Abstract 687: Yap is a Nuclear Target of Mst1/Lats2 Regulating Cell Size in Cardiac Myocytes.
Increasing lines of evidence suggest that mammalian STE20-like kinase 1 (Mst1), originally identified as a potent proapoptotic kinase in cardiac myocytes, also critically regulates cell size in cardiac myocytes. Lats2, a kinase stimulated by Mst1, plays an important role in determining cardiac myocyte size as well. However, the underlying molecular mechanisms are unknown. Yes-associated protein (Yap) is a transcriptional co-factor which associates with Tbx5 and Taffazin (Taz). Mutations in Tbx5 and Taz cause congenital heart defects associated with Holt-Oram and Barth syndrome, respectively. Since Yorkie, a homologue of Yap, is negatively regulated by homologues of Mst1 and Lats2 in Drosophila, we examined whether Yap is also regulated by Mst1-Lats2 in cardiac myocytes.Co-immunoprecipitation assays showed that Yap associates with Lats2 in cardiac myocytes. Serine phosphorylation of Yap was increased by co-expression of Lats2, suggesting that Yap is phosphorylated by Lats2. Yap is localized both in the nucleus and in the cytosol. Lats2 primarily localizes in the cytosol, whereas dominant negative (DN)-Lats2 is found predominantly in the nucleus, suggesting that Yap may be phosphorylated by Lats2 primarily in the cytosol. Yap increased the activity of atrial natriuretic factor (ANF)-luciferase (luc) (3.3 ± 0.2 fold, n = 4, p < 0.0001), mimicking the effect of Tbx5/Taz in cardiac myocytes. Co-expression of either Mst1 or Lats2 reduced the ANF-luc activity (0.7 ± 0.1 fold, 0.8 ± 0.1 fold vs. Yap single transfection, n = 4, p < 0.0001), whereas co-expression of DN-Lats2 increased ANF-luc (1.7 ± 0.2 fold, n = 4, p < 0.001), suggesting that the activity of Yap is negatively regulated by endogenous Lats2. Adenovirally expressed DN-Lats2 and Yap both significantly increased the cell size of cardiac myocytes (+44 ± 4%, +82 ± 4%, vs LacZ control, n = 4, 4, p < 0.0001), but there was no additive effect associated with co-expression of DN-Lats2 and Yap (+59 ± 4%. n = 4). Collectively, Yap stimulates both cell size and ANF transcription in cardiac myocytes, whereas the activity of Yap is negatively regulated by Mst1/Lats2. These results suggest that Yap is a nuclear target of the Mst1-Lats2 pathway regulating cell size and gene expression in cardiac myocytes.