Myst4 is required for neural differentiation of mouse embryonic stem cells

ECHEGARAY, CAMILA VAZQUEZ; MARCOS FRANCIA; COSENTINO, MARÍA SOLEDAD; ALEJANDRA GUBERMAN; SOLARI, CLAUDIA; WAISMAN, ARIEL; MIRIUKA, SANTIAGO; MARÍA VICTORIA PETRONE; BARAÑAO, LINO

Congreso; LXI Congreso de la Sociedad Argentina de Investigaciones Clínicas; 2016

The transition between transcriptional programs associated with Stem Cell (SC) differentiation isrelated to changes in chromatin structure. In this work we studied Myst4, a transcriptionalcoactivator with histone acetyltransferase activity. It was reported that Myst4 gene has aregulatory element highly occupied by Embryonic SC (ESC) key transcription factors (TFs), but therelevance of this protein in ESC remains to be established. Additionally, this gene is important forthe establishment and self-renewal of adult neural SC, and loss of only one allele in humans leadsto intellectual disability. We have previously shown that Myst4 is expressed in mouse ESC (mESC)and is repressed during differentiation. Moreover, our previous results also suggest that itsexpression is regulated by pluripotency TFs.To study the role of Myst4 in the maintenance of ESC?s properties, we used the CRISPR/Cas9strategy to generate mESC knock-out cell lines. Most of the clones analyzed presented indelmutations. We selected a clone with a frameshift mutation that generated a premature stopcodon in both alelles (M4 -/- ), and confirmed the lack of Myst4 protein expression by Westernblot. This clone displayed normal morphology and had no significant differences to the wild type(WT) control cells regarding pluripotency markers expression. Surprisingly, M4-/- mESC failed todifferentiate to neural derivatives during a directed differentiation protocol, with most of the cellsdying at day 12. Gene expression analysis during initial stages of neural differentiation showedthat M4 -/- mESC had lower expression of neural progenitor and neuron markers than WT cells.These results suggest that Myst4 is required for the differentiation of mESC to neurons, andprovides a platform to study the epigenetic mechanisms of normal development as well as humandisease. We consider that understanding the processes involved in ESC chromatin structureregulation is critical to their future application.