Abstract Dr. T. Hayes
Title of Project p107 Function on 3T3-L1 Differentiation
Abstract Obesity is a risk factor in many of the major chronic diseases that afflict our society, including coronary artery disease, diabetes mellitus, and cancer. The ability to regulate the cell cycle and differentiation of adipocytes is key in the development and physiology of obesity and also in the origin of cancer. Understanding these processes is critical to a rational approach to the treatment of obesity and cancer. 3T3-L1 cells are a well-studied model system for the differentiation of adipocytes. Quiescent 3T3-L1 cells express high levels of the Rb-related protein p130 but little if any p107. Over the first 24 hours after they are stimulated to differentiate, the expression of p107 increases to a high level while that of p130 drops. p107 and p130 return to their former levels as the cells complete differentiation. Data with protein kinase inhibitors suggest that the rapid induction of p107 is critical for differentiation but is not necessary for proliferation. Members of the Rb family function through specific interactions with a variety of proteins. The specific protein interactions that allow p107 to play its crucial role in the differentiation of preadipocytes will be determined. Biochemistry and mass spectrometry will be used to identify p107-interacting proteins from nuclear extracts of differentiating 3T3-L1 cells. The expression of these proteins will be knocked down using siRNA. This will allow us to identify the proteins whose interactions with p107 are critical for 3T3-L1 differentiation. The structural basis for the interaction of these critical proteins with p107 will be determined using deletions and targeted mutagenesis of these proteins and p107. This will enable us to investigate the role that post-translational modifications play in the regulation of these interactions. Finally, the role that these interactions play in the regulation of other genes involved in the differentiation of preadipocytes will be determined using microarray analysis on siRNA knock-down cells and comparing the results to exisiting microarray data from differentiating 3T3-L1 cells. This will enable the identification of the precise step in differentiation and cell cycle exit at which p107 acts in 3T3-L1 cells.
Updated 10/31/2005
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