Abstract 128: Nuclear Targeting of the α1A-Adrenergic Receptors Is Required for Cardiac Myocyte Contractility
In the heart, α1-adrenergic receptors (α1-ARs) regulate physiologic hypertrophy, cardiac myocyte survival, and contractility. Conventionally, G protein-coupled receptors such as α1-ARs are thought to localize and signal at the plasma membrane. However, we demonstrated that endogenous α1-ARs localize to the nucleus in wild-type (WT) adult cardiac myocytes, and identified nuclear localization sequences (NLS) in the α1A- and α1B-subtypes. Mutation of the NLS in each subtype resulted in mis-localization of the receptors and loss of signaling. Here, we examined how nuclear α1-ARs induce inotropic responses through regulation of cardiac troponin I (cTnI). In unloaded WT cardiac myocytes, the α1-agonist phenylephrine (PE) reduced sarcomeric shortening and increased cTnI phosphorylation at the protein kinase C (PKC) site threonine 144 (Thr144). PE had no effect in cardiac myocytes lacking both α1A- and α1B-subtypes (α1ABKO), but reconstitution of the α1A-subtype restored PE effects. Conversely, reconstitution of an NLS-mutant-α1A-subtype blunted the effects of PE on shortening and cTnI phosphorylation. Furthermore, in WT cardiac myocytes, the nuclear export inhibitor leptomycin B also blunted the effects of PE. With nuclei isolated from WT cardiac myocytes, we found high levels of PKCδ with lower levels of PKCα and ε, and that PE induced PKCδ activation in isolated nuclei. We also examined endogenous PKC translocation induced by PE in membrane, cytosolic, nuclear, and myofilament fractions, and found that PKCδ translocated from the nuclear to the myofilament fraction. Finally, expression of a dominant-negative PKCδ in WT myocytes blunted PE-induced effects on shortening and cTnI phosphorylation. Our data show that α1A-subtype nuclear localization is required for activation of PKCδ in the nucleus, subsequent phosphorylation of cTnI at Thr144 at the sarcomere to regulate myocyte contractility.
- © 2012 by American Heart Association, Inc.