal of Molecular Sciences 21, nr 3 (9.02.2020): 1153. http://dx.doi.org/10.3390/ijms21031153.
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Ovarian cancer (OC) is one of the deadliest cancers among women contributing to high risk of mortality, mainly owing to delayed detection. There is no specific biomarker for its detection in early stages. However, recent findings show that over-expression of specificity protein 1 (Sp1) is involved in many OC cases. The ubiquitous transcription of Sp1 apparently mediates the maintenance of normal and cancerous biological processes such as cell growth, differentiation, angiogenesis, apoptosis, cellular reprogramming and tumorigenesis. Sp1 exerts its effects on cellular genes containing putative GC–rich Sp1–binding site in their promoters. A better understanding of the mechanisms underlying Sp1 transcription factor (TF) regulation and functions in OC tumorigenesis could help identify novel prognostic markers, to target cancer stem cells (CSCs) by following cellular reprogramming and enable the development of novel therapies for future generations. In this review, we address the structure, function, and biology of Sp1 in normal and cancer cells, underpinning the involvement of Sp1 in OC tumorigenesis. In addition, we have highlighted the influence of Sp1 TF in cellular reprogramming of iPSCs and how it plays a role in controlling CSCs. This review highlights the drugs targeting Sp1 and their action on cancer cells. In conclusion, we predict that research in this direction will be highly beneficial for OC treatment, and chemotherapeutic drugs targeting Sp1 will emerge as a promising therapy for OC.
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Rodenburg, Richard J. T., P. Elly Holthuizen i John S. Sussenbach. "A Functional Sp1 Binding Site Is Essential for the Activity of the Adult Liver-Specific Human Insulin-Like Growth Factor II Promoter". Molecular Endocrinology 11, nr 2 (1.02.1997): 237–50. http://dx.doi.org/10.1210/mend.11.2.9888.
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Abstract The human gene encoding insulin-like growth factor II contains four promoters (P1–P4) that are differentially activated in various tissues during development. Expression of insulin-like growth factor II in adult liver tissue is directed by P1, which is activated by liver-enriched members of the CCAAT/enhancer binding protein family of transcription factors. In the present report we show that the region around −48 relative to the transcription start site contains a high affinity Sp1 binding site. This was demonstrated by electrophoretic mobility shift assays using nuclear extracts from Hep3B hepatoma cells and with specific antibodies directed against Sp1. Competition electrophoretic mobility shift assays revealed that the Sp1 binding site of P1 and a consensus Sp1 binding site bind Sp1 with comparable efficiencies. Mutation of the Sp1 binding site results in an 85% decrease in P1 promoter activity in transient transfection assays using two different cell lines, COS-7 and Hep3B. Investigation of P1 mutants in which the spacing of the Sp1 binding site and the transcription start site was increased showed that the role of the Sp1 binding site in regulation of P1 is position dependent. Interestingly, the Sp1-responsive element cannot be exchanged by a functional TATA box. Activation of P1 by transactivators CCAAT/enhancer binding protein-β and hepatocyte nuclear factor-3β is strongly impaired after mutation of the Sp1 binding site. These results demonstrate that the specific presence of a binding site for the ubiquitously expressed transcription factor Sp1 is of eminent importance for efficient activation of P1 by liver-enriched transactivators.
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Jiang, Jianhai, Yuanyan Wei, Jialin Shen, Dan Liu, Xiaoning Chen, Jin Zhou, Hongliang Zong i in. "Functional Interaction of E1AF and Sp1 in Glioma Invasion". Molecular and Cellular Biology 27, nr 24 (15.10.2007): 8770–82. http://dx.doi.org/10.1128/mcb.02302-06.
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ABSTRACT Transcription factor E1AF is widely known to play critical roles in tumor metastasis via directly binding to the promoters of genes involved in tumor migration and invasion. Here, we report for the first time E1AF as a novel binding partner for ubiquitously expressed Sp1 transcription factor. E1AF forms a complex with Sp1, contributes to Sp1 phosphorylation and transcriptional activity, and functions as a mediator between epidermal growth factor and Sp1 phosphorylation and activity. Sp1 functions as a carrier bringing E1AF to the promoter region, thus activating transcription of glioma-related gene for β1,4-galactosyltransferase V (GalT V; EC 2.4.1.38). Biologically, E1AF functions as a positive invasion regulator in glioma in cooperation with Sp1 partly via up-regulation of GalT V. This report describes a new mechanism of glioma invasion involving a cooperative effort between E1AF and Sp1 transcription factors.
