Published Online
on January 29, 2007

A more recent version of this article appeared on March 1, 2007

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Submitted on August 31, 2006
Revised on September 25, 2006

FK506 Binding Protein 12/12.6 Depletion Increases Endothelial Nitric Oxide Synthase Threonine 495 Phosphorylation and Blood Pressure

Cheng Long; Leslie G. Cook; Susan L. Hamilton; Gang-Yi Wu; and Brett M. Mitchell^*

From the Department of Molecular Physiology and Biophysics, Baylor College of Medicine, Houston, Tex.

^* To whom correspondence should be addressed. E-mail: brettm{at}bcm.tmc.edu.

Abstract--Chronic treatment with the immunosuppressive drug rapamycin leads to hypertension; however, the mechanisms are unknown. Rapamycin binds FK506 binding protein 12 and its related isoform 12.6 (FKBP12/12.6) and displaces them from intracellular Ca²⁺ release channels (ryanodine receptors) eliciting a Ca²⁺ leak from the endoplasmic/sarcoplasmic reticulum. We tested whether this Ca²⁺ leak promotes conventional protein kinase C-mediated endothelial NO synthase phosphorylation at Thr495, which reduces production of the vasodilator NO. Rapamycin treatment of control mice for 7 days, as well as genetic deletion of FKBP12.6, increased systolic arterial pressure significantly compared with controls. Untreated aortas from FKBP12.6^-/- mice and in vitro rapamycin-treated control aortas had similarly decreased endothelium-dependent relaxation responses and NO production and increased endothelial NO synthase Thr495 phosphorylation and protein kinase C activity. Inhibition of either conventional protein kinase C or ryanodine receptor restored endothelial NO synthase Thr495 phosphorylation and endothelial function to control levels. Rapamycin induced a small increase in basal intracellular Ca²⁺ levels in isolated endothelial cells, and rapamycin or FKBP12.6 gene deletion decreased acetylcholine-induced intracellular Ca²⁺ release, all of which were reversed by ryanodine. These data demonstrate that displacement of FKBP12/12.6 from ryanodine receptors induces an endothelial intracellular Ca²⁺ leak and increases conventional protein kinase C-mediated endothelial NO synthase Thr495 phosphorylation leading to decreased NO production and endothelial dysfunction. This molecular mechanism may, in part, explain rapamycin-induced hypertension.

Key words: endothelium • hypertension • experimental • NO • NO synthase • protein kinases • vasorelaxation

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