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A DDX5 S480A polymorphism is associated with increased transcription of fibrogenic genes in hepatic stellate cells.

ABSTRACT: We recently identified a missense single nucleotide polymorphism (SNP) in DDX5 (rs1140409, p.S480A) that enhances the risk of developing cirrhosis. DDX5 is an ATP-dependent RNA helicase and transcriptional modulator. We hypothesized that the activity of DDX5 in regulating fibrogenic gene transcription in hepatic stellate cells (HSCs) is altered by the S480A SNP. To test this, we employed two approaches: 1) transient overexpression of DDX5 cDNA or siRNA knockdown of endogenous DDX5, with replacement by either DDX5 wild type (WT) or SNP cDNA, or 2) stable expression of exogenous DDX5 WT and SNP in HSC lines. WT DDX5 mRNA in HSCs was inversely correlated with gene expression for alpha2(I) collagen, tissue inhibitor of metalloproteinase-1, and transforming growth factor-beta1. Stable DDX5 SNP-expressing cells had higher basal and transforming growth factor-beta1-stimulated expression and enhanced promoter activities of fibrogenic genes. DDX5 variant-expressing cells also had higher Smad3 and AP-1-responsive reporter activities. In a one-hybrid GAL4 system, co-expression of the DDX5 SNP variant with chimeras of GAL4 DNA binding domain linked to JunD or Sp1 displayed higher transactivation of a GAL4-responsive reporter than that of DDX5 WT. Increased fibrogenic gene expression in DDX5 SNP-expressing cells was associated with reduced recruitment of DDX5 homodimers to responsive promoters, but there was no difference in the recruitment of the co-repressor HDAC1 (histone deacetylase 1). These data suggest that DDX5 is a repressor of fibrogenic genes in HSCs through interaction with transcriptional complexes. The enhanced fibrogenic activity of the DDX5 risk variant is linked to a reduced repressive function toward these target genes.

SUBMITTER: Guo J

PROVIDER: S-EPMC2820771 | biostudies-literature | 2010 Feb

REPOSITORIES: biostudies-literature

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A DDX5 S480A polymorphism is associated with increased transcription of fibrogenic genes in hepatic stellate cells.

Guo Jinsheng J Hong Feng F Loke Johnny J Yea Steven S Lim Chooi Ling CL Lee Ursula U Mann Derek A DA Walsh Martin J MJ Sninsky John J JJ Friedman Scott L SL

The Journal of biological chemistry 20091217 8

We recently identified a missense single nucleotide polymorphism (SNP) in DDX5 (rs1140409, p.S480A) that enhances the risk of developing cirrhosis. DDX5 is an ATP-dependent RNA helicase and transcriptional modulator. We hypothesized that the activity of DDX5 in regulating fibrogenic gene transcription in hepatic stellate cells (HSCs) is altered by the S480A SNP. To test this, we employed two approaches: 1) transient overexpression of DDX5 cDNA or siRNA knockdown of endogenous DDX5, with replacem ...[more]

PMID: 20022962

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Project description:Background & aimsMethyl-CpG binding protein 2, MECP2, which binds to methylated regions of DNA to regulate transcription, is expressed by hepatic stellate cells (HSCs) and is required for development of liver fibrosis in mice. We investigated the effects of MECP2 deletion from HSCs on their transcriptome and of phosphorylation of MECP2 on HSC phenotype and liver fibrosis.MethodsWe isolated HSCs from Mecp2-/y mice and wild-type (control) mice. HSCs were activated in culture and used in array analyses of messenger RNAs and long noncoding RNAs. Kyoto Encyclopedia of Genes and Genomes pathway analyses identified pathways regulated by MECP2. We studied mice that expressed a mutated form of Mecp2 that encodes the S80A substitution, MECP2S80, causing loss of MECP2 phosphorylation at serine 80. Liver fibrosis was induced in these mice by administration of carbon tetrachloride, and liver tissues and HSCs were collected and analyzed.ResultsMECP2 deletion altered expression of 284 messenger RNAs and 244 long noncoding RNAs, including those that regulate DNA replication; are members of the minichromosome maintenance protein complex family; or encode CDC7, HAS2, DNA2 (a DNA helicase), or RPA2 (a protein that binds single-stranded DNA). We found that MECP2 regulates the DNA repair Fanconi anemia pathway in HSCs. Phosphorylation of MECP2S80 and its putative kinase, HAS2, were induced during transdifferentiation of HSCs. HSCs from MECP2S80 mice had reduced proliferation, and livers from these mice had reduced fibrosis after carbon tetrachloride administration.ConclusionsIn studies of mice with disruption of Mecp2 or that expressed a form of MECP2 that is not phosphorylated at S80, we found phosphorylation of MECP2 to be required for HSC proliferation and induction of fibrosis. In HSCs, MECP2 regulates expression of genes required for DNA replication and repair. Strategies to inhibit MECP2 phosphorylation at S80 might be developed for treatment of liver fibrosis.

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A DDX5 S480A polymorphism is associated with increased transcription of fibrogenic genes in hepatic stellate cells.

Publications

A DDX5 S480A polymorphism is associated with increased transcription of fibrogenic genes in hepatic stellate cells.

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