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Cyclic AMP Phosphodiesterase 4D4 Activity Critically Maintains Membrane cAMP in Lung Microvascular Endothelium
Author(s) -
Creighton Judy,
Zhu Bing,
Stevens Troy
Publication year - 2006
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.20.4.a748
Subject(s) - phosphodiesterase , microbiology and biotechnology , adenylyl cyclase , endothelium , chemistry , intracellular , endothelial stem cell , cytoskeleton , biochemistry , biology , signal transduction , endocrinology , enzyme , cell , in vitro
In pulmonary microvascular endothelial cells (PMVECs), calcium inhibited adenylyl cyclase type 6 (AC6) and phosphodiesterase type 4 (PDE4) activities coordinate membrane cAMP synthesis and degradation establishing a discrete cAMP pool important for endothelial barrier function. The PDE4D splice variant PDE4(D4) localizes to the membrane through its unique N‐terminus that interacts with the SH3 domain of non‐erythroid spectrin, a principal component of endothelial membrane cytoskeleton. We hypothesized that inhibition of endogenous PDE4(D4) activity would disrupt membrane cAMP sufficient to alter endothelial function. Expression of a catalytically inactive PDE4(D4) N‐terminus (a.a. 1‐166) peptide fused to GFP in PMVECs revealed localization with spectrin at the membrane. PDE activity studies confirmed a loss of PDE4 activity. However, membrane PDE4(D4) protein levels were not altered. PDE4(D4) functions as a dimer suggesting the PDE4D4(1‐166)/GFP construct acted as a dominate negative. Stimulation of cAMP synthesis initiated intercellular gap formation in PMVEC monolayers expressing the PDE4D4(1‐166)/GFP peptide, but not in control wild type monolayers. Collectively, these data support an essential role for PDE4(D4) activity in compartmentalizing membrane cAMP important for pulmonary endothelial barrier function. Supported by HL066299, HL 060024, and AHA 0415112B.