This is an open access article, free of all copyright, and may be freely reproduced, distributed, transmitted, modified, built upon, or otherwise used by anyone for any lawful purpose. The work is made available under the Creative Commons CC0 public domain dedication. β-adrenergic receptor (β-AR) stimulation induces cardiac myocyte apoptosis in vitro and in vivo. Neurofibromin 2 (NF2) 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. Here, we tested the hypothesis that β-AR stimulation induces post-translational modifications of NF2, and NF2 plays a pro-apoptotic role in β-AR-stimulated myocyte apoptosis. Methods and results Treatment of adult rat ventricular myocytes (ARVMs) with β-AR agonist (isoproterenol) for 15 min increased phosphorylation (serine-518) and sumoylation of NF2. Co-immunoprecipi-tation assay confirmed β-AR-stimulated sumoylation of NF2. β-AR stimulation enhanced nuclear translocation of phosphorylated and sumoylated NF2. Specific inhibition of β1-AR and protein kinase A (PKA) decreased β-AR-stimulated increase in NF2 post-translational modifications, while inhibition of β2-AR had no effect. Activation of adenylyl cyclase using forskolin (FSK) mimicked the effects of β-AR stimulation. β-AR stimulation and expression of wild-type (WT)-NF2 using adenoviruses increased phosphorylation of mammalian sterile like kinase-1/2 (MST1/2) and yes activated protein (YAP), downstream targets of NF2. Knockdown of NF2 using siRNA in H9C2 cardiomyocytes decreased β-AR-stimulated increase in NF2 and YAP phosphorylation. siRNA-mediated knockdown of NF2 decreased β-AR-stimulated increase in apoptosis, while expression of WT-NF2 induced apoptosis in ARVMs. Expression of WT-NF2 stimulated the mitochondrial death pathway as evidenced by activation of c-Jun N-terminal Kinases (JNKs), and increase in cytosolic cytochrome c levels and Bax expression. Conclusion β-AR stimulation affects post-translational modifications of NF2 via the involvement β1-AR/PKA/cAMP pathway, and NF2 plays a pro-apoptotic role in β-AR-stimulated myocyte apoptosis via the phosphorylation (inactivation) of YAP and involvement of mitochondrial death pathway.
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Dalal, Suman; Connelly, Barbara; Singh, Mahipal; and Singh, Krishna. 2018. NF2 Signaling Pathway Plays a Pro-Apoptotic Role in β-Adrenergic Receptor Stimulated Cardiac Myocyte Apoptosis. PLoS ONE. Vol.13(4). https://doi.org/10.1371/journal.pone.0196626 PMID: 29709009