Complex Phosphatase Regulation of Ca2+-activated Cl− Currents in Pulmonary Arterial Smooth Muscle Cells*
AUTOR(ES)
Ayon, Ramon
FONTE
American Society for Biochemistry and Molecular Biology
RESUMO
The present study was undertaken to determine whether the two ubiquitously expressed Ca2+-independent phosphatases PP1 and PP2A regulate Ca2+-activated Cl− currents (ICl(Ca)) elicited by 500 nm [Ca2+]i in rabbit pulmonary artery (PA) myocytes dialyzed with or without 3 mm ATP. Reverse transcription-PCR experiments revealed the expression of PP1α, PP1β/δ, PP1γ, PP2Aα, PP2Aβ, PP2Bα (calcineurin (CaN) Aα), and PP2Bβ (CaN Aβ) but not PP2Bγ (CaN Aγ) in rabbit PA. Western blot and immunofluorescence experiments confirmed the presence of all three PP1 isoforms and PP2A. Intracellular dialysis with a peptide inhibitor of calcineurin (CaN-AIP); the non-selective PP1/PP2A inhibitors okadaic acid (0.5, 10, or 30 nm), calyculin A (10 nm), or cantharidin (100 nm); and the selective PP1 inhibitor NIPP-1 (100 pm) potently antagonized the recovery of ICl(Ca) in cells dialyzed with no ATP, whereas the PP2A-selective antagonist fostriecin (30 or 150 nm) was ineffective. The combined application of okadaic acid (10 nm) and CaN-autoinhibitory peptide (50 μm) did not potentiate the response of ICl(Ca) in 0 ATP produced by maximally inhibiting CaN or PP1/PP2A alone. Consistent with the non-additive effects of either classes of phosphatases, the PP1 inhibitor NIPP-1 (100 pm) antagonized the recovery of ICl(Ca) induced by exogenous CaN Aα (0.5 μm). These results demonstrate that ICl(Ca) in PA myocytes is regulated by CaN and PP1 and/or PP2A. Our data also suggest the existence of a functional link between these two classes of phosphatases.
ACESSO AO ARTIGO
http://www.pubmedcentral.nih.gov/articlerender.fcgi?artid=2781665Documentos Relacionados
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