Abstract Body: Takotsuba Syndrome (TTS) is a type of emotional/physical stress-induced heart failure (HF). Stress-elevated catecholamines activate adrenergic signaling. Cardiac A-Kinase Anchoring Protein-1 (AKAP1) is known to regulate the adrenergic signaling via protein complex formation and phosphorylation. We recently discovered that activation of the stress response kinase JNK2 is critical in HF onset. Whether AKAP1 is involved with stress-activated JNK2 and HF onset remains completely unknown.

Isoproterenol (Iso; synthetic catecholamine) effects on JNK2 activation were assessed in left ventricles (LV) from Iso-treated wildtype (WT) mice (67 mg/kg i.p., 2 days) and in heterologous HEK-RyR2 cells (inducible cardiac ryanodine receptor-2; myocyte mimic; 5 mM Iso, 24 h). A JNK2 effect on cardiac function was reflected by cardiac output (CO) using echocardiography in cardiac-specific MKK7D (inducible overexpression of active MKK7D; a JNK upstream activator) and MKK7D-JNK2KD (50% JNK2 knockdown in MKK7D) mice. The AKAP1-JNK2 link was assessed using immunoprecipitation (IP) and immunoblotting in tGFP-JNK2 and/or FLAG-AKAP1 transfected HEK293 cells.

Iso-WT LV had increased JNK phosphorylation (JNK-p, activated) and reduced SERCA2 (a hallmark of HF) vs controls. Likely, MKK7-induction in MKK7D LV increased JNK2 and JNK-p (vs WT-littermates; p<0.05, n=5,4), which led to reduced CO by 68% (n=5,6, p<0.01) and downregulated SERCA2 by 79% (p<0.01, n=4,8), also evidenced by preserved CO and SERCA2 in MKK7D-JNK2KD mice (50% ablated JNK2; n=9). Moreover, Iso treatment in HEK-RyR2 cells activated JNK2, but not JNK1, compared to sham controls. Intriguingly, we found that JNK2 was associated with AKAP1, given the co-IP of tGFP-JNK2 with anti-FLAG antibody pulldown of FLAG-AKAP1 co-transfected HEK293 cells (n=3). Further, overexpressed FLAG-AKAP1 increased the phosphorylation of co-transfected tGFP-JNK2 as well as endogenous JNK (p<0.01 & p<0.001, n=3,3), while the notable JNK downstream target c-Jun also showed increased phosphorylation (c-Jun-p; p<0.05, n=3,3), supporting a previously unrecognized role for AKAP1 in JNK2 activation.

In total, our findings suggest that stress activates JNK2 via an adrenergic-AKAP1 activation pathway to drive HF/TTS onset.