All ETDs from UAB

Advisory Committee Chair

Scott M Wilson

Advisory Committee Members

Lucas Pozzo-Miller

Erik D Roberson

Michael Brenner

Lori L McMahon

Document Type

Dissertation

Date of Award

2015

Degree Name by School

Doctor of Philosophy (PhD) Heersink School of Medicine

Abstract

Loss of the deubiquitinating enzyme (DUB) USP14 in the ataxia (axJ) mice leads to altered neuromuscular junction (NMJ) structure, reduced synaptic transmission at the NMJ, and decreased mobility. However, the types of processes that USP14 regulates in the nervous system remain unclear. Because association with the proteasome stimulates USP14's ubiquitin hydrolase activity, it is thought to act primarily on proteasomal substrates. Therefore, one way for USP14 to support nervous system function is by modulating protein turnover. While a number of studies done in yeast and immortalized cell lines demonstrate that loss or inhibition of USP14 alters proteasome function, there is no consensus on whether inhibition of USP14 accelerates or delays protein degradation. A second possibility is that USP14's primary role is the maintenance of ubiquitin homeostasis. The proteasome-associated DUBs remove ubiquitin from substrates prior to their degradation and indeed, loss of USP14 leads to ubiquitin depletion in both yeast and the axJ mice. These two suggestions assume that, in keeping with its association with the proteasome, USP14 acts on proteasome-targeting ubiquitin chains. However, in the following pages, we demonstrate that USP14 is required to disassemble proteasome-independent ubiquitin chains and regulate kinase activation in the nervous system. Specifically, USP14's ubiquitin hydrolase activity regulates the ubiquitination of mixed linages kinase 3 (MLK3) to control the activation of its downstream target c-Jun N-terminal kinase (JNK), and over-activation of JNK contributes to the alteration of NMJ structure and motor function observed in a transgenic mouse expressing a dominant-negative, catalytically-inactive USP14 species in the nervous system. Moreover, we show that increasing the amount of ubiquitin conjugates in the nervous system alters motor function and muscle development even when not accompanied by depletion of free ubiquitin pools. These studies underline the importance of tightly regulated ubiquitin signaling at the NMJ.

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