The ubiquitous second messenger cAMP acts to integrate and translate information encoded by extracellular messenger molecules, including hormones and neurotransmitters. Intracellular cAMP concentrations are regulated through coordinated changes in the activities of adenylyl cyclases (ACs) and cyclic nucleotide phosphodiesterases (PDEs). Freely diffusing cAMP can reach concentrations sufficient to activate cAMP effector proteins, such as protein kinase A (PKA) or the exchange protein activated by cAMP (EPAC), except in defined compartments where PDEs are localized which allows for spatial and temporal control of the cAMP signal. In human aortic vascular endothelial cells (HAECs) and HEK293T cells we recently identified a macromolecular complex consisting of PDE3B and EPAC and showed that this complex coordinated cAMP-induced effects on adhesion of these cells to fibronectin-coated surfaces. Using “pull-down” assays and peptide array-based approaches we have identified the molecular determinants which coordinate the formation of this complex. Our evidence suggests that the extreme N-terminal 13 amino acids of PDE3B represent the portion of PDE3B that interacts with EPAC. In addition, although several EPAC-encoded peptides were shown to bind PDE3B, immunoprecipitation-based studies identified a region proximal to the cAMP-binding domains as likely to have a dominant role in this binding. Of functional relevance, a cell permeable peptide containing these amino-terminal 13 amino acids of PDE3B antagonizes PDE3B-EPAC interactions in cells. In addition, this peptide impacted the ability of cAMP-elevating agents to coordinate EPAC-dependent cell adhesions. / Thesis (Master, Pathology & Molecular Medicine) -- Queen's University, 2009-10-06 19:07:12.01
| Identifer | oai:union.ndltd.org:LACETR/oai:collectionscanada.gc.ca:OKQ.1974/5269 |
| Date | 07 October 2009 |
| Creators | PRITCHARD, LISA |
| Contributors | Queen's University (Kingston, Ont.). Theses (Queen's University (Kingston, Ont.)) |
| Source Sets | Library and Archives Canada ETDs Repository / Centre d'archives des thèses électroniques de Bibliothèque et Archives Canada |
| Language | English, English |
| Detected Language | English |
| Type | Thesis |
| Format | 1750754 bytes, application/pdf |
| Rights | This publication is made available by the authority of the copyright owner solely for the purpose of private study and research and may not be copied or reproduced except as permitted by the copyright laws without written authority from the copyright owner. |
| Relation | Canadian theses |
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