Access Type

Open Access Dissertation

Date of Award

January 2017

Degree Type

Dissertation

Degree Name

Ph.D.

Department

Pharmaceutical Sciences

First Advisor

Zhengping Yi

Abstract

Protein kinases play fundamental roles in regulation of biological processes and functions, such as insulin signaling and glucose metabolism. Dysregulation of protein kinases may cause impaired cell signaling and human diseases, such as metabolic syndrome and type 2 diabetes (T2D). Skeletal muscle is the main site responsible for insulin-stimulated glucose disposal, and insulin resistance in skeletal muscle is one of the key features of the pathogenesis of T2D. Therefore, malfunction of protein kinases and their interaction proteins may contribute to the molecular mechanism of insulin resistance in human skeletal muscle. However, no large scale profiling study has been reported to investigate the abnormal active kinases and their interaction partners that might cause skeletal muscle insulin resistance. Here, we present a high throughput platform coupling activity-based ATP probes and quantitative proteomics to globally profile functional kinome and its interactome in human skeletal muscle in 8 lean healthy and 8 obese insulin resistant participants. We identified 54 active protein kinases in human skeletal muscle, which is the largest catalog of experimental determined active kinases to date. Twenty-two out of the 54 active kinases displayed a significant change in insulin resistant obese subjects, including the protein kinases regulate JNK signaling, p38 MAPK signaling, Wnt signaling, AMPK signaling, ERK1/2 signaling and mTOR signaling. In addition, we identified 616 functional kinome interaction partners in human skeletal muscle, which is the largest human kinome interactome to date. Among the 616 interaction partners, 385 has a fold change > 1.5 or <0.67. Out of these 385 kinome interaction partners, 135 displayed significantly difference between lean heathy and insulin resistant obese subjects. The kinome interactome participate in various cell signaling pathways (i.e. diabetes pathway, mTOR pathway, insulin signaling pathway, etc.) and biofunctions (i.e. protein synthesis and degradation, cytoskeleton dynamics, and apoptosis). In summary, we generated the 1st global picture of functional kinome and kinome interactome in skeletal muscle in lean heathy individuals, and how it differs in obese insulin resistant participants. These results provide novel insights into molecular mechanism of skeletal muscle insulin resistance, and may facilitate identifying new targets for treating metabolic diseases.

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