Involvement of NF2 Signaling Pathway in β‐Adrenergic Receptor‐Stimulated Cardiac Myocyte Apoptosis

Abstract

β‐adrenergic receptor (β‐AR) stimulation induces cardiac myocyte apoptosis in vitro and in vivo. Tumor suppressor protein NF2 (neurofibromin 2, also known as merlin) is a member of the ezrin/radixin/moesin (ERM) family of proteins. Post‐translational modifications such as phosphorylation and sumoylation affect NF2 activity, subcellular localization and function. Role of NF2 in inhibition of cell growth and apoptosis is well established in a variety of cell types. However, the role of NF2 in β‐AR‐stimulated cardiac myocyte apoptosis remains to be investigated. Here we provide evidence that treatment of adult rat ventricular myocytes (ARVMs) with β‐AR agonist (isoproterenol, ISO, 10 μM) increases phosphorylation (serine 518) and sumoylation of NF2. Activation of adenylyl cyclase using forskolin mimicked the effects of ISO. ISO‐mediated increase in phosphorylation of NF2 was inhibited by protein kinase A inhibitor (H89) and β1‐AR receptor antagonist (CGP 20712A). Knockdown of NF2 using siRNA decreased ISO‐mediated increase in the number of apoptotic ARVMs (CTL, 8.3±0.6; ISO, 23.2±1.0*; negSiRNA+ISO, 21.3±1.2*; NF2‐SiRNA+ISO, 12.0±1.5$; *p<0.05 vs CTL; $p<0.05 vs ISO and negSiRNA+ISO, n=3–4). Adenoviral‐mediated expression of WT‐NF2 led to increased phosphorylation and sumoylation of NF2, and apoptosis. It also activated JNKs, a pro‐apoptotic kinase. Treatment with ISO and forskolin, or adenoviral‐mediated expression of NF2 increased phosphorylation (inactivation) of YAP, a downstream target of NF2. In H9C2 cardiomyocytes, knockdown of NF2 using siRNA decreased YAP phosphorylation. These data suggest that β‐AR stimulation affects post‐translational modification of NF2 via the involvement of PKA pathway, and NF2 plays a pro‐apoptotic role in β‐AR‐stimulated myocyte apoptosis.Support or Funding InformationSupported by Department of Veterans Affairs (BX002332), NIH R15HL129140 (NHLBI) and Institutional Research and Improvement account.