Abstract 896: Ser9/Ser21 Phosphorylation of GSK-3 Regulates Physiological Growth and Electrical Function in the Heart
Glycogen synthase kinase (GSK3), a unique serine/threonine kinase, is active even in un-stimulated cells and negatively regulates cardiac hypertrophy. Ser9(S9)/Ser21(S21) phosphorylation of GSK3 by upstream kinases, such as Akt, inhibits GSK3 by competing for substrate binding. Hypertrophic stimuli inactivate GSK3, thereby removing the negative constraint of GSK3 upon cardiac hypertrophy. To delineate the role of S9 /S21 phosphorylation of GSK3 in mediating the cardiac effect of GSK3, we characterized homozygous knockin mice in which Akt phosphorylation sites on GSK3α (S21) and GSK3β (S9) were changed to Ala (GSK3-DKI). As expected, basal phosphorylation of S9/S21 was completely eliminated in GSK3-DKI mice. At 4 weeks of age, the mean body weight (BW) of GSK3-DKI is less than that of wild type control (WT) mice (15.4±1.0 vs 17.9±0.5g, p<0.05, n=15,14). Heart weight/tibial length (HW/TL) was significantly smaller in GSK3-DKI than in WT mice (male 46.6±2.4 vs 54.1±2.3, p<0.05, n=9,10; female 46.6±1.8 vs 56.2±2.1, p<0.01, n=6,4). Liver weight (LiW)/TL was also significantly smaller (male 470.4±50.2 vs 608.8±30.4, p<0.05, n=9,10; female 449.6±31.0 vs 616.9±38.2, p<0.01, n=6,4), but Lung weight/TL was normal in GSK3-DKI mice. Left ventricular ejection fraction in GSK3-DKI was not significantly different from that in WT mice. By 7 weeks, BW, HW/TL and LiW/TL in GSK3-DKI were normalized. Under light anesthesia, GSK3-DKI have a significantly slower heart rate than WT at 12 weeks (378.2±14.8 vs 461.9±12.1 bpm, p<0.01, n=4,8). An electrocardiogram (n=7) showed that GSK3-DKI frequently exhibited arrhythmias, including bradycardia, siginificantly widened PR interval (p<0.05), junctional rhythm (1/7) and atrial fibrillation (1/7). Despite the complete lack of S9/S21 phosphorylation of GSK3, β-catenin was slightly upregulated in GSK3-DKI, suggesting that the cellular level of β-catenin is regulated by phosphorylation of S9/S21 GSK3-independent mechanisms. These results suggest that S9/S21 phosphorylation of GSK3 is required not only for physiological growth of the heart in the early stages of life, but also for correct rhythm formation and cardiac conduction. These effects appear to be mediated by β-catenin-independent signaling mechanisms.