A-kinase Anchoring Protein 100 (AKAP100) is Localized in Multiple Subcellular Compartments in the Adult Rat Heart

doi:10.1083/jcb.142.2.511

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© The Rockefeller University Press, 0021-9525/1998//511 $5.00
The Journal of Cell Biology, Volume 142, Number 2, , 1998 511-522

Articles

A-kinase Anchoring Protein 100 (AKAP100) is Localized in Multiple Subcellular Compartments in the Adult Rat Heart

Jiacheng Yang^*, Judith A. Drazba, Donald G. Ferguson, and Meredith Bond^*^,||

^* Department of Molecular Cardiology and Department of Neuroscience, The Lerner Research Institute, Cleveland Clinic Foundation, Cleveland, OH 44195; and Department of Anatomy and ^|| Department of Physiology and Biophysics, School of Medicine, Case Western Reserve University, Cleveland, Ohio 44106

Stimulation of β-adrenergic receptors activates type Iand II cyclic AMP–dependent protein kinase A, resultingin phosphorylation of various proteins in the heart. It hasbeen proposed that PKA II compartmentalization by A-kinase–anchoringproteins (AKAPs) regulates cyclic AMP–dependent signalingin the cell. We investigated the expression and localizationof AKAP100 in adult hearts. By immunoblotting, we identifiedAKAP100 in adult rat and human hearts, and showed that typeI and II regulatory (RI and II) subunits of PKA are presentin the rat heart. By immunofluorescence and confocal microscopyof rat cardiac myocytes and cryostat sections of rat left ventriclepapillary muscles, we localized AKAP100 to the nucleus, sarcolemma,intercalated disc, and at the level of the Z-line. After doubleimmunostaining of transverse cross-sections of the papillarymuscles with AKAP100 plus ${alpha}$ -actinin–specific antibodiesor AKAP100 plus ryanodine receptor–specific antibodies,confocal images showed AKAP100 localization at the region ofthe transverse tubule/junctional sarcoplasmic reticulum. RIis distributed differently from RII in the myocytes. RII, but not RI, was colocalized with AKAP100 in the rat heart. Ourstudies suggest that AKAP100 tethers PKA II to multiple subcellularcompartments for phosphorylation of different pools of substrateproteins in the heart.

Key Words: AKAP • PKA • RI • RII • heart

Abbreviations used in this paper: AKAP, A-kinase anchoring protein; C, PKA catalytic subunit; ECL, enhanced chemiluminescence; LRSC, lissamine rhodamine B sulfonyl chloride; PKA, cAMP-dependent protein kinase A; PKC, protein kinase C; RyR, ryanodine receptor; SR, sarcoplasmic reticulum; T-tubule, transverse tubule.

Address all correspondence to Meredith Bond, Ph.D., Departmentof Molecular Cardiology, FF10, The Lerner Research Institute,The Cleveland Clinic Foundation, 9500 Euclid Avenue, Cleveland,OH 44195. Tel.: 216-444-3734; Fax: 216-445-6062; e-mail: bondm{at}cesmtp.ccf.org

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