Date of Award

Fall 2014

Degree Type

Thesis

Degree Name

Master of Science (MS)

Department

Biochemistry

First Advisor

Clint Chapple

Committee Member 1

Scott Briggs

Committee Member 2

Joe Kappock

Committee Member 3

Maureen McCann

Abstract

The Mediator complex is a multi-protein co-regulator of eukaryotic transcription which plays a role in the expression of many, if not most, genes of the cell. Two Mediator subunits, REF4 and RFR1, were demonstrated to be important for the normal regulation of phenylpropanoid metabolism in Arabidopsis. Phenylpropanoids are a family of specialized plant metabolites derived from the amino acid phenylalanine and are involved in defense against pathogens, UV protection and structural support. In order to understand how Mediator regulates phenylpropanoid metabolism through REF4 and RFR1, a better basic understanding of this protein complex is required. Here I provide data on the initial steps in characterizing the Arabidopsis Mediator complex. First, I performed partial purification of Mediator using ion-exchange chromatography followed by LC-MS analysis of cation-exchange purified fraction. Then we generated and evaluated antibodies against specific Mediator subunits. Lastly, I used yeast two-hybrid to evaluate previously identified putative interactors of REF4, evaluated interaction between Mediator tail subunits with REF4 and RFR1, and identified new interaction candidates for RFR1.

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