title: L-Proline induces differentiation of ES cells: a novel role for an amino acid in the regulation of pluripotent cells in culture.
abstract: The development of cell therapeutics from embryonic stem (ES) cells will require technologies that direct cell differentiation to specific somatic cell lineages in response to defined factors. The initial step in formation of the somatic lineages from ES cells, differentiation to an intermediate, pluripotent primitive ectoderm-like cell, can be achieved in vitro by formation of early primitive ectoderm-like (EPL) cells in response to a biological activity contained within the conditioned medium MEDII. Fractionation of MEDII has identified two activities required for EPL cell formation, an activity with a molecular mass of <3 kDa and a second, much larger species. Here, we have identified the low-molecular-weight activity as l-proline. An inhibitor of l-proline uptake, glycine, prevented the differentiation of ES cells in response to MEDII. Supplementation of the culture medium of ES cells with >100 M l-proline and some l-proline-containing peptides resulted in changes in colony morphology, cell proliferation, gene expression, and differentiation kinetics consistent with differentiation toward a primitive ectoderm-like cell. This activity appeared to be associated with l-proline since other amino acids and analogs of proline did not exhibit an equivalent activity. Activation of the mammalian target of rapamycin (mTOR) signaling pathway was found to be necessary but not sufficient for l-proline activity; addition of other activators of the mTOR signaling pathway failed to alter the ES cell phenotype. This is the first report describing a role for amino acids in the regulation of pluripotency and cell differentiation and identifies a novel role for the imino acid l-proline.
authors.affiliation: Univ. of Melbourne, Victoria, Australia.
authors.forename: Jennifer M
authors.initials: JM
authors.lastname: Washington
authors.forename: Joy
authors.initials: J
authors.lastname: Rathjen
authors.forename: Fernando
authors.initials: F
authors.lastname: Felquer
authors.forename: Ana
authors.initials: A
authors.lastname: Lonic
authors.forename: Michael D
authors.initials: MD
authors.lastname: Bettess
authors.forename: Nancy
authors.initials: N
authors.lastname: Hamra
authors.forename: Ljiljana
authors.initials: L
authors.lastname: Semendric
authors.forename: Boon Siang Nicholas
authors.initials: BS
authors.lastname: Tan
authors.forename: Julie-Anne
authors.initials: JA
authors.lastname: Lake
authors.forename: Rebecca A
authors.initials: RA
authors.lastname: Keough
authors.forename: Michael B
authors.initials: MB
authors.lastname: Morris
authors.forename: Peter D
authors.initials: PD
authors.lastname: Rathjen
mesh_terms: D000818:Animals; D002460:Cell Line; D053595:Embryonic Stem Cells / Q000166:cytology* / Q000502:physiology*; D005786:Gene Expression Regulation; D005998:Glycine / Q000494:pharmacology; D006801:Humans; D047908:Intracellular Signaling Peptides and Proteins / Q000235:genetics / Q000378:metabolism; D007930:Leucine / Q000494:pharmacology; D051379:Mice; D039904:Pluripotent Stem Cells / Q000166:cytology* / Q000502:physiology*; D011392:Proline / Q000378:metabolism / Q000494:pharmacology*; D017346:Protein Serine-Threonine Kinases / Q000235:genetics / Q000378:metabolism; D015398:Signal Transduction; D058570:TOR Serine-Threonine Kinases
chemical_list: D047908:Intracellular Signaling Peptides and Proteins; D011392:Proline; C546842:MTOR protein, human; C546843:mTOR protein, mouse; D017346:Protein Serine-Threonine Kinases; D058570:TOR Serine-Threonine Kinases; D007930:Leucine; D005998:Glycine