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Journal of Biochemistry Advance Access originally published online on February 14, 2008
Journal of Biochemistry 2008 143(5):675-683; doi:10.1093/jb/mvn019
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© 2008 The Japanese Biochemical Society.

Transcriptional Regulation of the Human PNRC Promoter by NFY in HepG2 Cells

Yan Zhang, Bin Chen, Yuping Li, Jian Chen, Guiyu Lou, Min Chen and Dujin Zhou*

Department of Biochemistry and Molecular Biology, Third Military Medical University, Chongqing 400038, China

*To whom correspondence should be addressed. Tel: +86 2368752940, Fax: +86 23 68752940, E-mail: dujinzhou{at}yahoo.com

Received January 18, 2008; Accepted February 7, 2008


  Abstract

PNRC (proline-rich nuclear receptor co-activator) was previously identified using bovine SF-1 (steroidogenic factor 1) as the bait in a yeast two-hybrid screening of a human mammary gland cDNA expression library. PNRC has been demonstrated to be a novel co-activator for multiple nuclear receptors. To understand the molecular mechanisms that regulate the expression of human PNRC gene, in this study, potential transcriptional start site was determined by 5' RACE analysis. Functional analysis of the 5' flanking region of the human PNRC gene by deletion mutagenesis, transient transfection and luciferase assays revealed that the –123/+27 region is the minimal promoter of the human PNRC gene. Within this promoter region, there is one putative binding site for the transcription factor NFY (nuclear factor Y). EMSA and ChIP analyses demonstrated the specific binding of NFY protein to the human PNRC promoter. Transient transfection and luciferase assays further revealed that over-expression of NFY represses promoter activity of PNRC gene in a dose-dependent manner. These results indicate that the transcription factor NFY specifically binds to promoter region of PNRC and negatively regulates the transcription of the human PNRC gene.

Key Words: nuclear receptor co-activator, NFY, PNRC, promoter analysis, transcriptional regulation

Abbreviations: ChIP, chromatin immunoprecipitaion; EMSA, electrophoresis mobility shift and super-shift assay; NFY, nuclear factor Y; PNRC, proline-rich nuclear receptor co-activator; RACE, rapid amplification of cDNA ends; SF1, steroidogenic factor 1; SH3, Src homology domain-3


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