title: RhoA inhibits neural differentiation in murine stem cells through multiple mechanisms.
abstract: Spontaneous neural differentiation of embryonic stem cells is induced by Noggin-mediated inhibition of bone morphogenetic protein 4 (BMP4) signaling. RhoA is a guanosine triphosphatase (GTPase) that regulates cytoskeletal dynamics and gene expression, both of which control stem cell fate. We found that disruption of Syx, a gene encoding a RhoA-specific guanine nucleotide exchange factor, accelerated retinoic acid-induced neural differentiation in murine embryonic stem cells aggregated into embryoid bodies. Cells from Syx(+/+) and Syx(-/-) embryoid bodies had different abundances of proteins implicated in stem cell pluripotency. The differentiation-promoting proteins Noggin and RARγ (a retinoic acid receptor) were more abundant in cells of Syx(-/-) embryoid bodies, whereas the differentiation-suppressing proteins SIRT1 (a protein deacetylase) and the phosphorylated form of SMAD1 (the active form of this transcription factor) were more abundant in cells of Syx(+/+) embryoid bodies. These differences were blocked by the overexpression of constitutively active RhoA, indicating that the abundance of these proteins was maintained, at least in part, by RhoA activity. The peripheral stress fibers in cells from Syx(-/-) embryoid bodies were thinner than those in Syx(+/+) cells. Furthermore, less Noggin and fewer vesicles containing Rab3d, a GTPase that mediates Noggin trafficking, were detected in cells from Syx(-/-) embryoid bodies, which could result from increased Noggin exocytosis. These results suggested that, in addition to inhibiting Noggin transcription, RhoA activity in wild-type murine embryonic stem cells also prevented neural differentiation by limiting Noggin secretion.
authors.affiliation: Cardeza Foundation for Hematologic Research, Department of Medicine, Sidney Kimmel Medical College, Thomas Jefferson University, Philadelphia, PA 19107, USA.
authors.forename: Junning
authors.initials: J
authors.lastname: Yang
authors.affiliation: Department of Molecular Cardiology, Cleveland Clinic Foundation, Cleveland, OH 44195, USA.
authors.forename: Chuanshen
authors.initials: C
authors.lastname: Wu
authors.affiliation: Cardeza Foundation for Hematologic Research, Department of Medicine, Sidney Kimmel Medical College, Thomas Jefferson University, Philadelphia, PA 19107, USA.
authors.forename: Ioana
authors.initials: I
authors.lastname: Stefanescu
authors.affiliation: Department of Medical Biochemistry and Biophysics, Karolinska Institute, Stockholm 17177, Sweden.
authors.forename: Lars
authors.initials: L
authors.lastname: Jakobsson
authors.affiliation: Division of Biostatistics, Department of Pharmacology and Experimental Therapeutics, Sidney Kimmel Medical College, Thomas Jefferson University, Philadelphia, PA 19107, USA.
authors.forename: Inna
authors.initials: I
authors.lastname: Chervoneva
authors.affiliation: Cardeza Foundation for Hematologic Research, Department of Medicine, Sidney Kimmel Medical College, Thomas Jefferson University, Philadelphia, PA 19107, USA. arie.horowitz@jefferson.edu.
authors.forename: Arie
authors.initials: A
authors.lastname: Horowitz
mesh_terms: D000818:Animals; D002352:Carrier Proteins / Q000096:biosynthesis / Q000235:genetics; D002454:Cell Differentiation / Q000502:physiology*; D020662:Guanine Nucleotide Exchange Factors / Q000235:genetics / Q000378:metabolism; D051379:Mice; D018345:Mice, Knockout; D000066450:Mouse Embryonic Stem Cells / Q000166:cytology / Q000378:metabolism*; D009474:Neurons / Q000166:cytology / Q000378:metabolism*; D014158:Transcription, Genetic / Q000502:physiology; D020692:rab3 GTP-Binding Proteins / Q000235:genetics / Q000378:metabolism; D020741:rho GTP-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D020742:rhoA GTP-Binding Protein; D000099176:Noggin Protein
chemical_list: D002352:Carrier Proteins; D020662:Guanine Nucleotide Exchange Factors; D020692:rab3 GTP-Binding Proteins; D020741:rho GTP-Binding Proteins; D020742:rhoA GTP-Binding Protein; D000099176:Noggin Protein; C511269:Syx protein, mouse; C116711:Rab3d protein, mouse; C526380:RhoA protein, mouse