Abstract 1833: Dual-Specificity Tyrosine Phosphorylation-Regulated Kinase 1A is a NFAT Kinase Mediating Negative Feedback on Calcineurin/NFAT Signaling.
The Calcineurin and NFAT transcriptional pathway plays a pivotal role in cardiac hypertrophy and heart failure. Dual-specificity tyrosine phosphorylation-regulated kinases (DYRK) were identified recently as NFAT priming kinases in non-cardiac cells. We examined DYRK isoforms for a potential role in regulating cardiac Calcineurin/NFAT signaling. Expression levels of all known DYRK isoforms DYRK1A, 1B, -2, -3, -4 and -6 were determined in human cardiac tissue samples by quantitative real-time PCR. Expression was highest for DYRK1A with 3,61±0,7x10–2, with the remaining DYRK isoforms decreasing in the order DYRK-6,-2,-1B,-4,-3. Adenoviruses for overexpression of DYRK1A and a kinase-dead mutant (DYRKK188R) were created to study the effect of DYRK in isolated rat cardiac myocytes on nuclear translocation of GFP-tagged NFATc3. Under basal culture conditions NFAT-GFP was almost exclusively cytoplasmic (1,3±1,0 % nuclear NFAT) and remained cytoplasmic with either co-expression of Ad-DYRK1A (4,0±5,1%) or Ad-DYRK1AK188R (2,6±2,4%). However, upon co-expression of constitutively active Calcineurin, where NFAT-GFP was located predominantly nuclear (82,03±1,94%), co-expression of Ad-DYRK1A (0,83±1,16%), but not Ad-DYRK1AK188R (71,96±6,13%); p<0.01 led to a profound reduction in nuclear NFAT localization, highlighting a powerful inhibitory effect of DYRK1A on NFAT. Expression of Calcineurin in neonatal rat cardiomyocytes increased DYRK1A expression significantly by 29,7% compared to LacZ control-transfected cells, suggesting DYRK1A is part of a negative feedback controlling Calcineurin/NFAT signaling. Likewise, in vivo, mice subjected to aortic banding exhibited a significantly higher DYRK1A mRNA expression compared to sham operated animals. DYRK1A represents the DYRK isoform with highest expression in Cardiomyocytes. DYRK1A, but not kinase-dead DYRK robustly reduces nuclear NFAT localisation in Calcineurin stimulated cells and is part of a previously unknown negative feedback, limiting Calcineurin/NFAT signaling in cardiomyocytes.