Abstract
Regulatory networks for differentiation and pluripotency in embryonic stem (ES) cells have long been suggested to be mutually exclusive. However, with the identification of many new components of these networks ranging from epigenetic, transcriptional, and translational to even post-translational mechanisms, the cellular states of pluripotency and early differentiation might not be strictly bi-modal, but differentiating stem cells appear to go through phases of simultaneous expression of stemness and differentiation genes. Translational regulators such as RNA binding proteins (RBPs) and micro RNAs (miRNAs) might be prime candidates for guiding a cell from pluripotency to differentiation. Using Trim71, one of two members of the Tripartite motif (Trim) protein family with RNA binding activity expressed in murine ES cells, we demonstrate that Trim71 is not involved in regulatory networks of pluripotency but regulates neural differentiation. Loss of Trim71 in mES cells leaves stemness and self-maintenance of these cells intact, but many genes required for neural development are up-regulated at the same time. Concordantly, Trim71−/− mES show increased neural marker expression following treatment with retinoic acid. Our findings strongly suggest that Trim71 keeps priming steps of differentiation in check, which do not pre-require a loss of the pluripotency network in ES cells.
Highlights
Regulatory networks for differentiation and pluripotency in embryonic stem (ES) cells have long been suggested to be mutually exclusive
Most strikingly, starting from E9.5 all Trim71−/− embryos exhibit a severe neural tube closure defect characterized by an open cranial neural tube extending from forebrain to hindbrain (Fig. 1c,d)
We found that Plexin B2 shows a higher surface expression in Trim71−/− murine embryonic stem (mES) cells on the entire cell population resulting in a right-shift of the histogram without split signals (Fig. 6b)
Summary
Regulatory networks for differentiation and pluripotency in embryonic stem (ES) cells have long been suggested to be mutually exclusive. With the identification of many new components of these networks ranging from epigenetic, transcriptional, and translational to even post-translational mechanisms, the cellular states of pluripotency and early differentiation might not be strictly bimodal, but differentiating stem cells appear to go through phases of simultaneous expression of stemness and differentiation genes Translational regulators such as RNA binding proteins (RBPs) and micro RNAs (miRNAs) might be prime candidates for guiding a cell from pluripotency to differentiation. De-repression of markers for primitive endoderm (Gata6/4), mesoderm/visceral endoderm (Bmp2) and neuroectoderm (Isl1) was observed[11] These findings suggested that the switch from pluripotency to early-differentiated cells is not following mutually exclusive and binary cell specification states but may rather be described as phases of overlapping programs with several checkpoints that need to be overcome to initiate final differentiation of mES cells. When monitoring loss of Nanog over time, it became apparent that only half of the genes changed upon loss of Nanog are regulated by chromatin modification and transcription, while the remaining genes appear to be regulated by post-transcriptional, translational and post-translational regulation[31,28]
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