Date Written

4-28-2019

Document Type

Distinction Paper

Degree Name

Biochemistry and Molecular Biology-BS

Department

Biochemistry and Molecular Biology

Advisor

Dr. David Sheridan, Ph.D.

First Committee Member

Dr. Jennifer Bennett, Ph.D.

Second Committee Member

Dr. David Robertson, Ph.D.

Keywords

Calcium Channels, Glucocorticoid Receptor, Cortisol, Stress

Subject Categories

Biochemistry, Biophysics, and Structural Biology | Molecular Biology

Abstract

L-type calcium channels couple membrane depolarization to muscle contraction and aid in signal conduction between cells. Cav1.3 calcium channel isoform is expressed in neurons and endocrine cells. Nervous and endocrine tissues also express both Glucocorticoid (GR) and Mineralocorticoid receptors (MR) that are responsible for binding serum cortisol to induce immediate and long term changes within the cell. While neurons co-express both GRs and MRs, GR activation occurs in response to high concentrations of cortisol. There is a direct correlation between GR activation and the amplitude of calcium currents (Champeau, 2007). To determine potential interactions between the GR and the calcium channel isoforms, we will co-express a GFP-tagged GR, purchased from Addgene, and a mRubyC1-tagged L-type calcium channel in a simplified, non-native cellular system, Chinese Hamster Ovary (CHO) cells. Specifically targeted primers were designed to clone the Cav1.3 sequence in preparation for its insertion into the mRubyC1 red fluorescent vector. The Cav1.3+mRubyC1 construct was then transfected into CHO cells expressing GFP-tagged GR. Injection of a high concentration of cortisol into the cellular environment will activate the GR receptor, and fluorescence capture imaging will determine if there is an in vitro interaction between Cav1.3 and GR.

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