Fig. 1.

Inactivation of the cohesin regulatory subunit STAG2 in primary human cells results in intra-S-phase cell cycle arrest and senescence, whereas the core cohesin subunits SMC1A, SMC3, and RAD21 are essential for cell viability. a Sequence alignment of STAG2 genomic DNA isolated from RPE cells (top) with wildtype STAG2 and after Cas9 cleavage with gRNA targeting the STAG2 locus (bottom). b Phase contrast image of RPE cells following ectopic expression of Cas9 and STAG2 gRNA, which uniformly showed morphologic features of senescence and failure to proliferate. c Immunoblot of total lysate from RPE cells following shRNA depletion of STAG2 expression using two independent shRNA sequences (sh1221 and sh3782). d Flow cytometry plots of RPE cells following lentiviral transduction with empty pLKO.1 or STAG2 shRNA demonstrating an abnormal accumulation of cells within S-phase after STAG2 depletion. e Phase contrast image of RPE cells following lentiviral transduction with empty pLKO.1 or STAG2 shRNA demonstrating morphologic features of senescence and failure to proliferate. f RPE cells at 14 days following lentiviral transduction with STAG2 shRNA show β-galactosidase activity characteristic of senescence. g Phase contrast image of RPE cells following lentiviral transduction with shRNAs against cohesin components (left), and summary of observed cellular phenotype (right). Source data are provided as a Source Data file