How mutations in proinsulin cause diabetes: a protein-misfolding disease Insulin plays a central role in the regulation of vertebrate metabolism. The hormone, the post-translational product of a single-chain precursor (proinsulin), is a globular protein containing two chains, A (21 residues) and B (30 residues). Recent advances in human genetics have identified dominant negative mutations in the insulin gene causing permanent neonatal-onset diabetes mellitus (DM). The objective of this interdisciplinary application is to investigate the biochemical, structural, and cell-biological mechanisms of this syndrome as a model disease of protein misfolding. We hypothesize that the clinical mutations block folding of the precursor in the endoplasmic reticulum (ER) of pancreatic ?-cells. Structural analysis of the mutant proinsulins will provide insight into native determinants of foldability. Although expression of the wild-type allele would in other circumstances be sufficient to maintain homeostasis, studies of a corresponding mouse model motivate the hypothesis that the misfolded variant perturbs wild-type biosynthesis through formation of non-native aggregates containing both wild-type and mutant polypeptides. Impaired ?-cell secretion is associated with ER stress, distorted organelle architecture, and eventual cell death. To test this central hypothesis and to define the structural bases of pathological misfolding, a team has been assembled at CWRU, University of Michigan, and University of Chicago to bring to bear the combined power of biochemistry, biophysics, structural biology, cell biology, and transgenic mouse models. This collaborative proposal thus offers the exciting possibility of deciphering the molecular basis of a human disease of protein misfolding. Although neonatal diabetes is uncommon, the proposed contribution of proinsulin misfolding and ER stress to the mechanism of ?-cell dysfunction in the metabolic syndrome and type 2 diabetes mellitus extends the significance of this application to diverse human populations.

Public Health Relevance

Our goal is to determine the molecular bases of toxic proinsulin misfolding in the pathogenesis of neonatal-onset diabetes mellitus due to mutations in the insulin gene. An interdisciplinary strategy is proposed that integrates structural biology with biochemistry, synthetic chemistry, and cell biology. The results promise to eludidate the structural principles of native disulfide pairing in the biosynthesis of insulin.

Agency
National Institute of Health (NIH)
Institute
National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK)
Type
Research Project (R01)
Project #
5R01DK069764-08
Application #
8448597
Study Section
Molecular and Cellular Endocrinology Study Section (MCE)
Program Officer
Sechi, Salvatore
Project Start
2004-09-30
Project End
2015-03-31
Budget Start
2013-04-01
Budget End
2014-03-31
Support Year
8
Fiscal Year
2013
Total Cost
$443,546
Indirect Cost
$117,437
Name
Case Western Reserve University
Department
Biochemistry
Type
Schools of Medicine
DUNS #
077758407
City
Cleveland
State
OH
Country
United States
Zip Code
44106
Cui, Jingqiu; Chen, Wei; Sun, Jinhong et al. (2015) Competitive Inhibition of the Endoplasmic Reticulum Signal Peptidase by Non-cleavable Mutant Preprotein Cargos. J Biol Chem 290:28131-40
Avital-Shmilovici, Michal; Whittaker, Jonathan; Weiss, Michael A et al. (2014) Deciphering a molecular mechanism of neonatal diabetes mellitus by the chemical synthesis of a protein diastereomer, [D-AlaB8]human proinsulin. J Biol Chem 289:23683-92
Avital-Shmilovici, Michal; Mandal, Kalyaneswar; Gates, Zachary P et al. (2013) Fully convergent chemical synthesis of ester insulin: determination of the high resolution X-ray structure by racemic protein crystallography. J Am Chem Soc 135:3173-85
Wright, Jordan; Birk, Julia; Haataja, Leena et al. (2013) Endoplasmic reticulum oxidoreductin-1α (Ero1α) improves folding and secretion of mutant proinsulin and limits mutant proinsulin-induced endoplasmic reticulum stress. J Biol Chem 288:31010-8
Weiss, Michael A (2013) Diabetes mellitus due to the toxic misfolding of proinsulin variants. FEBS Lett 587:1942-50
Kiselar, Janna G; Datt, Manish; Chance, Mark R et al. (2011) Structural analysis of proinsulin hexamer assembly by hydroxyl radical footprinting and computational modeling. J Biol Chem 286:43710-6
Liu, Ming; Haataja, Leena; Wright, Jordan et al. (2010) Mutant INS-gene induced diabetes of youth: proinsulin cysteine residues impose dominant-negative inhibition on wild-type proinsulin transport. PLoS One 5:e13333
Luisier, Samuel; Avital-Shmilovici, Michal; Weiss, Michael A et al. (2010) Total chemical synthesis of human proinsulin. Chem Commun (Camb) 46:8177-9
Liu, Ming; Hua, Qing-xin; Hu, Shi-Quan et al. (2010) Deciphering the hidden informational content of protein sequences: foldability of proinsulin hinges on a flexible arm that is dispensable in the mature hormone. J Biol Chem 285:30989-1001
Phillips, Nelson B; Wan, Zhu-li; Whittaker, Linda et al. (2010) Supramolecular protein engineering: design of zinc-stapled insulin hexamers as a long acting depot. J Biol Chem 285:11755-9

Showing the most recent 10 out of 15 publications