Biomedical subjects

Kenneth L Wright

Publications and source records attributed to Kenneth L Wright.

9 recordsLinked to original sources

PubMed2006

Epigenetic regulation of MHC-II and CIITA genes.

This review describes recent advances in understanding how epigenetic events control MHC-class-II-family (MHC-II) gene expression. To address this issue, two phases of gene transcription have to be considered. First, the control of MHC-II by chromatin-modifying events such as histone acetylation, methylation, deacetylation, ubiquitination and the interplay between these different epigenetic events will be examined. The interactions of chromatin-modifying enzymes with class II transactivator (CIITA) and relevant DNA-binding proteins for activating and silencing MHC-II gene transcription will be reviewed. Second, the transcriptional control of the promoter of CIITA, the master regulator of MHC-II, by DNA methylation and chromatin modification will be discussed, and the novel role of noncoding RNA will be explored. Finally, the relevance of these findings to infection, transplantation and cancer will be reviewed.

Animals↗

PubMed2006

Positive regulatory domain I-binding factor 1 mediates repression of the MHC class II transactivator (CIITA) type IV promoter.

MHC class II transactivator (CIITA), a co-activator that controls MHC class II (MHC II) transcription, functions as the master regulator of MHC II expression. Persistent activity of the CIITA type III promoter (pIII), one of the four potential promoters of this gene, is responsible for constitutive expression of MHC II by B lymphocytes. In addition, IFN-gamma induces expression of CIITA in these cells through the type IV promoter (pIV). Positive regulatory domain 1-binding factor 1 (PRDI-BF1), called B lymphocyte-induced maturation protein 1 (Blimp-1) in mice, represses the expression of CIITA pIII in plasma and multiple myeloma cells. To investigate regulation of CIITA pIV expression by PRDI-BF1 in the B lymphocyte lineage, protein/DNA-binding studies, and functional promoter analyses were performed. PRDI-BF1 bound to the IFN regulatory factor-element (IRF-E) site in CIITA pIV. Ectopic expression of either PRDI-BF1 or Blimp-1 repressed this promoter in B lymphocytes. In vitro binding and functional analyses of CIITA pIV demonstrated that the IRF-E is the target of this repression. In vivo genomic footprint analysis demonstrated protein binding at the IRF-E site of CIITA pIV in U266 myeloma cells, which express PRDI-BF1. PRDI-BF1beta, a truncated form of PRDI-BF1 that is co-expressed in myeloma cells, also bound to the IRF-E site and repressed CIITA pIV. These findings demonstrate for the first time that, in addition to silencing expression of CIITA pIII in B lymphocytes, PRDI-BF1 is capable of binding and suppressing CIITA pIV.

B-Lymphocytes↗

PubMed2005

Identification of the ankyrin repeat proteins ANKRA and RFXANK as novel partners of class IIa histone deacetylases.

Eighteen human histone deacetylases (HDACs) have been identified, and according to their sequence similarity to yeast homologs, these enzymes are grouped into distinct classes. Within class II, HDAC4, HDAC5, HDAC7, and HDAC9 share similar domain organization both within the N-terminal extension and the C-terminal catalytic domain, thus forming a subclass known as class IIa. These HDACs function as signal-responsive transcriptional corepressors. To gain further insight into their function and regulation, we utilized an N-terminal fragment of HDAC4 as bait in yeast two-hybrid screens, which uncovered myocyte enhancer factor 2C, 14-3-3zeta, and ankyrin repeat family A protein (ANKRA). ANKRA is a poorly characterized protein with an ankyrin repeat domain similar to RFXANK, a subunit of the trimeric transcription factor RFX. Mutations on genes of the RFX subunits and the coactivator CIITA are responsible for the bare lymphocyte syndrome, an immunodeficiency disorder attributed to the lack of major histocompatibility complex class II (MHCII) antigens. Through its ankyrin repeat domain, RFXANK interacted with HDAC4. Two RFXANK-binding sites were found on HDAC4 with one located within residues 118-279 and another within residues 448-666. Interestingly, this deacetylase also interacted with CIITA. Consistent with the physical interaction with RFXANK and CIITA, HDAC4 and homologs repressed MHCII expression. These results identify ANKRA, RFXANK, and CIITA as novel targets of class IIa HDACs and suggest that these deacetylases play a role in regulating MHCII expression.

