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Hypoxia Increases Labile Zinc in Cultured Sheep Pulmonary Artery Endothelial Cells (SPAEC) in a Nitric Oxide Dependent Fashion: Association with Activation of PKC Episilon
Author(s) -
Cao Rong,
Wang Q. Jane,
Chen Jun,
Wasserloos Karla J,
Leelavanichkul Karanee,
Manzano Elisenda Lopez,
Pitt Bruce R,
St Croix Claudette M
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.a750-a
Subject(s) - protein kinase c , nitric oxide , zinc , chemistry , chromosomal translocation , western blot , hypoxia (environmental) , metallothionein , transfection , fusion protein , microbiology and biotechnology , signal transduction , biophysics , biochemistry , oxygen , biology , gene , organic chemistry , recombinant dna
We hypothesized that hypoxia may affect [Zn 2+ ] I by activating a nitric oxide ‐ metallothionein‐zinc (NO‐MT‐Zn) pathway in SPAEC and labile [Zn 2+ ] I in turn may affect targets such as PKC‐ε. Microspectrofluorometric analysis of SPAEC exposed to low oxygen (P O2 =24mmHg) revealed: a) an acute increase in labile (i.e. TPEN chelatable) [Zn 2+ ] I that was sensitive to L‐NAME (and partially reversible with supplemental L‐arginine); and b) an unfolding (or release of zinc) of MT as determined from an acute decrease in efficiency of fluorescence resonance energy transfer (FRET) of an infected chimeric FRET. MT reporter. Exposure of SPAEC to exogenous zinc (1‐10 μM) was associated with translocation of a transfected GFP‐PKC‐ε fusion protein to perinuclear localization. Biochemical confirmation revealed increases in immunoreactive PKC‐ε in membrane compartments via Western blot from protein isolated from SPAEC exposed to exogenous zinc. Collectively these results are consistent with hypoxia causing S‐nitrosation of MT. The resultant increase in labile [Zn 2+ ] I may potentially result in activation of PKC‐ε and accordingly, this potential new signaling pathway (NO‐MT‐Zn) in endothelial cells may affect PKC‐ε dependent cellular responses to hypoxia. This research was supported in part by NIH HL 65697 (BRP) and Parker B. Francis FNDT (CMS)