Date of Award

2019

Document Type

Thesis

Degree Name

Master of Science (M.S.)

Department

Biological Sciences

First Advisor

Douglas Weiser

First Committee Member

Lisa A. Wrischnik

Second Committee Member

Jane Khudyakov

Abstract

Failure to balance synthesis, folding and degradation of secreted proteins results in the accumulation of unfolded proteins in the endoplasmic reticulum, termed ER stress. Cells respond to ER stress by activating a signaling pathway known as the unfolded protein response (UPR). The UPR induces phosphorylation of eIF2α to attenuate global protein translation, allowing the ER to clear misfolded proteins. Opposing this function, eIF2α phosphatases contain a catalytic subunit, Protein Phosphatase 1, and either of two homologous scaffolding subunits, GADD34 and CReP. Inhibition of eIF2α phosphatases has been shown to prolong UPR signaling and promote survival in many cells types and has reduced the progression of neurodegenerative diseases in several mouse models. Despite the clinical significance of GADD34 and CReP, their precise roles in UPR signaling and ER stress-induced apoptosis are largely unknown. Zebrafish are an ideal model for studying eIF2ɑ phosphatases in the UPR due to the efficiency of reverse genetics and the susceptibility of the caudal fin epidermal cells to ER stress-induced apoptosis. In acute ER stress, these cells undergo apoptosis mediated by p63-puma, but under chronic stress, apoptosis is mediated by CHOP, a downstream UPR target. We sought to determine the role of eIF2α phosphatases in both apoptotic programs by inducing eIF2α phosphatase loss- or gain-of-function and measuring the effect on apoptosis in response to ER stress. Inhibition of both eIF2ɑ phosphatases protected cells against both apoptotic responses, primarily through GADD34 inhibition. We speculate that survival is promoted through enhanced eIF2ɑ phosphorylation, stalling the UPR in an early state and preventing accumulation of the late-stage target, CHOP. GADD34 overexpression protected cells against acute apoptosis, which we hypothesize is through a transient promotion of autophagy. Our results indicate that altering the influence of eIF2α phosphatases in the UPR may be a promising therapeutic for diseases exacerbated by ER stress.

Pages

97

Available for download on Friday, June 25, 2021

Included in

Biology Commons

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