41-OR: Roles of Transcription Factor EB (TFEB) in ß-Cell Function and Survival Under Stress

Abstract

Autophagy can help protect stressed β-cells. We investigated if disruption of Transcription Factor EB (Tfeb), a transcriptional master-regulator of lysosomes and autophagy, contributes to β-cell failure and death under various ER stress-inducing conditions. Short-term β-cell autophagy-deficiency (inducible Atg5 knockout) reduced β-cell survival under hypoxic stress in vitro and impaired the ability of syngeneic marginal mass islet grafts to maintain glucose homeostasis in streptozotocin (STZ)-induced diabetic mice, with >80% of recipients of Atg5-deficient islets returning to diabetes within 10 days post-transplant (vs. 30% of controls, p=0.05). Hypoxic culture activated pro-apoptotic ER stress pathways, and confocal analysis of islet cells from LC3-RFP-EGFP autophagy reporter mice demonstrated that autophagic flux was interrupted. Hypoxia-induced impairment of autophagy was associated with reduced lysosomal cathepsin B activity and a significant loss of Tfeb, both of which could be partially rescued by Torin-1. Lipotoxicity and thapsigargin-induced ER-stress similarly reduced Tfeb protein and cell viability, suggesting ER stress-specific mechanisms may drive dysregulation of Tfeb and lysosomes. Conversely, overexpression of Tfeb:GFP reduced thapsigargin-induced death in MIN6 cells. To directly study the consequences of Tfeb loss we generated mice with Ins1Cre-driven, β-cell-specific, Tfeb deletion (TfebβKO). Notably, both male and female TfebβKO mice had normal glucose tolerance, but cultured Tfeb KO β-cells showed impaired lysosomal cathepsin B activity and TfebβKO mice had worsened glucose tolerance compared to Tfeb flox littermates following injection with multiple low-doses STZ. In summary, autophagy protects β-cells under hypoxia but is impaired under prolonged hypoxic stress, possibly through Tfeb dysregulation. Our data further suggest loss of Tfeb is a common feature of ER stress and that this loss exacerbates stress-induced β-cell failure and death. Disclosure Y. Zou: None. D. Pasula: None. B. Verchere: Advisory Panel; Self; Integrated Nanotherapeutics Inc., Sirona Biochem, Stock/Shareholder; Self; Integrated Nanotherapeutics Inc. D. S. Luciani: None. Funding JDRF (2-2013-50)