14-3-3 Proteins↗

PubMed2005

Expression of the MHC class II transactivator (CIITA) type IV promoter in B lymphocytes and regulation by IFN-gamma.

The MHC class II transactivator (CIITA), the master regulator of MHC class II (MHC II) expression, is a co-activator that controls MHC II transcription. Human B lymphocytes express MHC II constitutively due to persistent activity of CIITA promoter III (pIII), one of the four potential promoters (pI-pIV) of this gene. Although increases in MHC II expression in B cells in response to cytokines have been observed and induction of MHC II and CIITA by IFN-gamma has been studied in a number of different cell types, the specific effects of IFN-gamma on CIITA expression in B cells have not been studied. To investigate the regulation of CIITA expression by IFN-gamma in B cells, RT-PCR, in vivo and in vitro protein/DNA binding studies, and functional promoter analyses were performed. Both MHC II and CIITA type IV-specific RNAs increased in human B lymphocytes in response to IFN-gamma treatment. CIITA promoter analysis confirmed that pIV is IFN-gamma inducible in B cells and that the GAS and IRF-E sites are necessary for full induction. DNA binding of IRF-1 and IRF-2, members of the IFN regulatory factor family, was up-regulated in B cells in response to IFN-gamma and increased the activity of CIITA pIV. In vivo genomic footprint analysis demonstrated proteins binding at the GAS, IRF-E and E box sites of CIITA pIV. Although CIITA pIII is considered to be the hematopoietic-specific promoter of CIITA, these findings demonstrate that pIV is active in B lymphocytes and potentially contributes to the expression of CIITA and MHC II in these cells.

B-Lymphocytes↗

PubMed2005

Role of Stat3 in regulating p53 expression and function.

Loss of p53 function by mutation is common in cancer. However, most natural p53 mutations occur at a late stage in tumor development, and many clinically detectable cancers have reduced p53 expression but no p53 mutations. It remains to be fully determined what mechanisms disable p53 during malignant initiation and in cancers without mutations that directly affect p53. We show here that oncogenic signaling pathways inhibit the p53 gene transcription rate through a mechanism involving Stat3, which binds to the p53 promoter in vitro and in vivo. Site-specific mutation of a Stat3 DNA-binding site in the p53 promoter partially abrogates Stat3-induced inhibition. Stat3 activity also influences p53 response genes and affects UV-induced cell growth arrest in normal cells. Furthermore, blocking Stat3 in cancer cells up-regulates expression of p53, leading to p53-mediated tumor cell apoptosis. As a point of convergence for many oncogenic signaling pathways, Stat3 is constitutively activated at high frequency in a wide diversity of cancers and is a promising molecular target for cancer therapy. Thus, repression of p53 expression by Stat3 is likely to have an important role in development of tumors, and targeting Stat3 represents a novel therapeutic approach for p53 reactivation in many cancers lacking p53 mutations.

Animals↗

PubMed2004

PRDI-BF1 recruits the histone H3 methyltransferase G9a in transcriptional silencing.

PRDI-BF1, the human ortholog of mouse Blimp-1, is a DNA-binding protein involved in postinduction repression of interferon-beta gene transcription in response to viral infection. PRDI-BF1 also has an essential function in driving terminal differentiation of B lymphocytes and therein silences multiple genes. Here we show PRDI-BF1 assembles silent chromatin over the interferon-beta promoter in the osteosarcoma cell line U2OS through recruitment of the histone H3 lysine methyltransferase G9a. G9a is recruited only when in a complex with PRDI-BF1. G9a catalytic activity is required for the accumulation of methylated histone H3 and transcriptional silencing mediated by PRDI-BF1 in vivo. This establishes a mechanism for the recruitment of G9a, the main mammalian euchromatic methyltransferase, and defines nonembryonic targets of G9a.

Amino Acid Sequence↗

PubMed2003

Identification of a functionally impaired positive regulatory domain I binding factor 1 transcription repressor in myeloma cell lines.