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Gordon-Shaag, Ariela, Orly Ben-Nun-Shaul, Vered Roitman, Yael Yosef i Ariella Oppenheim. "Cellular Transcription Factor Sp1 Recruits Simian Virus 40 Capsid Proteins to the Viral Packaging Signal, ses". Journal of Virology 76, nr 12 (15.06.2002): 5915–24. http://dx.doi.org/10.1128/jvi.76.12.5915-5924.2002.
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ABSTRACT Simian virus 40 (SV40) capsid assembly occurs in the nucleus. All three capsid proteins bind DNA nonspecifically, raising the dilemma of how they attain specificity to the SV40 minichromosome in the presence of a large excess of genomic DNA. The SV40 packaging signal, ses, which is required for assembly, is composed of multiple DNA elements that bind transcription factor Sp1. Our previous studies showed that Sp1 participates in SV40 assembly and that it cooperates in DNA binding with VP2/3. We hypothesized that Sp1 recruits the capsid proteins to the viral minichromosome, conferring upon them specific DNA recognition. Here, we have tested the hypothesis. Computer analysis showed that the combination of six tandem GC boxes at ses is not found at cellular promoters and therefore is unique to SV40. Cooperativity in DNA binding between Sp1 and VP2/3 was not abolished at even a 1,000-fold excess of cellular DNA, providing strong support for the recruitment hypothesis. Sp1 also binds VP1 and cooperates with VP1 in DNA binding. VP1 pentamers (VP15) avidly interact with VP2/3, utilizing the same VP2/3 domain as described for polyomavirus. We conclude that VP15-VP2/3 building blocks are recruited by Sp1 to ses, where they form the nucleation center for capsid assembly. By this mechanism the virus ensures that capsid formation is initiated at a single site around its minichromosome. Sp1 enhances the formation of SV40 pseudovirions in vitro, providing additional support for the model. Analyses of Sp1 and VP3 deletion mutants showed that Sp1 and VP2/3 bind one another and cooperate in DNA binding through their DNA-binding domains, with additional contacts outside these domains. VP1 contacts Sp1 at residues outside the Sp1 DNA-binding domain. These and additional data allowed us to propose a molecular model for the VP15-VP2/3-DNA-Sp1 complex.
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Sriraman, Venkataraman, S. Chidananda Sharma i JoAnne S. Richards. "Transactivation of the Progesterone Receptor Gene in Granulosa Cells: Evidence that Sp1/Sp3 Binding Sites in the Proximal Promoter Play a Key Role in Luteinizing Hormone Inducibility". Molecular Endocrinology 17, nr 3 (1.03.2003): 436–49. http://dx.doi.org/10.1210/me.2002-0252.
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Abstract LH induction of the progesterone receptor (PR) in granulosa cells is a central event in ovulation. To identify critical regions of the mouse PR promoter that confer LH inducibility in granulosa cells, a mouse PR promoter (−384/+680) genomic fragment was ligated to a luciferase reporter construct and transfected into primary cultures of granulosa cells. Forskolin/phorbol myristate (PMA) induced PR promoter-luciferase reporter activity in granulosa cells greater than 15-fold. A deletion construct comprised only of the distal promoter alone (−348/+64) was inactive. Conversely, deletion constructs eliminating putative distal promoter-regulatory elements that bind Sp1, nuclear factor Y, and GATA-4 as well as the transcription start site (+1) failed to reduce forskolin/PMA activation of reporter activity. Additional 5′-deletions identified a minimal promoter region (+420/+680) sufficient to bestow cAMP responsiveness approximately 8- to 10-fold. Two GC-rich regions Sp1(A)[+440/+461] and Sp1(B) [+473/+490] bound Sp1/Sp3. Site-directed mutagenesis of Sp1(A) and Sp1(B) reduced activity of the proximal (+357/+680) promoter reporter construct approximately 50% and 99%, respectively. When the same Sp1(B) mutation was introduced into the intact promoter (−145/+680), forskolin/PMA induction of promoter activity was reduced by 70–80%. When the distal GC box as well as the proximal Sp1(B) site were both mutated in the context of the intact promoter, inducibility of the transgene was even more severely reduced. The importance of these Sp1/Sp3 binding regions was confirmed in human MCF-7 cells and Drosophila SL2 cells. Collectively, these results indicate that the Sp1/Sp3 binding sites within the mouse PR proximal promoter are essential for transactivation of the gene by agonists in granulosa cells. The molecular mechanisms by which LH activates Sp1/Sp3 at this region within the PR gene remain unknown but do not involve changes in the binding of Sp1/Sp3 to the critical GC boxes. Rather, Sp1/Sp3 appear to recruit other factors to the promoter.