title: Regulation of pluripotent cell differentiation by a small molecule, staurosporine.
abstract: Research in the embryo and in culture has resulted in a sophisticated understanding of many regulators of pluripotent cell differentiation. As a consequence, protocols for the differentiation of pluripotent cells generally rely on a combination of exogenous growth factors and endogenous signalling. Little consideration has been given to manipulating other pathways to achieve pluripotent cell differentiation. The integrity of cell:cell contacts has been shown to influence lineage choice during pluripotent cell differentiation, with disruption of cell:cell contacts promoting mesendoderm formation and maintenance of cell:cell contacts resulting in the preferential formation of neurectoderm. Staurosporine is a broad spectrum inhibitor of serine/threonine kinases which has several effects on cell function, including interruption of cell:cell contacts, decreasing focal contact size, inducing epithelial to mesenchyme transition (EMT) and promoting cell differentiation. The possibility that staurosporine could influence lineage choice from pluripotent cells in culture was investigated. The addition of staurosporine to differentiating mouse EPL resulted in preferential formation of mesendoderm and mesoderm populations, and inhibited the formation of neurectoderm. Addition of staurosporine to human ES cells similarly induced primitive streak marker gene expression. These data demonstrate the ability of staurosporine to influence lineage choice during pluripotent cell differentiation and to mimic the effect of disrupting cell:cell contacts. Staurosporine induced mesendoderm in the absence of known inducers of formation, such as serum and BMP4. Staurosporine induced the expression of mesendoderm markers, including markers that were not induced by BMP4, suggesting it acted as a broad spectrum inducer of molecular gastrulation. This approach has identified a small molecule regulator of lineage choice with potential applications in the commercial development of ES cell derivatives, specifically as a method for forming mesendoderm progenitors or as a culture adjunct to prevent the formation of ectoderm progenitors during pluripotent cell differentiation.
authors.affiliation: School of Molecular and Biomedical Science, University of Adelaide, Adelaide, South Australia 5005, Australia.
authors.forename: James Nicholas
authors.initials: JN
authors.lastname: Hughes
authors.affiliation: Department of Zoology, University of Melbourne, Parkville, Victoria 3010 Australia; Australian Stem Cell Centre, Monash University, Clayton, 3800 Victoria, Australia.
authors.forename: Chong Kum Edwin
authors.initials: CK
authors.lastname: Wong
authors.affiliation: Department of Zoology, University of Melbourne, Parkville, Victoria 3010 Australia.
authors.forename: Kevin Xiuwen
authors.initials: KX
authors.lastname: Lau
authors.affiliation: Department of Zoology, University of Melbourne, Parkville, Victoria 3010 Australia; The Menzies Research Institute Tasmania, University of Tasmania, 17 Liverpool Street, Hobart, Tasmania 7000, Australia. Electronic address: joy.rathjen@utas.edu.au.
authors.forename: Peter David
authors.initials: PD
authors.lastname: Rathjen
authors.affiliation: Department of Zoology, University of Melbourne, Parkville, Victoria 3010 Australia; The Menzies Research Institute Tasmania, University of Tasmania, 17 Liverpool Street, Hobart, Tasmania 7000, Australia. Electronic address: joy.rathjen@utas.edu.au.
authors.forename: Joy
authors.initials: J
authors.lastname: Rathjen
mesh_terms: D000818:Animals; D002454:Cell Differentiation / Q000187:drug effects*; D002460:Cell Line; D053595:Embryonic Stem Cells / Q000166:cytology* / Q000187:drug effects; D006801:Humans; D008648:Mesoderm / Q000166:cytology; D051379:Mice; D054258:Neural Plate / Q000166:cytology; D039904:Pluripotent Stem Cells / Q000166:cytology* / Q000187:drug effects; D047428:Protein Kinase Inhibitors / Q000494:pharmacology*; D019311:Staurosporine / Q000494:pharmacology*
chemical_list: D047428:Protein Kinase Inhibitors; D019311:Staurosporine
keywords: Differentiation; Embryonic stem cell; Mesoderm; Neural ectoderm; Primitive streak; Staurosporine