Date

7-2013

Document Type

Dissertation

Degree Name

Ph.D.

Department

Dept. of Cell and Developmental Biology

Institution

Oregon Health & Science University

Abstract

Mechanosensation in hair cells is a fascinating and intricately regulated process. Not only are the components of the mechanotransduction complex themselves critical for faithful transduction of mechanical stimuli, but this process also depends on the trafficking of those components to their correct cellular location in hair cells. Correct sorting of proteins within hair cells is important for a number of processes including ion homeostasis. In Chapters 3 and 4 I explore the consequences of the zebrafish mutations in the adaptor protein 1 beta 1 (ap1b1) gene the beta-subunit of the AP-1 clathrin dependent sorting complex on mechanotransduction and regulation of intracellular Na+ concentrations. I show that the basolateral protein, the Na+/K+ATPase pump, is missorted to apical compartments of hair cells in ap1b1 mutants. I also demonstrate that ap1b1 mutants fail to regulate intracellular Na+ levels, likely a consequence of reduced Na+/K+ATPase pump at the basolateral membrane. In Chapters 2 and 5, I investigate the roles of two putative members of the mechanotransduction complex on transduction of mechanical stimuli. In Chapter 2, I show that tmhsa expression in hair cells is necessary for auditory response behaviors in zebrafish. I also show that expression of tmhsa specifically in hair cells in the mutant background is sufficient to rescue the auditory and vestibular phenotypes. Additionally, I demonstrate that the GFP-Tmhsa transgenic construct localizes to the tips of stereocilia in zebrafish hair cells, a characteristic that is consistent with the protein being apart of the transduction complex. In Chapter 5, I provide evidence that zebrafish Tmc2a is involved in hair-cell mechanotransduction. I show that while exogenous expression of the cytoplasmic, N-terminal fragment of viii Tmc2a reduces mechanosensitivity, exogenous expression of the cytoplasmic, Cterminal fragment of Tmc2a increases mechanosensitivity, suggesting that Tmc2a is involved in mechanotransduction, but that the N- and C-terminal domains play different roles in this process.

Identifier

doi:10.6083/M4RR1WJJ

School

School of Medicine

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