Mammalian target of rapamycin (mTOR), a central component of a signaling pathway that coordinates cell growth with nutrient availability, is a promising target for the treatment of diabetes and obesity. Mounting evidence supports that mTOR signaling plays a crucial role in the development of the metabolic diseases, but it has not yet been possible to obtain a clear understanding of the mechanism underlying mTOR-mediated pathogenesis of the diseases. The objective of this application is to identify proteins that transmit mTOR signals downstream to the key nuclear processes involved in nutrient metabolism and gene expression. The central hypothesis for the proposed research is that mTOR triggers a cascade of cellular processes that lead to nuclear localization of proteins that activate or repress expression of important metabolic or regulatory genes. The rationale for the proposed research is that, once the nuclear factors that shuttle between nucleus and cytoplasm under the control of mTOR are discovered, a significant insight will be obtained into the mechanism responsible for mTOR-regulated nutrient metabolism and pathogenesis of metabolic diseases.
The specific aims are: (1) To systematically identify mTOR-regulated nuclear proteins in adipocytes. The research will focus on the identification of nuclear proteins whose nuclear localization is regulated by mTOR signaling in adipocytes, the cells derived from the adipose tissue that governs a whole body energy homeostasis using a novel quantitative proteomics approach; (2) To systematically identify mTOR-regulated DNA-binding proteins in adipocytes.
The aim of this study will focus on the identification of mTOR-regulated DNA-binding proteins in adipocytes, which will be achieved through quantitative proteomics approach coupled with whole chromatin isolation; (3) To determine functions of identified mTOR-regulated nuclear proteins. ~20 proteins will be selected among proteins identified in the aims #1 and #2, and their sub-nuclear localization and functions will be analyzed through biochemical and genetic approaches, which include the use of the YFP protein and RNAi as well as biochemical and reporter gene assay for glucose and fat metabolism. The research proposed in this application is significant, because it will allow us to obtain the new knowledge on the mechanism that is responsible for connecting mTOR activity to the nuclear events involving the regulation of metabolic gene expression, and therefore it will provide crucial regulatory proteins that we can target for the treatment of metabolic diseases.
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