Emma Mastroianni
Pronouns: She/her
Research Mentor(s): Anoop Arunagiri
Research Mentor School/College/Department: Int. Med / Medicine
Program:
Authors: Emma Mastroianni, Anoop Arunagiri, Peter Arvan
Session: Session 4: 1:40 pm – 2:30 pm
Poster: 11
Abstract
Diabetes is caused when the pancreas cannot synthesize a hormone called insulin that regulates blood glucose. Insulin originates in the pancreatic beta cells from its precursor protein called proinsulin. When proinsulin fails to form three intramolecular disulfide bonds in their structure correctly, they oligomerize via intermolecular disulfide links and further larger aggregates develop that cannot be exported out of the endoplasmic reticulum (ER) to their proper destination in the beta cells to produce insulin (Arunagiri et al, Proinsulin misfolding is an early event in the progression to type 2 diabetes, Elife 2019 Jun 11:8:e44532). In the present work, we aimed to determine the correlation between proinsulin misfolding and beta cell ER function. Proinsulin misfolding can be a result of increased protein synthesis, decreased degradation, or impaired forward trafficking. To increase proinsulin synthesis, INS-1E beta cells were exposed to different concentrations of nutrients viz. glucose or glutamine. Additionally, Brefeldin-A was applied to the cells which prevent protein translocation from ER to the Golgi apparatus. Pharmacological inhibition of PERK kinase activity in beta cells, another established approach in our laboratory to cause proinsulin misfolding, was also applied in the current study. All of these conditions resulted in moderate to severe proinsulin folding impediments and ER homeostasis defects. It is also known that one or more of these conditions would cause ER stress. We postulate that improving the oxidative environment of the beta cells ER would lead to proper folding and trafficking of proinsulin. To test this hypothesis, we allowed overexpression of an ER oxidoreductase ERO-1? in beta cells (by transfection) or acutely treated the cells with a strong oxidant, Diamide. A western blotting approach, developed in our lab, was then used to determine the quantity of proinsulin aggregates formed. Lower exogenous expression of ERO-1 alpha appeared to improve proinsulin folding, while Diamide treatment caused a decrease in nonnative monomers and disulfide-linked dimers. The ER redox towards a more oxidative state seemed favorable to proinsulin folding. Insulin biosynthesis and secretion as a result of these treatments would be worthy of monitoring next. Based on the literature and previous work done in the lab, we surmise the ER chaperone protein BiP, ER-resident oxidoreductases, and ER-associated protein degradation (or ERAD) are all potential targets for improving proinsulin folding and ER homeostasis.
