Zhangfei: a second cellular protein interacts with herpes simplex virus accessory factor HCF in a manner similar to Luman and VP16.
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Host cell factor (HCF, C1, VCAF or CFF) is a cellular protein that is required for transcription activation of herpes simplex virus (HSV) immediate-early (IE) genes by the virion protein VP16. The biological function of HCF remains unclear. Recently we identified a cellular transcription activator, Luman. As with VP16, the transactivation function of Luman is also regulated by HCF. Here we report a second human protein, Zhangfei (ZF) that interacts with HCF in a fashion similar to Luman and VP16. Although ZF shares no significant sequence homology with Luman, the two proteins have some structural similarities. These include: a basic domain-leucine zipper (bZIP) region, an acidic activation domain and a consensus HCF-binding motif. Unlike Luman, or most other bZIP proteins, ZF by itself did not appear to bind consensus bZIP-binding sites. It was also unable to activate promoters containing these response elements. Although in transient expression assays ectopically expressed ZF was unable to block transactivation by VP16 of a HSV IE promoter, ZF could prevent the expression of several HSV proteins in cells infected with the virus. The ability of ZF to block the synthesis of the HSV IE protein ICP0 relied on its binding to HCF, since a mutant of ZF that was unable to bind HCF was also unable to prevent viral IE protein expression. (+info)
Repression of IL-2 promoter activity by the novel basic leucine zipper p21SNFT protein.
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IL-2 is the major autocrine and paracrine growth factor produced by T cells upon T cell stimulation. The inducible expression of IL-2 is highly regulated by multiple transcription factors, particularly AP-1, which coordinately activate the promoter. Described here is the ability of the novel basic leucine zipper protein p21SNFT to repress AP-1 activity and IL-2 transcription. A detailed analysis of the repression by p21SNFT repression on the IL-2 promoter distal NF-AT/AP-1 site demonstrates that it can bind DNA with NF-AT and Jun, strongly suggesting that it represses NF-AT/AP-1 activity by competing with Fos proteins for Jun dimerization. The importance of this repression is that p21SNFT inhibits the trans-activation potential of protein complexes that contain Jun, thereby demonstrating an additional level of control for the highly regulated, ubiquitous AP-1 transcription factor and the IL-2 gene. (+info)
The leucine zipper of NRL interacts with the CRX homeodomain. A possible mechanism of transcriptional synergy in rhodopsin regulation.
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Photoreceptor-specific expression of rhodopsin is mediated by multiple cis-acting elements in the proximal promoter region. NRL (neural retina leucine zipper) and CRX (cone rod homeobox) proteins bind to the adjacent NRE and Ret-4 sites, respectively, within this region. Although NRL and CRX are each individually able to induce rhodopsin promoter activity, when expressed together they exhibit transcriptional synergy in rhodopsin promoter activation. Using the yeast two-hybrid method and glutathione S-transferase pull-down assays, we demonstrate that the leucine zipper of NRL can physically interact with CRX. Deletion analysis revealed that the CRX homeodomain (CRX-HD) plays an important role in the interaction with the NRL leucine zipper. Although binding with the CRX-HD alone was weak, a strong interaction was detected when flanking regions including the glutamine-rich and the basic regions that follow the HD were included. A reciprocal deletion analysis showed that the leucine zipper of NRL is required for interaction with CRX-HD. Two disease-causing mutations in CRX-HD (R41W and R90W) that exhibit reduced DNA binding and transcriptional synergy also decrease its interaction with NRL. These studies suggest novel possibilities for protein-protein interaction between two conserved DNA-binding motifs and imply that cross-talk among distinct regulatory pathways contributes to the establishment and maintenance of photoreceptor function. (+info)
HMG-I/Y, a new c-Myc target gene and potential oncogene.
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The HMG-I/Y gene encodes the HMG-I and HMG-Y proteins, which function as architectural chromatin binding proteins important in the transcriptional regulation of several genes. Although increased expression of the HMG-I/Y proteins is associated with cellular proliferation, neoplastic transformation, and several human cancers, the role of these proteins in the pathogenesis of malignancy remains unclear. To better understand the role of these proteins in cell growth and transformation, we have been studying the regulation and function of HMG-I/Y. The HMG-I/Y promoter was cloned, sequenced, and subjected to mutagenesis analysis. A c-Myc-Max consensus DNA binding site was identified as an element important in the serum stimulation of HMG-I/Y. The oncoprotein c-Myc and its protein partner Max bind to this site in vitro and activate transcription in transfection experiments. HMG-I/Y expression is stimulated by c-Myc in a Myc-estradiol receptor cell line in the presence of the protein synthesis inhibitor cycloheximide, indicating that HMG-I/Y is a direct c-Myc target gene. HMG-I/Y induction is decreased in Myc-deficient fibroblasts. HMG-I/Y protein expression is also increased in Burkitt's lymphoma cell lines, which are known to have increased c-Myc protein. Like Myc, increased expression of HMG-I protein leads to the neoplastic transformation of both Rat 1a fibroblasts and CB33 cells. In addition, Rat 1a cells overexpressing HMG-I protein form tumors in nude mice. Decreasing HMG-I/Y proteins using an antisense construct abrogates transformation in Burkitt's lymphoma cells. These findings indicate that HMG-I/Y is a c-Myc target gene involved in neoplastic transformation and a member of a new class of potential oncogenes. (+info)
CDC6 interacts with c-Myc to inhibit E-box-dependent transcription by abrogating c-Myc/Max complex.
