Post-transcriptional Regulation of PPAR-g Expression by 5'-Untranslated Regions

California State University Northridge, Northridge CA

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Abstract

PPAR-g is a member of the nuclear receptor family of transcription factors and is known to regulate[unreadable] many different genes with diverse physiological functions. The presence of a total of seven PPAR-g[unreadable] transcript isoforms has previously been demonstrated in monkey and human macrophages. Most of the[unreadable] variability between different PPAR-g transcripts is in the 5'-untranslated region (5'-UTR), such that the[unreadable] seven transcripts encode for only 3 different protein isoforms.[unreadable] Recently, 5'-UTRs have emerged as major modulators of cytoplasmic mRNA processing in eukaryotic[unreadable] cells. Such post-transcriptional regulation allows for rapid adjustments in protein expression in response to[unreadable] various stimuli. Based on experimental evidence, we hypothesize that sequence variations in the 5' UTR of[unreadable] PPAR-g transcripts may regulate mRNA stability ortranslational efficiency.[unreadable] The proposed studies focus on identifying mechanisms for the post-transcriptional regulation of PPARg[unreadable] expression by PPAR-g 5'-UTRs. The translation of different Lentivirus-derived PPAR-g transcript isoforms[unreadable] will be compared in THP-1 macrophages. A requirement for any macrophage-specific or ligand-induced[unreadable] factors will also be ascertained. Effect of PPAR-g 5'-UTR on translational efficiency will be investigated by[unreadable] in-vitro and in-vivo translation of full-length PPAR-g transcripts and of chimeric constructs of different PPARg[unreadable] 5'-UTR cloned upstream of the luciferase reporter gene. The stability (half-life) and decay rates of PPARg[unreadable] transcripts with different 5'-UTRs will be determined in the presence of transcription inhibitors by RT-PCR,[unreadable] Northern blot analysis and pulse-chase radiolabeling experiments. The presence of PPAR-g 5'-UTRspecific[unreadable] cytosolic RNA-binding proteins will be identified by electrophoretic mobility shift assays, UV crosslinking[unreadable] and affinity chromatography. The proposed studies will explain the biological significance of multiple[unreadable] PPAR-g transcripts and facilitate the use of PPAR-g 5'-UTR as targets for specific therapeutic outcomes.[unreadable] Relevance to Public Health: PPAR-g are implicated in many human diseases including[unreadable] atherosclerosis, diabetes, obesity and certain cancers. Information about posttranscriptional regulation of[unreadable] PPAR-g function is vital for efficient and selective modulation of different PPAR-g isoforms.