Characterization and functional analysis of Usp14

Abstract

The ubiquitin proteasome system (UPS) is essential for regulated protein degradation, a requirement for numerous neuronal process, including vesicle cycling, neurotransmitter release, spine morphology, and synaptic plasticity. Better understanding of UPS function in neurons will increase our knowledge of neurological diseases caused by alterations in ubiquitin signaling.

I utilized the ataxia (ax^J) mice as a tool to study the UPS in the nervous system. The ax^J mice have lowered expression of ubiquitin-specific protease 14 (Usp14) in all tissues; this decreased expression results from an intracisternal-A particle insertion into Usp14. The ax^J mice are phenotypically indistinguishable from wild type littermates at birth. By 2 weeks, ax^J mice are smaller in size and display a resting tremor. Muscle wasting and tremor become more pronounced until 6 weeks of age, and death occurs by 8 weeks. It was suggested that the ax^J phenotypes are caused by dystrophic Purkinje cell axons. However, the global loss of Usp14 indicates that the ax ^J phenotypes result from multiple organ system failure.

To understand Usp14’s function in neurons, I characterized the expression and activity of Usp14 gene products. Loss of Usp14 results in a decrease in steady state levels of monomeric ubiquitin. Because Usp14’s catalytic activity is stimulated by proteasome binding, loss of Usp14 on the proteasome implies altered proteasome function; evidence of which is strengthened by ubiquitin loss in ax^J mice. Furthermore, I specifically expressed Usp14 in ax^J neurons to test the hypothesis that the ax^J phenotype results from loss of Usp14 expression in these cells. Neuronal-specific expression of Usp14 rescued the premature death, muscle wasting, and tremor independently of Purkinje cells.

Here we demonstrate an essential requirement for Usp14 in the mammalian nervous system. Surprisingly, Usp14 appears dispensable in all tissues except neurons, indicating the UPS may have nervous system specific functions not required in other cell types. Our data also provide the first evidence for a primary defect in proteasome function as the underlying cause of a neurological disease in mammals. Through increasing our understanding of Usp14 in neurons, we can better understand UPS function in other diseases; illuminating new therapeutic targets for neurological disorders.