REGULATION OF HPSC FATE BY SMARCB1

SMARCB1 is one of the core subunits of the SWI/SNF chromatin-remodeling complex. Therefore, loss of function (LOF) or gain of function (GOF) of this protein might have significant effects on the epigenetic state of the cells and on their phenotype. Indeed, SMARCB1 LOF is the solely genetic lesion in Rhabdoid tumor of the kidney and in AT/RT tumor in the brain. SMARCB1 is also required for early embryonic development as it was shown that SMARCB1-/- mouse blastocysts lose their hatching capacity and that SMARCB1 regulates the expression of Nanog during the differentiation of mouse embryonic stem cells.Here we aimed to study the effect of SMARCB1 LOF and SMARCB1 GOF on human pluripotent stem cells (hPSCs). For this purpose, we introduced a conditional SMARCB1 expressing cassette into the AAVS1 locus and targeted the endogenous SMARCB1 gene. Using this system, we show that both SMARCB1 GOF and SMARCB1 LOF lead to rapid changes in cell fate, however in a different manner. SMARCB1 GOF leads to down-regulation of pluripotency markers such as Oct4 and to spontaneous differentiation of the cells mainly into neuronal lineage. By contrast, SMARCB1 LOF significantly affects the morphology of the cell colonies, which start to grow as 3D structures, but does not appear to directly affect the expression of pluripotency markers. These morphological changes suggest that SMARCB1 LOF affects the interactions between the cells and the extra cellular matrix. Indeed, global gene expression analysis reveals significant down-regulation in pathways related to the extra cellular matrix upon SMARCB1 LOF. To further study the effect of SMARCB1 LOF on the pluripotency of the cells, we evaluated their in-vitro and in-vivo differentiation capacity. Notably, while the LOF cells were able to differentiate into early progenitors cells of the ectoderm, endoderm and mesoderm layers, they failed to further differentiate into more mature cells at least in the neuronal lineage. Taken together, our results reveal the crucial role of SMARCB1 in human pluripotent stem cell biology and show that SMARCB1 has to be strictly regulated to maintain the self-renewal and differentiation capacity of the cells.