Secretory Pathway Protein Degradation Maintains Insulin Biogenesis + Secretion - ABSTRACT Pancreatic insulin mass (a combination of ß-cell mass and function in the production and storage of insulin) is linked to blood glucose regulation. Normal islet ß-cells contain more than a billion insulin molecules per cell. To maintain this amount involves the synthesis of ≥ 6,000 new molecules per second. Insulin synthesis begins with the translocation of preproinsulin into the endoplasmic reticulum (ER) triggering the formation of proinsulin, which must fold in order to advance through the secretory pathway. Things get “tricky” because not all newly-made proinsulin folds properly. Improperly-folded proinsulin cannot pass ER quality control for export from the ER. Further, misfolded proinsulin has a propensity to associate with bystander proinsulin molecules, forming non-native protein complexes that can propagate the misfolding. Because failed proinsulin molecules do not undergo the forward trafficking needed to become mature insulin, these unsuccessful molecules must be degraded to limit the amount of proinsulin misfolding that can predispose to diabetes. In the last cycle of this grant, our laboratories provided incontrovertible evidence that misfolded proinsulin molecules are targeted for intracellular degradation, and this function is essential for both normal insulin production and the health and survival of pancreatic ß-cells. This competing continuation provides a deeper dive, at the molecular level, to explore three essential secretory pathway quality control mechanisms: ER-Associated Degradation (ERAD); ER-autophagy (ER-to-lysosome, known as ER-phagy); and delivery of (pre)proinsulin into the ER. The three tightly collaborative investigators (Qi, Tsai, Arvan) are experts in these processes and they share highly successful interactions. Specifically, in recent years, the Arvan lab identified the contribution of proteasomal degradation to the maintenance of steady state proinsulin levels in ß-cells, while the Qi lab identified the importance of Sel1L-HRD1 ERAD in maintenance of pancreatic insulin mass [and crucially in additional studies, identified that the ER-based Sigma-1 Receptor (product of the SIGMAR1 / ΣR1 gene) is an endogenous ERAD substrate — upon diminished ERAD, ΣR1 (and other key gene products) are impacted to regulate ER homeostasis]. Curiously, we find that when ERAD capacity or proteasomal clearance is insufficient, proinsulin levels in ß-cells do not rise even though proinsulin is also an ERAD substrate — rather, these levels actually fall, and enhanced ER-phagy is one of the key contributors to this phenotype. Further, our preliminary data (and research plan) highlight perturbed preproinsulin translocation in addition to ΣR1-dependent upregulation of ER-phagy — demonstrating that ERAD crosstalk intersects with several key elements of ß-cell ER function. Thus, we offer a strong rationale for studying ER quality control mechanisms that are linked to development/progression, and possible treatment, of pancreatic ß-cell failure.
