Type 2 diabetes (T2D) is characterized by an abnormal elevation of blood glucose levels resulting from defects in secretion and action of insulin and the incretins. The two incretin hormones glucose-dependent insulinotropic polypeptide (GIP) and glucagon-like peptide-1 (GLP-1) are secreted from intestinal K and L cells, respectively, in response to nutrient ingestion. Once secreted, GIP and GLP-1 act at pancreatic Beta-cells to enhance glucose-stimulated insulin secretion (GSIS). Although GLP-1-based agents are currently used for the treatment of T2D, significant lapses in our molecular mechanistic understanding of GLP-1 secretion from the intestine and GLP-1 action at the Beta-cell remain. Toward this, the involvement of a new signaling axis involving Cdc42-Pak1 proteins in regulating insulin-induced GLP-1 secretion in vitro was recently revealed. Intriguingly, this pathway resembled the Cdc42-Pak1 pathway our lab discovered in 2007 to regulate second-phase insulin secretion from the pancreatic islet beta cell. However, it remains untested as to whether GLP-1 interfaces with the Cdc42-Pak1 signaling pathway in the Beta-cell to augment insulin secretion. Moreover, insulin is not a canonical GLP-1 secretagogue, such that the physiological relevance of this pathway in the intestinal L cell remains unclear. Thus, the objective of this application is to define the role of the Cdc42-Pak1 signaling pathway in the incretin effect (incretin secretion plus incretin action). I hypothesize that the Cdc42-Pak1 pathway is required for both the secretion of GLP-1 induced by glucose and the enhancement of GSIS by GLP-1, and defects in this signaling pathway contribute to the aberrant glucose homeostasis characterized by T2D. This hypothesis will be tested in three Specific Aims: 1) Establish the requirement of the Cdc42-Pak1 pathway in glucose-induced GLP-1 secretion, 2) Elucidate how GLP-1 interfaces with the Cdc42-Pak1 pathway to potentiate GSIS in Beta-cells, and 3) Delineate the in vivo role of Pak1 signaling in the incretin effect, using a new model of T2D, the Pak1[+/-] heterozygous knockout mouse. Preliminary data show a paucity of Pak1 protein in islets from type 2 diabetic human patients, supporting the physiological relevance of these studies and use of the heterozygous Pak1 knockout mouse. Experimental methods will include siRNA-mediated knockown in intestinal L cells and islet beta cells with ?rescue? strategies, confocal and TIRF microscopy techniques to examine insulin granule mobilization and F-actin remodeling, Rho GTPase activation assays, paired with in vivo and ex vivo analyses of GLP-1 release and incretin-induced insulin secretion. Importantly, my work will provide crucial missing molecular mechanistic data, with future use towards accomplishing my long-term goal of manipulating the incretin system for therapies that more accurately mimic endogenous incretin and insulin secretion to safely restore glucose homeostasis in diabetic patients.

Public Health Relevance

The prevalence of type 2 diabetes has reached pandemic proportions. The current treatments fail to mimic physiological conditions and have many side-effects. In this proposal, I will research a novel cellular pathway (Cdc42-Pak1) in the intestine and pancreas that may have promise as a better drug target for type 2 diabetes.

Agency
National Institute of Health (NIH)
Institute
National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK)
Type
Postdoctoral Individual National Research Service Award (F32)
Project #
5F32DK094488-02
Application #
8334737
Study Section
Special Emphasis Panel (ZDK1-GRB-9 (O1))
Program Officer
Castle, Arthur
Project Start
2011-09-06
Project End
2014-09-05
Budget Start
2012-09-06
Budget End
2013-09-05
Support Year
2
Fiscal Year
2012
Total Cost
$49,214
Indirect Cost
Name
Indiana University-Purdue University at Indianapolis
Department
Pediatrics
Type
Schools of Medicine
DUNS #
603007902
City
Indianapolis
State
IN
Country
United States
Zip Code
46202
Yoder, Stephanie M; Dineen, Stacey L; Wang, Zhanxiang et al. (2014) YES, a Src family kinase, is a proximal glucose-specific activator of cell division cycle control protein 42 (Cdc42) in pancreatic islet ? cells. J Biol Chem 289:11476-87
Ramalingam, Latha; Yoder, Stephanie M; Oh, Eunjin et al. (2014) Munc18c: a controversial regulator of peripheral insulin action. Trends Endocrinol Metab 25:601-8
Kalwat, Michael A; Yoder, Stephanie M; Wang, Zhanxiang et al. (2013) A p21-activated kinase (PAK1) signaling cascade coordinately regulates F-actin remodeling and insulin granule exocytosis in pancreatic ? cells. Biochem Pharmacol 85:808-16