Author Date

2023-06-10

Degree Name

Department

Neuroscience

College

Life Sciences

Defense Date

2023-05-31

Publication Date

2023-06-10

First Faculty Advisor

Dr. Arminda Suli

First Faculty Reader

Dr. Jeffery Barrow

Honors Coordinator

Dr. Rebekka Matheson

Keywords

Zebrafish, auxin, protein, degradation, TIR, safe harbor

Abstract

The Auxin-Inducible Degron (AID) system is a protein degradation system naturally occurring in plants. In the presence of the hormone auxin, plant proteins conjugated with the AID degron (a short amino acid sequence which signals for the protein’s degradation) are recruited via an interaction with the protein TIR1, into an E3 ubiquitin ligase protein complex. Following this interaction, AID-tagged proteins are ubiquitinated and sent for degradation. When auxin is metabolized or transported out of the cell through efflux transporters, the transcript of the protein of interest is translated without being degraded, allowing the concentration of the targeted protein to return to normal levels. Several groups have already adapted the AID system to study protein function in different model organisms including yeast, mammalian cell lines, and C. elegans. However, successfully adapting this system to zebrafish has yet to be accomplished. We have shown that co-expression of exogenous AID-tagged GFP and TIR1 within the mechanosensory hair cells (HC) or spinal cord neurons of zebrafish larvae leads to GFP degradation in the presence of auxin. Surprisingly, GFP-AID is degraded at higher rates in some tissues, such as HCs, compared to other tissues, such as spinal cord neurons. In this thesis, I explore some of the possibilities for incomplete functioning of the AID system in the zebrafish model organism which include: insufficient auxin flowthrough, low concentrations of exogenous TIR, and poor integration of designed plasmids into the host genome.