Distinct roles of glycogen synthase kinase (GSK)-3alpha and GSK-3beta in mediating cardiomyocyte differentiation in murine bone marrow-derived mesenchymal stem cells.
The signaling mechanisms facilitating cardiomyocyte (CM) differentiation from bone marrow (BM)-derived mesenchymal stem cells (MSCs) are not well understood. 5-Azacytidine (5-Aza), a DNA demethylating agent, induces expression of cardiac-specific genes, such as Nkx2.5 and alpha-MHC, in mouse BM-derived MSCs. 5-Aza treatment caused significant up-regulation of glycogen synthase kinase (GSK)-3beta and down-regulation of beta-catenin, whereas it stimulated GSK-3alpha expression only modestly. The promoter region of GSK-3beta was heavily methylated in control MSCs, but was demethylated by 5-Aza. Although overexpression of GSK-3beta potently induced CM differentiation, that of GSK-3alpha induced markers of neuronal and chondrocyte differentiation. GSK-3 inhibitors, including LiCl, SB 216743, and BIO, abolished 5-Aza-induced up-regulation of CM-specific genes, suggesting that GSK-3 is necessary and sufficient for CM differentiation in MSCs. Although specific knockdown of endogenous GSK-3beta abolished 5-Aza-induced expression of cardiac specific genes, surprisingly, that of GSK-3alpha facilitated CM differentiation in MSCs. Although GSK-3beta is found in both the cytosol and nucleus in MSCs, GSK-3alpha is localized primarily in the nucleus. Nuclear-specific overexpression of GSK-3beta failed to stimulate CM differentiation. Down-regulation of beta-catenin mediates GSK-3beta-induced CM differentiation in MSCs, whereas up-regulation of c-Jun plays an important role in mediating CM differentiation induced by GSK-3alpha knockdown. These results suggest that GSK-3alpha and GSK-3beta have distinct roles in regulating CM differentiation in BM-derived MSCs. GSK-3beta in the cytosol induces CM differentiation of MSCs through down-regulation of beta-catenin. In contrast, GSK-3alpha in the nucleus inhibits CM differentiation through down-regulation of c-Jun.
Cho
Jaeyeaon
J
Department of Cell Biology and Molecular Medicine, Cardiovascular Research Institute, University of Medicine and Dentistry of New Jersey, Newark, New Jersey 07103.
Rameshwar
Pranela
P
Department of Medicine, New Jersey Medical School, University of Medicine and Dentistry of New Jersey, Newark, New Jersey 07103.
Sadoshima
Junichi
J
Department of Cell Biology and Molecular Medicine, Cardiovascular Research Institute, University of Medicine and Dentistry of New Jersey, Newark, New Jersey 07103. Electronic address: sadoshju@umdnj.edu.
D000818:Animals; D001374:Azacitidine / Q000494:pharmacology; D001854:Bone Marrow Cells / Q000166:cytology / Q000201:enzymology*; D002454:Cell Differentiation / Q000502:physiology*; D002467:Cell Nucleus / Q000201:enzymology / Q000235:genetics; D019175:DNA Methylation / Q000187:drug effects / Q000502:physiology; D004791:Enzyme Inhibitors / Q000494:pharmacology; D015971:Gene Expression Regulation, Enzymologic / Q000187:drug effects / Q000502:physiology; D038362:Glycogen Synthase Kinase 3 / Q000096:biosynthesis* / Q000235:genetics; D000071679:Glycogen Synthase Kinase 3 beta; D000072576:Homeobox Protein Nkx-2.5; D018398:Homeodomain Proteins / Q000235:genetics / Q000378:metabolism; D059630:Mesenchymal Stem Cells / Q000166:cytology / Q000201:enzymology*; D051379:Mice; D008822:Mice, Transgenic; D032383:Myocytes, Cardiac / Q000166:cytology / Q000201:enzymology*; D009928:Organ Specificity / Q000187:drug effects / Q000502:physiology; D016755:Proto-Oncogene Proteins c-jun / Q000235:genetics / Q000378:metabolism; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism; D015854:Up-Regulation / Q000187:drug effects / Q000502:physiology; D051176:beta Catenin / Q000235:genetics / Q000378:metabolism
D004791:Enzyme Inhibitors; D000072576:Homeobox Protein Nkx-2.5; D018398:Homeodomain Proteins; C484469:Nkx2-5 protein, mouse; D016755:Proto-Oncogene Proteins c-jun; D014157:Transcription Factors; D051176:beta Catenin; D000071679:Glycogen Synthase Kinase 3 beta; C000605507:Gsk3b protein, mouse; D038362:Glycogen Synthase Kinase 3; C448021:glycogen synthase kinase 3 alpha; D001374:Azacitidine