Myst4 is required for neural differentiation of mouse embryonic stem cells

VAZQUEZ ECHEGARAY C; WAISMAN A; FRANCIA M; PETRONE MV; GUBERMAN A; BARAÑAO L; COSENTINO S; SOLARI C; MIRIUKA S

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Congreso; saic 2016; 2016

The transition between transcriptional programsassociated with Stem Cell (SC) differentiation is related to changes inchromatin structure. In this work we studied Myst4, a transcriptional coactivator with histone acetyltransferaseactivity. It was reported that Myst4 gene has a regulatory element highlyoccupied by Embryonic SC (ESC) key transcription factors (TFs), but the relevance of thisprotein in ESC remains to be established. Additionally, this gene is importantfor theestablishment and self-renewal of adult neural SC, and loss of only one allelein humans leads to intellectual disability. We have previously shown that Myst4 is expressed in mouse ESC (mESC) andis repressed during differentiation. Moreover, our previous results alsosuggest that its expression is regulated by pluripotency TFs. To study the role of Myst4in the maintenance of ESC?s properties, we used the CRISPR/Cas9 strategy togenerate a mESC knock-out cell line. Mostof the clones analyzed presented indel mutations. We selected a clone with aframeshift mutation that generated a premature stop codon in both alelles (M4-/-), and confirmed the lack of Myst4 protein expression by Western blot. Thisclone displayed normal morphology and had no significant differences to thewild type (WT) control cells regarding pluripotency markers expression.Surprisingly, M4-/- mESC failed to differentiate to neural derivatives during adirected differentiation protocol, with most of the cells dying at day 12. Geneexpression analysis during initial stages of neural differentiation showed thatM4 -/- mESC had lower expression of neural progenitor and neuron markers than WTcells. These results suggestthat Myst4 is required for the differentiation of mESC to neurons, and providesa platform to study the epigenetic mechanisms of normal development as well ashuman disease. We consider that understanding the processes involved in ESC chromatinstructure regulation is critical to their future application.