Targeting of protein kinase A by muscle A kinase-anchoring protein (mAKAP) regulates phosphorylation and function of the skeletal muscle ryanodine receptor.
Protein kinase A anchoring proteins (AKAPs) tether cAMP-dependent protein kinase (PKA) to specific subcellular locations. The muscle AKAP, mAKAP, co-localizes with the sarcoplasmic reticulum Ca2+ release channel or ryanodine receptor (RyR). The purpose of this study was to determine whether anchoring of PKA by mAKAP regulates RyR function. Either mAKAP or mAKAP-P, which is unable to anchor PKA, was expressed in CHO cells stably expressing the skeletal muscle isoform of RyR (CHO-RyR1). Immunoelectron microscopy showed that mAKAP co-localized with RyR1 in disrupted skeletal muscle. Following the addition of 10 microm forskolin to activate adenylyl cyclase, RyR1 phosphorylation in CHO-RyR1 cells expressing mAKAP increased by 42.4 +/- 6.6% (n = 4) compared with cells expressing mAKAP-P. Forskolin treatment alone did not increase the amplitude of the cytosolic Ca2+ transient in CHO-RyR1 cells expressing mAKAP or mAKAP-P; however, forskolin plus 10 mm caffeine elicited a cytosolic Ca2+ transient, the amplitude of which increased by 22% (p < 0.05) in RyR1/mAKAP-expressing cells compared with RyR1/mAKAP-P-expressing cells. Therefore, localization of PKA by mAKAP at RyR1 increases both PKA-dependent RyR phosphorylation as well as efflux of Ca2+ through the RyR. Therefore, RyR1 function is regulated by mAKAP targeting of PKA, implying an important functional role for PKA phosphorylation of RyR in skeletal muscle.
Ruehr
Mary L
ML
Department of Molecular Cardiology, Lerner Research Institute, Cleveland, Ohio 44195, USA.
Russell
Mary A
MA
Ferguson
Donald G
DG
Bhat
Manju
M
Ma
Jianjie
J
Damron
Derek S
DS
Scott
John D
JD
Bond
Meredith
M
D000818:Animals; D016466:CHO Cells; D002118:Calcium / Q000378:metabolism; D002352:Carrier Proteins / Q000378:metabolism*; D006224:Cricetinae; D009124:Muscle Proteins / Q000378:metabolism; D018482:Muscle, Skeletal / Q000378:metabolism*; D010766:Phosphorylation; D051381:Rats; D019837:Ryanodine Receptor Calcium Release Channel / Q000378:metabolism*
D002352:Carrier Proteins; D009124:Muscle Proteins; D019837:Ryanodine Receptor Calcium Release Channel; D002118:Calcium