title: Quantitative proteomics identify DAB2 as a cardiac developmental regulator that inhibits WNT/β-catenin signaling.
abstract: To reveal the molecular mechanisms involved in cardiac lineage determination and differentiation, we quantified the proteome of human embryonic stem cells (hESCs), cardiac progenitor cells (CPCs), and cardiomyocytes during a time course of directed differentiation by label-free quantitative proteomics. This approach correctly identified known stage-specific markers of cardiomyocyte differentiation, including SRY-box2 (SOX2), GATA binding protein 4 (GATA4), and myosin heavy chain 6 (MYH6). This led us to determine whether our proteomic screen could reveal previously unidentified mediators of heart development. We identified Disabled 2 (DAB2) as one of the most dynamically expressed proteins in hESCs, CPCs, and cardiomyocytes. We used clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated protein 9 (Cas9) mutagenesis in zebrafish to assess whether DAB2 plays a functional role during cardiomyocyte differentiation. We found that deletion of Dab2 in zebrafish embryos led to a significant reduction in cardiomyocyte number and increased endogenous WNT/β-catenin signaling. Furthermore, the Dab2-deficient defects in cardiomyocyte number could be suppressed by overexpression of dickkopf 1 (DKK1), an inhibitor of WNT/β-catenin signaling. Thus, inhibition of WNT/β-catenin signaling by DAB2 is essential for establishing the correct number of cardiomyocytes in the developing heart. Our work demonstrates that quantifying the proteome of human stem cells can identify previously unknown developmental regulators.
authors.affiliation: Department of Pathology, University of Washington, Seattle, WA 98109; Center for Cardiovascular Biology, University of Washington, Seattle, WA 98109; Institute for Stem Cell and Regenerative Medicine, University of Washington, Seattle, WA 98109;
authors.forename: Peter
authors.initials: P
authors.lastname: Hofsteen
authors.affiliation: Institute for Stem Cell and Regenerative Medicine, University of Washington, Seattle, WA 98109; Department of Pharmacology, University of Washington, Seattle, WA 98109;
authors.forename: Aaron M
authors.initials: AM
authors.lastname: Robitaille
authors.affiliation: Department of Pathology, University of Washington, Seattle, WA 98109; Center for Cardiovascular Biology, University of Washington, Seattle, WA 98109;
authors.forename: Daniel Patrick
authors.initials: DP
authors.lastname: Chapman
authors.affiliation: Institute for Stem Cell and Regenerative Medicine, University of Washington, Seattle, WA 98109; Department of Pharmacology, University of Washington, Seattle, WA 98109; Howard Hughes Medical Institute, Chevy Chase, MD 20815; rtmoon@uw.edu murry@uw.edu.
authors.forename: Randall T
authors.initials: RT
authors.lastname: Moon
authors.affiliation: Department of Pathology, University of Washington, Seattle, WA 98109; Center for Cardiovascular Biology, University of Washington, Seattle, WA 98109; Institute for Stem Cell and Regenerative Medicine, University of Washington, Seattle, WA 98109; Department of Bioengineering, University of Washington, Seattle, WA 98109; Division of Cardiology, Department of Medicine, University of Washington, Seattle, WA 98109 rtmoon@uw.edu murry@uw.edu.
authors.forename: Charles E
authors.initials: CE
authors.lastname: Murry
mesh_terms: D048868:Adaptor Proteins, Signal Transducing / Q000502:physiology*; D000818:Animals; D051017:Apoptosis Regulatory Proteins; D002454:Cell Differentiation; D002478:Cells, Cultured; D053595:Embryonic Stem Cells / Q000166:cytology; D005260:Female; D006321:Heart / Q000196:embryology*; D006801:Humans; D036341:Intercellular Signaling Peptides and Proteins / Q000502:physiology; D032383:Myocytes, Cardiac / Q000166:cytology; D040901:Proteomics; D025521:Tumor Suppressor Proteins / Q000502:physiology*; D060449:Wnt Signaling Pathway / Q000502:physiology*; D015027:Zebrafish / Q000196:embryology; D051176:beta Catenin / Q000502:physiology*
chemical_list: D048868:Adaptor Proteins, Signal Transducing; D051017:Apoptosis Regulatory Proteins; C495270:CTNNB1 protein, human; C102885:DAB2 protein, human; C501126:DKK1 protein, human; D036341:Intercellular Signaling Peptides and Proteins; D025521:Tumor Suppressor Proteins; D051176:beta Catenin
keywords: WNT/β-catenin; cardiomyocyte; embryonic stem cell; quantitative proteomics; zebrafish