Abstract 1067: CamKII Regulates NFAT Nuclear Translocation in a Calcineurin Independent Manner
Pathologic cardiac myocyte hypertrophy involves stress induced increases in intracellular Ca2+, which binds to calmodulin (CaM) and then activates both the phosphatase calcineurin (Cn) and CaM kinase (CaMKII). In hypertrophy, Cn dephosphorylates the cytoplasmic nuclear factor of activated T-cells (NFAT), inducing nuclear translocation where it activates hypertrophic target genes. While GSK3, p38, and JNK are kinases that have been shown to directly phosphorylate NFAT, resulting in nuclear export, the role of cytoplasmic CamKII in NFAT translocation is unknown. This study explored the hypothesis that cytoplasmic CamKII directly induces NFAT cytoplasmic localization which antagonizes Cn mediated nuclear NFAT translocation.
Methods: Adenovirus gene delivery was used to express GFP-tagged NFATc3 (m.o.i - 100, 12h) in either cultured neonatal rat (NRVM) or in adult feline ventricular myocytes (AFVM). Myocytes were co-infected with either constitutively active (CamKII-CA) or dominant negative (CamKII-DN) mutants of cytoplasmic targeted CamKIIδc (m.o.i - 100, 12 h). FK506 (2uM) or CsA (2uM) were used to inhibit Cn. The % of myocytes with nuclear translocated NFAT is reported.
Results: In NRVM (and in AFVM), NFAT nuclear translocation (Control: 14.9±1%) was significantly increased (29±2%, p<0.05 vs control) by raising the bath [Ca2+] by 2X. This Ca2+ mediated NFAT translocation was blocked by FK506 (3.8±1%) and CsA (7.3±2%, p<0.05 vs control). CamKII-DN (CaMKII inhibition) increased (44.9±3%, P<0.001 vs control) and CamKII-CA (CaMKII activation) decreased (6.8±2%, P<0.01 vs control) NFAT translocation. FK506 and CsA did not block NFAT translocation in CamKII-DN NRVM (FK506: 50.7±2, CsA: 46.1±2%). CamKII-DN also increased NFAT translocation (50.5±3% vs 24.8±2% in control, P<0.01) in AFVM. CsA treatment also failed to reduce CamKII-DN mediated NFAT translocation (59.6±9, P<0.01 vs control) in adult myocytes.
Conclusion: These data show for the first time that activation of cytoplasmic CaMKII inhibits NFAT nuclear translocation in both neonatal and adult ventricular myocytes, possibly by direct NFAT phosphorylation. These effects are in parallel with but independent of Ca2+ dependent Cn signaling.