B cell differentiation into a plasma cell requires expression of the positive regulatory domain zinc finger protein 1 gene (PRDM1) that encodes the positive regulatory domain I binding factor 1 (PRDI-BF1 or Blimp-1) protein. It represses the transcription of specific target genes, including c-myc, the MHC class II trans-activator, Pax-5, and CD23b. In this study we demonstrate the presence of an alternative protein product of the PRDM1 gene. The new protein, PRDI-BF1 beta, has a disrupted PR domain and lacks the amino-terminal 101 aa of the originally described protein. PRDI-BF1 beta has a dramatic loss of repressive function on multiple target genes, but maintains normal DNA-binding activity, nuclear localization, and association with histone deacetylases and deacetylase activity. Myeloma cell lines express the highest levels of PRDM1 beta mRNA relative to the full-length form, while primary cells and several other cell lines have very low, but detectable, levels of PRDM1 beta. RNA analysis and analysis of the PRDM1 promoters demonstrate that PRDI-BF1 beta is generated from the same gene by alternative transcription initiation using an internal promoter. These newly described features of the PRDM1 gene are highly analogous to the PRDM2 (RIZ) and PRDM3 (MDS1-EVI1) genes, in which each express a truncated protein missing the PR domain. The expression of each of the truncated proteins is elevated in cancerous cells and may play an important role in the disease.

Alternative Splicing↗

PubMed2002

Regulation and specificity of MHC2TA promoter usage in human primary T lymphocytes and cell line.

Although activated human T cells express MHC class II antigens, the regulation of these antigens in T cells is poorly understood. This study focuses on the control of the MHC2TA gene in these cells. MHC2TA encodes the transcriptional master regulator of MHC class II, the class II trans-activator (CIITA). It has at least three distinct promoters (PI, PIII, and PIV), each active in an overlapping subset of cell types and directing a slightly different product. This report used highly purified blood T cells prepared by negative immunoselection to analyze CIITA. Real-time PCR analysis indicates that resting T cells do not express detectable CIITA transcript, while activated T cells express the PIII CIITA form. Transient transfection of activated blood T cells using wild-type and mutant PIII promoter-reporter constructs shows that two promoter elements, activation response element-1 (ARE-1) and ARE-2, are important for PIII function. cAMP response element binding protein, a known activator of gene expression in activated T cells, activates PIII in primary T cells. However, an intact ARE-2 site is not required for this activation, indicating that cAMP response element binding protein does not activate via this site. EMSAs indicate that an activating transcription factor/cAMP response element binding protein/cAMP response element modulator family member, but not phosphorylated cAMP response element binding protein-1, binds to ARE-2. ARE-2 also forms a complex with an unidentified protein. The ARE-2 binding protein is constitutively expressed in a DR(+) T cell line, reflecting differences between the DR(+) cell line and primary blood lymphocytes. These results show that MHC2TA PIII is induced in activated T lymphocytes, and that the induced binding of ARE-2 is a crucial step in this process.

Acyltransferases↗

PubMed2002

Constitutive Stat3 activity up-regulates VEGF expression and tumor angiogenesis.

Non-receptor and receptor tyrosine kinases, such as Src and EGF receptor (EGFR), are major inducers of vascular endothelial growth factor (VEGF), one of the most potent mediators of angiogenesis. While tyrosine kinases signal through multiple pathways, signal transducer and activation of transcription 3 (Stat3) is a point of convergence for many of these and is constitutively activated with high frequency in a wide range of cancer cells. Here, we show that VEGF expression correlates with Stat3 activity in diverse human cancer cell lines. An activated Stat3 mutant (Stat3C) up-regulates VEGF expression and stimulates tumor angiogenesis. Stat3C-induced VEGF up-regulation is abrogated when a Stat3-binding site in the VEGF promoter is mutated. Furthermore, interrupting Stat3 signaling with dominant-negative Stat3 protein or Stat3 antisense oligonucleotide in tumor cells down-regulates VEGF expression. Consistent with an important role of Stat3 in VEGF up-regulation induced by various oncogenic tyrosine kinases, v-Src-mediated VEGF expression is inhibited when Stat3 signaling is blocked. Moreover, chromatin immunoprecipitation assays indicate that Stat3 protein binds to the VEGF promoter in vivo and mutation of a Stat3-binding site in the VEGF promoter abrogates v-Src-induced VEGF promoter activity. These studies provide evidence that the VEGF gene is regulated directly by Stat3 protein, and indicate that Stat3 represents a common molecular target for blocking angiogenesis induced by multiple signaling pathways in human cancers.

3T3 Cells↗