Overexpression of FK506-Binding protein FKBP12.6 in cardiomyocytes reducesryanodine receptor-mediated Ca2+ leak from the sarcoplasmic reticulum and increases contractility

The FK506-binding protein FKBP12.6 is tightly associated with the cardiac s arcoplasmic reticulum (SR) Ca2+-release channel (ryanodine receptor type 2 [RyR2]), but the physiological function of FKBP12.6 is unclear. We used ade novirus (Ad)-mediated gene transfer to overexpress FKBP12.6 in adult rabbit cardiomyocytes. Western immunoblot and reverse transcriptase-polymerase ch ain reaction analysis revealed specific overexpression of FKBP12.6, with un changed expression of endogenous FKBP12. FKBP12.6-transfected myocytes disp layed a significantly higher (21%) fractional shortening (FS) at 48 hours a fter transfection compared with Ad-GFP-infected control cells (4.8+/-0.2% F S versus 4+/-0.2% FS, respectively; n=79 each; P=0.001). SR-Ca2+ uptake rat es were monitored in beta -escin-permeabilized myocytes using Fura-2. Ad-FK BP12.6-infected cells showed a statistically significant higher rate of Ca2 + uptake of 0.8+/-0.09 nmol/s(-1)/10(6) cells (n = 8, P<0.05) compared with 0.52+/-0.1 nmol/s(-1)/10(6) cells in sham-infected cells (n = 8) at a [Ca2 +] of 1 <mu>mol/L. In the presence of 5 mu mol/L ruthenium red to block Ca2 + efflux via RyR2, SR-Ca2+ uptake rates were not significantly different be tween groups. From these measurements, we calculate that SR-Ca2+ leak throu gh RyR2 is reduced by 53% in FKBP12.6-overexpressing cells. Caffeine-induce d contractures were significantly larger in Ad-FKBP12.6-infected myocytes c ompared with Ad-GFP-infected control cells, indicating a higher SR-Ca2+ loa d. Taken together, these data suggest that FKBP12.6 stabilizes the closed c onformation state of RyR2. This may reduce diastolic SR-Ca2+ leak and conse quently increase SR-Ca2+ release and myocyte shortening.