Hofsteen Peter, Robitaille Aaron Mark, Strash Nicholas, Palpant Nathan, Moon Randall T, Pabon Lil, Murry Charles E
Department of Pathology, School of Medicine, University of Washington, 850 Republican Street, Brotman Building Room 453, Seattle, WA 98109, USA; Center for Cardiovascular Biology, School of Medicine, University of Washington, Seattle, WA 98109, USA; Institute for Stem Cell and Regenerative Medicine, School of Medicine, University of Washington, Seattle, WA 98109, USA.
Department of Pharmacology, School of Medicine, University of Washington, Seattle, WA 98109, USA; Institute for Stem Cell and Regenerative Medicine, School of Medicine, University of Washington, Seattle, WA 98109, USA.
iScience. 2018 Apr 27;2:88-100. doi: 10.1016/j.isci.2018.03.010.
Cardiac development requires coordinated biphasic regulation of the WNT/β-catenin signaling pathway. By intersecting gene expression and loss-of-function siRNA screens we identified Alpha Protein Kinase 2 (ALPK2) as a candidate negative regulator of WNT/β-catenin signaling in cardiogenesis. In differentiating human embryonic stem cells (hESCs), ALPK2 is highly induced as hESCs transition from mesoderm to cardiac progenitors. Using antisense knockdown and CRISPR/Cas9 mutagenesis in hESCs and zebrafish, we demonstrate that ALPK2 promotes cardiac function and cardiomyocyte differentiation. Quantitative phosphoproteomics, protein expression profiling, and β-catenin reporter assays demonstrate that loss of ALPK2 led to stabilization of β-catenin and increased WNT signaling. Furthermore, cardiac defects attributed to ALPK2 depletion can be rescued in a dose-dependent manner by direct inhibition of WNT signaling through the small molecule XAV939. Together, these results demonstrate that ALPK2 regulates β-catenin-dependent signaling during developmental commitment of cardiomyocytes.
心脏发育需要WNT/β-连环蛋白信号通路的双相协调调节。通过基因表达分析和功能丧失性siRNA筛选，我们确定α蛋白激酶2（ALPK2）是心脏发生过程中WNT/β-连环蛋白信号的候选负调节因子。在分化的人类胚胎干细胞（hESC）中，随着hESC从中胚层向心脏祖细胞转变，ALPK2被高度诱导。通过在hESC和斑马鱼中使用反义敲低和CRISPR/Cas9诱变，我们证明ALPK2促进心脏功能和心肌细胞分化。定量磷酸蛋白质组学、蛋白质表达谱分析和β-连环蛋白报告基因检测表明，ALPK2的缺失导致β-连环蛋白的稳定和WNT信号增加。此外，通过小分子XAV939直接抑制WNT信号，可以剂量依赖性方式挽救因ALPK2缺失导致的心脏缺陷。这些结果共同表明，ALPK2在心肌细胞发育过程中调节β-连环蛋白依赖性信号。
