Unknown,Transcriptomics,Genomics,Proteomics

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Transcription profiling of human heat shocked HeLa cell line with siRNA knockdown of HSF1

ABSTRACT: Although HSF1 is known to play an important role in regulating the cellular response to proteotoxic stressors, little is known about the structure and function of the HSF1 signaling network under both stressed and unstressed conditions. In this study, we used a combination of chromatin immunoprecipitation (ChIP) microarray analysis and time course gene expression microarray analysis with and without siRNA-mediated inhibition of HSF1 comprehensively identify genes directly and indirectly regulated by HSF1 and examine the structure of the extended HSF1 signaling network. Correlation between promoter binding and gene expression was not significant for all genes bound by HSF1 suggesting that HSF1 binding per se is not sufficient for expression. However, the correlation with promoter binding was significant for genes identified as HSF1-regulated following siRNA knockdown allowing the identification of direct transcriptional targets of HSF1. Among promoters bound by HSF1 following heat shock, a gene ontology (GO) analysis showed significant enrichment only in categories related to protein folding. In contrast, analysis of the extended HSF1 signaling network showed enrichment in a variety of categories related to protein folding, anti-apoptosis, RNA splicing, ubiquitination and others, highlighting a complex transcriptional program directly and indirectly regulated by HSF1.,SUBMITTER_CITATION: Genome-wide analysis of human HSF1 signaling reveals a transcriptional program linked to cellular adaptation and survival Authors: Todd J. Page, Devanjan Sikder, Longlong Yang, Linda Pluta, Russell D. Wolfinger, Thomas Kodadek, and Russell S. Thomas Journal: Molecular Biosystems 2:627-639

ORGANISM(S): Homo sapiens

SUBMITTER: Todd Page

PROVIDER: E-GEOD-3697 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Project description:Mutations in the RMRP gene are the origin of cartilage-hair hypoplasia. Cartilage-hair hypoplasia is associated with severe dwarfism caused by impaired skeletal development. However, it is not clear why mutations in the RMRP gene lead to skeletal dysplasia. Viperin is a known substrate of RMRP. Since chondrogenic differentiation of the growth plate is required for development of the long bones, we hypothesized that viperin functions as a chondrogenic regulator downstream of RMRP. Viperin protein is expressed throughout the stages of chondrogenic differentiation in vivo. Viperin gene expression is increased during knockdown of Rmrp RNA in the ATDC5 model for chondrogenic differentiation. Viperin is expressed during ATDC5 chondrogenic differentiation. Viperin knockdown reduces, while viperin overexpression increases overall protein secretion, with CXCL10 identified as a potential target via mass spectrometry-proteomics. CXCL10 protein expression is reduced during knockdown and increased during overexpression of viperin and CXCL10 protein expression coincides with viperin expression in ATDC5 chondrogenic differentiation. Viperin knockdown induces, while viperin overexpression reduces TGFβ activity. Furthermore, viperin knockdown conditioned media increases, while viperin overexpression conditioned media reduces chondrogenic differentiation of ATDC5 cells. TGFβ target genes Pai1 and Smad7 are increased during knockdown and reduced during overexpression of viperin. Moreover, TGFβ activity is reduced when differentiating ATDC5 cells are exposed to CXCL10 and, acting as a viperin overexpression mimic, CXCL10 similarly reduces chondrogenic differentiation of ATDC5. Lastly, we show that in CHH patient cells, RMRP expression is reduced and viperin expression is increased, coinciding with reduced chondrogenic differentiation and increased CXCL10 expression, possibly explaining the CHH phenotype. Together our data show that viperin may play a pivotal role in chondrogenic differentiation, with potential consequences for cartilage-hair hypoplasia pathobiology.

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Transcription profiling of human heat shocked HeLa cell line with siRNA knockdown of HSF1

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