Date of Award

12-2013

Document Type

Dissertation

Degree Name

Doctor of Philosophy (PhD)

Program

Biomedical Sciences

Track

Molecular Therapeutics and Cell Signaling

Research Advisor

Rennolds Ostrom, Ph.D.

Committee

Elizabeth Fitzpatrick, Ph.D. Edwards Park, Ph.D. Steven Tavalin, Ph.D. Christopher Waters, Ph.D.

Abstract

Adenylyl cyclase (AC) catalyzes the formation of the ubiquitous second messenger cAMP. AC isoforms differ in their tissue distribution, cellular localization, regulation, and protein interactions, and most cells express multiple isoforms. We hypothesized that cAMP produced by different AC isoforms regulates unique cellular responses. Overexpression of individual isoforms had distinct effects on forskolin (Fsk)-induced expression of a number of known cAMP-responsive genes in human bronchial smooth muscle cells (BSMC) and human embryonic kidney cells (HEK-293). Most notable, in BSMC overexpression and activation of AC2 enhanced interleukin 6 (IL-6) expression, but overexpression of AC3 or AC6 had no effect. IL-6 production by BSMC was induced by Fsk and select G protein-coupled receptor (GPCR) agonists, though IL-6 levels did not directly correlate with intracellular cAMP levels. At low cAMP concentrations exchange protein directly activated by cAMP (Epac) predominated in mediating the IL-6 response, but at higher cAMP concentrations protein kinase A (PKA) assumed the larger role. IL-6 promoter mutations demonstrated that activator protein 1 (AP-1) and cAMP responsive element (CRE) transcription sites were required for cAMP mediated induction. Our findings indicate that AC2 participates in a cAMP-signaling compartment that specifically regulates IL-6 expression in BSMC and that other AC isoforms are excluded from this compartment.

DOI

10.21007/etd.cghs.2013.0029

Comments

Six month embargo expired June 2014

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