Identification and characterization of a novel interaction between peroxisome-proliferator activated receptor gamma (PPARγ) and mesoderm induction-early response: implications for adipogenesis

Parsons, Amanda (2008) Identification and characterization of a novel interaction between peroxisome-proliferator activated receptor gamma (PPARγ) and mesoderm induction-early response: implications for adipogenesis. Masters thesis, Memorial University of Newfoundland.

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Abstract

PPARγ is a nuclear hormone receptor and master regulator of lipid metabolism and adipogenesis. PPARγ regulates these processes through the recruitment of a diverse set of transcriptional coregulators in a tissue and time specific manner. This study focused on characterizing the interaction between PPARγ and mesoderm induction early response 1 (MIER1), a transcription factor that has been shown to interact with other nuclear hormone receptors and regulate target gene expression. Glutathione S transferase pull-down assays have revealed that PPARγ interacts with both MIER1α and β through the common SANT domain. Coimmunoprecipitations in HEK-293 (human embryonic kidney cells) confirmed that this interaction occurs in vivo. A transcriptional reporter assay using luciferase regulated by the PPAR response element (PPRE) demonstrated that MIER1α and β cause a ligand-independent 2-fold activation of PPAR-driven transcriptional activity and this was similar to the 3-fold activation observed with a known PPARγ coactivator (PGC1-α). Thus, MIER1 interacts with PPAR in a ligand-independent manner through its SANT domain, and activates PPARγ -mediated transcriptional activity. PCR analysis showed that MIER1 mRNA expression is upregulated in 3T3-L1 pre-adipocytes during their differentiation into adipocytes. As well, immunocytochemistry revealed that MIER1α is highly expressed specifically in differentiated 3T3-L1s. Future work will determine the exact role of MIER1 in adipogenesis, using shRNA to knockdown MIER1α in the well-established 3T3-L1 differentiation system.

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