ABSTRACT
Mouse embryonic stem cells (mESCs) are expanded and maintained pluripotent in vitro in the presence of leukemia inhibitory factor (LIF), an IL6 cytokine family member which displays pleiotropic functions, depending on both cell maturity and cell type. LIF withdrawal leads to heterogeneous differentiation of mESCs with a proportion of the differentiated cells apoptosising. During LIF withdrawal, cells sequentially enter a reversible and irreversible phase of differentiation during which LIF addition induces different effects. However the regulators and effectors of LIF-mediated reprogramming are poorly understood. By employing a LIF-dependent 'plasticity' test, that we set up, we show that Klf5, but not JunB is a key LIF effector. Furthermore PI3K signaling, required for the maintenance of mESC pluripotency, has no effect on mESC plasticity while displaying a major role in committed cells by stimulating expression of the mesodermal marker Brachyury at the expense of endoderm and neuroectoderm lineage markers. We also show that the MMP1 metalloproteinase, which can replace LIF for maintenance of pluripotency, mimics LIF in the plasticity window, but less efficiently. Finally, we demonstrate that mESCs maintain plasticity and pluripotency potentials in vitro under hypoxic/physioxic growth conditions at 3% O2 despite lower levels of Pluri and Master gene expression in comparison to 20% O2.
Subject(s)
Cell Plasticity/drug effects , Embryonic Stem Cells/metabolism , Hypoxia/metabolism , Kruppel-Like Transcription Factors/metabolism , Matrix Metalloproteinase 1/pharmacology , Animals , Cell Differentiation/drug effects , Embryonic Stem Cells/drug effects , Enzyme Inhibitors/pharmacology , Kruppel-Like Transcription Factors/genetics , Leukemia Inhibitory Factor/pharmacology , Mice , Phosphoinositide-3 Kinase Inhibitors , Phosphorylation/drug effects , Proto-Oncogene Proteins c-akt/metabolism , RNA, Small Interfering , Signal Transduction/drug effects , Transcription Factors/genetics , Transcription Factors/metabolismABSTRACT
The Oct4 protein, encoded by the Pou5f1 gene was the very first master gene, discovered 25 years ago, to be absolutely required for the stemness properties of murine and primate embryonic stem cells. This transcription factor, which has also been shown to be essential for somatic cell reprogrammation, displays various functions depending upon its level of expression and has been quoted as a "rheostat" gene. Oct4 protein is in complexes with many different partners and its activity depends upon fine post-translational modifications. This review aims at revisiting some properties of this protein, which has not yet delivered all its potentialities.