Faculty Bibliography

PURPOSE: The HP1 family of evolutionarily conserved proteins regulates heterochromatin packaging, in addition to a less defined role in the regulation of euchromatic genes. To examine the possible role of HP1 proteins in fetal prostate development and prostate cancer the protein expression of HP1alpha, beta and gamma was evaluated in human archival tissue. MATERIALS AND METHODS: Tissue sections from human prostate cancer and fetal prostate were examined using antibodies against HP1 isoforms to evaluate HP1 modulation in cancer and development. Western blot analysis of HP1 proteins was also performed in extracts of cultured prostate cancer cells. RESULTS: HP1alpha, beta and gamma are differentially regulated in various cellular compartments in prostate development. HP1alpha is not expressed at 14 or 24 weeks of prostate development but it is expressed in adult prostate tissue. HP1beta is highly expressed at 14 and 24 weeks, and it appears predominantly in epithelial cells compared to HP1gamma, which is expressed at equal levels in epithelial and stromal cells. All 3 HP1 isoforms show altered expression in prostate cancer compared to that in normal adult prostate tissue. CONCLUSIONS: HP1 proteins are tightly regulated during prostate development. In the adult prostate HP1alpha, beta and gamma antibodies detect high levels of HP1 antigen in a contiguous layer of epithelial cells. However, the detection of HP1 in prostate cancer ranges from undetectable to inconsistent staining of noncontiguous epithelial cells

c-Jun is a component of the heterodimeric transcription factor AP-1 that is rapidly activated in response to ultraviolet light (UV). In unstressed cells, c-Jun activity is negatively regulated by transcriptional repressor complexes. Here we show that the F-box protein Fbl10/JHDM1B interacts with c-Jun and represses c-Jun-mediated transcription. Chromatin-immunoprecipitation assays demonstrate that Fbl10 is present at the c-jun promoter, and that c-Jun is required for the recruitment of Fbl10. Fbl10 binds to the unmethylated CpG sequences in the c-jun promoter through the CxxC zinc finger and tethers transcriptional repressor complexes. Suppression of Fbl10 expression by RNA interference (RNAi) induces transcription of c-jun and other c-Jun-target genes, and causes an aberrant cell-cycle progression and increased UV-induced cell death. Furthermore, Fbl10 protein and messenger RNA are downregulated in response to UV in an inverse correlation with c-Jun. Taken together, our results demonstrate that Fbl10 is a key regulator of c-Jun function

TBP-associated factor 4 (TAF4), an essential subunit of the TFIID complex acts as a coactivator for multiple transcriptional regulators, including Sp1 and CREB. However, little is known regarding the structural properties of the TAF4 subunit that lead to the coactivator function. Here, we report the crystal structure at 2.0-A resolution of the human TAF4-TAFH domain, a conserved domain among all metazoan TAF4, TAF4b, and ETO family members. The hTAF4-TAFH structure adopts a completely helical fold with a large hydrophobic groove that forms a binding surface for TAF4 interacting factors. Using peptide phage display, we have characterized the binding preference of the hTAF4-TAFH domain for a hydrophobic motif, DPsiPsizetazetaPsiPhi, that is present in a number of nuclear factors, including several important transcriptional regulators with roles in activating, repressing, and modulating posttranslational modifications. A comparison of the hTAF4-TAFH structure with the homologous ETO-TAFH domain reveals several critical residues important for hTAF4-TAFH target specificity and suggests that TAF4 has evolved in response to the increased transcriptional complexity of metazoans

Huntington's disease (HD) is an inherited neurodegenerative disease caused by a glutamine repeat expansion in huntingtin protein. Transcriptional deregulation and altered energy metabolism have been implicated in HD pathogenesis. We report here that mutant huntingtin causes disruption of mitochondrial function by inhibiting expression of PGC-1alpha, a transcriptional coactivator that regulates several metabolic processes, including mitochondrial biogenesis and respiration. Mutant huntingtin represses PGC-1alpha gene transcription by associating with the promoter and interfering with the CREB/TAF4-dependent transcriptional pathway critical for the regulation of PGC-1alpha gene expression. Crossbreeding of PGC-1alpha knockout (KO) mice with HD knockin (KI) mice leads to increased neurodegeneration of striatal neurons and motor abnormalities in the HD mice. Importantly, expression of PGC-1alpha partially reverses the toxic effects of mutant huntingtin in cultured striatal neurons. Moreover, lentiviral-mediated delivery of PGC-1alpha in the striatum provides neuroprotection in the transgenic HD mice. These studies suggest a key role for PGC-1alpha in the control of energy metabolism in the early stages of HD pathogenesis