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The c-myc oncogene product (c-Myc) is a transcription factor that dimerizes with Max and recognizes the E-box sequence. It plays key functions in cell proliferation, differentiation and apoptosis. It is generally thought that c-Myc transactivates genes encoding proteins essential to cell-cycle progression by binding to the E-boxes that control them. The functions of c-Myc are also thought to be modulated by its associated proteins, several of which have recently been identified. In this study, we found that c-Myc directly bound in vivo and in vitro to the N-terminal region of human CDC6, a component of the pre-replication complex, and that both co-localized in cell nuclei. CDC6 bound to the C-proximal region of c-Myc, thereby competing with Max on the E-box sequence and changing c-Myc/Max heterodimer to a Max/Max homodimer. In consequence, the E-box-dependent transcription activity of c-Myc was abrogated. These results suggest that, in addition to its DNA replication activity, CDC6 also has a role as a transcriptional suppressor of c-Myc. (+info)
Mlx, a new Max-like bHLHZip family member: the center stage of a novel transcription factors regulatory pathway?
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The Myc proto-oncogene family members have been identified as the cellular homologs of the transforming oncogene of avian retroviruses. They encode central regulators of mammalian cell proliferation and apoptosis, and they associate with the bHLHZip protein Max to bind specific DNA sequences and regulate the expression of genes important for cell cycle progression. The other family members, Mad1, Mxi1, Mad3, Mad4 and Rox (Mnt) antagonize their activities. The Mads and Rox compete with Myc in heterodimerizing with Max and in binding to the same specific target sequences. These Mads:Max and Rox:Max dimers repress transcription through binding to the mSIN3 corepressor protein and by tethering histone deacetylase-containing complexes to the DNA. In a screen for Rox interactors we isolated Mlx, a bHLHZip protein previously identified in a screen for Mad1 interactors. In the present work we extend the known dimerization partners of Mlx by demonstrating its ability to interact with Rox. Moreover, we show that contrary to previous reports Mlx is able to homodimerize and to bind E-box sequences at low concentration levels. The possible role of Mlx in an emerging regulatory pathway and acting parallel to the Max driven network is discussed. (+info)
The Drosophila PAR domain protein 1 (Pdp1) gene encodes multiple differentially expressed mRNAs and proteins through the use of multiple enhancers and promoters.
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Transcription factors are often expressed at several times and in multiple tissues during development and regulate diverse sets of downstream target genes by varying their combinatorial interactions with other transcription factors. The Drosophila Tropomyosin I (TmI) gene is regulated by a complex of proteins within the enhancer that synergistically interacts with MEF2 to activate TmI transcription as muscle cells fuse and differentiate. One of the components of this complex is PDP1 (PAR domain protein 1), a basic leucine zipper transcription factor that is highly homologous to three vertebrate genes that are members of the PAR domain subfamily. We have isolated and describe here the structure of the Pdp1 gene. The Pdp1 gene is complex, containing at least four transcriptional start sites and producing at least six different mRNAs and PDP1 isoforms. Five of the PDP1 isoforms differ by the substitution or insertion of amino acids at or near the N-terminal of the protein. At least three of these alternately spliced transcripts are differentially expressed in different tissues of the developing embryo in which PDP1 expression is correlated with the differentiation of different cell types. A sixth isoform is produced by splicing out part of the PAR and basic DNA binding domains, and DNA binding and transient transfection experiments suggest that it functions as a dominant negative inhibitor of transcription. Furthermore, two enhancers have been identified within the gene that express in the somatic mesodermal precursors to body wall muscles and fat body and together direct expression in other tissues that closely mimics that of the endogenous gene. These results show that Pdp1 is widely expressed, including in muscle, fat, and gut precursors, and is likely involved in the transcriptional control of different developmental pathways through the use of differentially expressed PDP1 isoforms. Furthermore, the similarities between Pdp1 and the other PAR domain genes suggest that Pdp1 is the homologue of the vertebrate genes. (+info)
Protein kinase-A dependent phosphorylation of transcription enhancer factor-1 represses its DNA-binding activity but enhances its gene activation ability.
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The cAMP-dependent signaling pathway has been implicated in cardiac cell growth/differentiation and muscle gene transcription. Previously, we have identified a cAMP-inducible E-box/M-CAT hybrid motif in the cardiac alpha-myosin heavy chain (alpha-MHC) gene promoter. The two factors, TEF-1 and Max, that bind to this motif are found to physically associate with each other and exert a positive cooperative effect for gene regulation. Here we show that TEF-1, but not Max, is a substrate for protein kinase-A (PK-A)-dependent phosphorylation. TEF-1 is phosphorylated by PK-A at residue serine-102. This post-translational modification of TEF-1 repressed its DNA-binding activity, but not its ability to interact with the Max protein. Replacement of serine-102 in TEF-1 by a neutral or a charged amino acid did not abolish its DNA-binding ability, suggesting that changing a charge at the 102 amino-acid position of TEF-1 was not sufficient to inhibit its DNA-binding activity. We also show that PK-A response of the alpha-MHC gene is stimulated by the presence of wild-type TEF-1 but not by mutant TEF-1 having serine-102 replaced by alanine, suggesting that phosphorylation at this residue accounts for the cAMP/PK-A response of the gene. Thus, these data demonstrate that TEF-1 is a direct target of cAMP/PK-A signaling in cardiac myocytes. (+info)