Biomedical Sciences ETDs

Clark Bird

Publication Date

12-1-2012

Abstract

The KH-type splicing regulatory protein (KSRP) promotes the decay of AU-rich element (ARE) containing mRNAs. Although KSRP is expressed in the developing and mature nervous system, very little is known about its role in regulating gene expression in the brain. In this study, we utilized in vitro binding and decay studies to examine whether KSRP regulates the stability of the GAP-43 transcript, an ARE-containing neuronal mRNA whose protein product plays a role in axonal growth and synaptic plasticity. We found KSRP destabilizes GAP-43 mRNA by binding to the GAP-43 ARE, a process that depends on the presence of the fourth KH domain in the protein. Furthermore, KSRP competed with another GAP-43 mRNA binding protein, the stabilizing factor HuD, for binding to these ARE sequences. Given that GAP-43 expression is crucial for accurate axonal outgrowth during neuronal development, we also examined the functional consequences of KSRP overexpression and depletion on the axonal outgrowth from primary hippocampal neurons. Overexpression of either full length KSRP or KSRP without the nuclear localization signal hindered axonal outgrowth in these cultures, while overexpression of a mutant protein without the KH4 domain did not have any effect. In contrast, depletion of KSRP led to a dramatic increase in axonal length. Concurrent overexpression of GAP-43 and KSRP rescued the axonal outgrowth deficits seen with KSRP overexpression, but only when the GAP-43 mRNA was targeted to axons using GAP-43 or amphoterin 3 UTR sequences. Together, our results suggest that KSRP is an important regulator of GAP-43 mRNA stability and neuronal differentiation that works in direct opposition to HuD.

Keywords

GAP-43, KSRP, Neuron outgrowth

Sponsors

NIH Grant NS30255-18

Document Type

Dissertation

Language

English

Degree Name

Biomedical Sciences

Level of Degree

Doctoral

Department Name

Biomedical Sciences Graduate Program

First Committee Member (Chair)

Valenzuela, Fernando

Second Committee Member

Caldwell, Kevin

Third Committee Member

Hartley, Rebecca

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