Kaposi's sarcoma-associated herpesvirus (KSHV) maintains a latent infection in primary effusion lymphoma cells but can be induced to enter full lytic replication by exposure to a variety of chemical inducing agents or by expression of the KSHV-encoded replication and transcription activator (RTA) protein. During latency, only a few viral genes are expressed, and these include the three genes of the so-called latency transcript (LT) cluster: v-FLIP (open reading frame 71 [ORF71]), v-cyclin (ORF72), and latency-associated nuclear antigen (ORF73). During latency, all three open reading frames are transcribed from a common promoter as part of a multicistronic mRNA. Subsequent alternative mRNA splicing and internal ribosome entry allows for the expression of each protein. Here, we show that transcription of LT cassette mRNA can be induced by RTA through the activation of a second promoter (LT(i)) immediately downstream of the constitutively active promoter (LT(c)). We identified a minimal cis-regulatory region, which overlaps with the promoter for the bicistronic K14/v-GPCR delayed early gene that is transcribed in the opposite direction. In addition to a TATA box at -30 relative to the LT(i) mRNA start sites, we identified three separate RTA response elements that are also utilized by the K14/v-GPCR promoter. Interestingly, LT(i) is unresponsive to sodium butyrate, a potent inducer of lytic replication. This suggests there is a previously unrecognized class of RTA-responsive promoters that respond to direct, but not indirect, induction of RTA. These studies highlight the fact that induction method can influence the precise program of viral gene expression during early events in reactivation and also suggest a mechanism by which RTA contributes to establishment of latency during de novo infections

Huntington's disease (HD) is an inherited neurodegenerative disease caused by expansion of a polyglutamine tract in the huntingtin protein. Transcriptional dysregulation has been implicated in HD pathogenesis. Here, we report that huntingtin interacts with the transcriptional activator Sp1 and coactivator TAFII130. Coexpression of Sp1 and TAFII130 in cultured striatal cells from wild-type and HD transgenic mice reverses the transcriptional inhibition of the dopamine D2 receptor gene caused by mutant huntingtin, as well as protects neurons from huntingtin-induced cellular toxicity. Furthermore, soluble mutant huntingtin inhibits Sp1 binding to DNA in postmortem brain tissues of both presymptomatic and affected HD patients. Understanding these early molecular events in HD may provide an opportunity to interfere with the effects of mutant huntingtin before the development of disease symptoms

Type I interferon (IFN) stimulates transcription through a heteromeric transcription factor that contains tyrosine-phosphorylated STAT2. We show that STAT2 recruits histone acetyltransferases (HAT) through its transactivation domain, resulting in localized transient acetylation of histones. GCN5, but not p300/CBP or PCAF, is required for STAT2 function. However, GCN5 function is impaired by the transcriptional antagonist, adenovirus E1A oncoprotein. The TFIID component TAFII130 potentiates STAT2 function, but TAFII28 or the HAT activity of TAFII250 do not, and transcriptional induction can proceed independently of the TATA-binding protein, TBP. Moreover, IFN-stimulated transcription was resistant to poliovirus-targeted degradation by TBP, and continued despite host-cell transcriptional shutoff during poliovirus infection. We conclude that a non-classical transcriptional mechanism combats an anticellular action of poliovirus, through a TBP-free TAF-containing complex and GCN5

The Sonic hedgehog (Shh)-Gli signaling pathway regulates development of many organs, including teeth. We cloned a novel gene encoding a transcription factor that contains a zinc finger domain with highest homology to the Gli family of proteins (61-64% amino acid sequence identity) from incisor pulp. Consistent with this sequence conservation, gel mobility shift assays demonstrated that this new Gli homologous protein, GliH1, could bind previously characterized Gli DNA binding sites. Furthermore, transfection assays in dental pulp cells showed that whereas Gli1 induces a nearly 50-fold increase in activity of a luciferase reporter containing Gli DNA binding sites, coexpression of Gli1 with Gli3 and/or GliH1 results in inhibition of the Gli1-stimulated luciferase activity. In situ hybridization analysis of mouse embryos demonstrated that GliH1 expression is initiated later than the three Gli genes and has a more restricted expression pattern. GliH1 is first detected diffusely in the limb buds at 10.0 days post coitus and later is expressed in the branchial arches, craniofacial interface, ventral part of the tail, whisker follicles and hair, intervertebral discs, teeth, eyes and kidney. LacZ was inserted into the GliH1 allele in embryonic stem cells to produce mice lacking GliH1 function. While this produced indicator mice for GliH1-expression, analysis of mutant mice revealed no discernible phenotype or required function for GliH1. A search of the Celera Genomics and associated databases identified possible gene sequences encoding a zinc finger domain with approximately 90% homology to that of GliH1, indicating there is a family of GliH genes and raising the possibility of overlapping functions during development