Approximately 23.6 million Americans (7.8% of the population) have diabetes - most needing to produce more insulin because of insulin resistance, yet nearly all with pancreatic ?-cell dysfunction. In ?-cells, insulin synthesis begins with the precursor, preproinsulin, which must undergo co-translational translocation into the endoplasmic reticulum (ER), signal peptide (SP) cleavage, and downstream proinsulin folding. These earliest events are critical to insulin biosynthesis, but they are relatively understudied. Over the past three years, four preproinsulin SP mutations have been reported to cause human diabetes that make investigation of these earliest events especially timely. While it is known that insulin haploinsufficiency does not cause diabetes, patients with preproinsulin SP mutations are heterozygotes, suggesting that mutants act in a dominant- negative fashion. The molecular mechanisms of ?-cell failure caused by these mutants remain unknown. Interestingly, diabetes phenotypes associated with the SP mutants ranges from severe neonatal-onset diabetes caused by A(SP24)D, to mild adult-onset diabetes associated with R(SP6)C or H. I hypothesize that these two classes of SP mutants cause ?-cell failure through two distinct mechanisms. In one case, I propose that inefficient co-translational translocation [of R(SP6)C] causes cytosolic accumulation of untranslocated mutant that is slowly toxic to ? -cells, leading to adult-onset diabetes that may be akin to the pathogenesis of Alzheimer's and some other neurodegenerative diseases. In the other case, I propose that failed SP cleavage [of A(SP24)D] disturbs downstream proinsulin folding, causing ER retention of the mutant that abnormally interacts with co-expressed wild-type (WT) proinsulin and blocks its ER exit in trans, decreasing insulin production and initiating severe insulin-deficient diabetes in early life. This proposal aims to better understand the coordination of the earliest events of insulin biosynthesis and define the molecular mechanisms of ?-cell failure caused by defects of those events. The ultimate goal is to develop novel strategies to prevent development of diabetes caused by misfolded (pre)proinsulin.
Three Specific Aims are proposed: 1) To examine coordination of preproinsulin co-translational translocation, SP cleavage, and downstream proinsulin folding;2) To define genetic manipulations that would allow WT proinsulin to escape from blockade caused by mutant (pre)proinsulins;3) To identify small molecules that could prevent ?-cell failure caused by mutant preproinsulins. Accumulating evidence suggests that ER stress and proinsulin misfolding plays a role in the pathogenesis of the most common form of diabetes (type 2) which does not involve any preproinsulin coding sequence mutations. These new diabetogenic mutants, in which underlying (pre)proinsulin mishandling-misfolding is unequivocal, are ideal models for understanding molecular mechanisms of ?-cell failure, and for testing experimental therapies aiming at preventing diabetes.

Public Health Relevance

To make bioactive insulin in pancreatic beta cells, the precursor preproinsulin must undergo a series of sequential processing events. Defects in these early processing events have been reported to cause diabetes in humans. This proposal aims to elucidate the molecular pathogenesis of diabetes linked to such defects, and to develop novel strategies to prevent pancreatic beta cell failure.

Agency
National Institute of Health (NIH)
Institute
National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK)
Type
Research Project (R01)
Project #
5R01DK088856-03
Application #
8460916
Study Section
Membrane Biology and Protein Processing (MBPP)
Program Officer
Haft, Carol R
Project Start
2011-08-01
Project End
2016-04-30
Budget Start
2013-05-01
Budget End
2014-04-30
Support Year
3
Fiscal Year
2013
Total Cost
$262,601
Indirect Cost
$93,726
Name
University of Michigan Ann Arbor
Department
Internal Medicine/Medicine
Type
Schools of Medicine
DUNS #
073133571
City
Ann Arbor
State
MI
Country
United States
Zip Code
48109
Liu, Ming; Wright, Jordan; Guo, Huan et al. (2014) Proinsulin entry and transit through the endoplasmic reticulum in pancreatic beta cells. Vitam Horm 95:35-62
Guo, Huan; Xiong, Yi; Witkowski, Piotr et al. (2014) Inefficient translocation of preproinsulin contributes to pancreatic ? cell failure and late-onset diabetes. J Biol Chem 289:16290-302
Alejandro, Emilyn U; Gregg, Brigid; Wallen, Taylor et al. (2014) Maternal diet-induced microRNAs and mTOR underlie ? cell dysfunction in offspring. J Clin Invest 124:4395-410
Haataja, Leena; Snapp, Erik; Wright, Jordan et al. (2013) Proinsulin intermolecular interactions during secretory trafficking in pancreatic ? cells. J Biol Chem 288:1896-906
Liu, Ming; Lara-Lemus, Roberto; Shan, Shu-ou et al. (2012) Impaired cleavage of preproinsulin signal peptide linked to autosomal-dominant diabetes. Diabetes 61:828-37