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

Type 2 Diabetes is a metabolic disorder associated with insulin resistance and consequent high blood glucose levels. Under normal conditions, in response to high blood glucose levels, pancreatic beta cells produce insulin. The secreted insulin is dis-tributed to tissues thereby stimulating insulin stimulated glucose uptake. However, maximum glucose disposal takes place in skeletal muscle. Thus, studying beta cells and skeletal muscle in respect to diabetes is crucial. Protein Phosphatase 2A (PP2A) is one of the major serine/threonine phosphatases belonging to PhosphoProteinPhospha-tase (PPP) family. It constitutes about 80% of all serine/threonine phosphatases. It is regulated by numerous regulatory subunits as well as other substrate molecules and post translational modifications. This alters their localization, activity and its target molecules. Many evidences show the effect of insulin on PP2Ac and its abnormal reg-ulation in conditions of glucolipotoxicity. Thus, studying PP2Ac interaction partners in respect to type 2 diabetes will give insight into its role in insulin resistance.

Here, we studied interaction partners of PP2Ac in both beta cells and human skeletal muscle. INS-1 832/13 insulin secreting cells are used to study beta cell which are treated with basal and high glucose for 48hrs which are then harvested and ana-lyzed. Skeletal muscle biopsies are collected from human subjects. Two biopsies are collected from each individual, basal and insulin stimulated using hyperinsulenemic euglycemic clamp technique. We collected biopsies from individuals characterized in three different groups, lean controls, obese/overweight insulin resistant, and type 2 diabetics. Both beta cells and human skeletal muscle biopsies are analyzed using a sim-ilar proteomics approach using ESI-HPLC-MS/MS. Using this technique, we identi-fied 514 partners in INS-1 832/13 cells with 89 partners classified as glucose respon-sive. Similarly, 211 interaction partners are identified in human skeletal muscle biop-sies and 69 proteins presented a significant difference among three gropus. Several important PP2Ac interaction partners were identified which included some known partners (identified in other cell types) as well. Many proteins involved in insulin se-cretion are found as PP2Ac partners in beta cells whereas several vital molecules in-volved in insulin signaling pathway are identified in skeletal muscle biopsies. Some important molecules like Rac1, Limk1, Akt2, MAPK are identified among others. Pro-teins that effect PP2Ac post translational modification, such as PPME-1, are also identified and presented with a significant change. Further validation of these partners will help with a better understanding of the role and regulation of PP2Ac in diabetes.

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