Fig. 1: A CRISPR/Cas9 screen identifies miRNAs as regulators of mammalian XCI.
A Schematic summary of the CRISPR/Cas9 screen in female mouse fibroblast BMSL2 cells. The active X chromosome (Xa) harbors the deletion of Xist. B, C BMSL2 cells expressing a sgRNA against the indicated miRNAs or, as a control, a non-silencing (NS) sgRNA were selected in the HAT medium. The representative images are shown (B), and the results are quantified from three experiments after crystal violet staining (C). Unpaired, two-sided t-test with no multiple adjustments. For miR106a, *p = 0.017; miR363, *p = 0.0303; miR340, *p = 0.026; miR34b, ***p = 4.21 × 10-5; miR30e, *p = 0.02; miR181a, **p = 0.0032. D–G Two-color RNA FISH monitoring expression of G6pdx (Red) and Mecp2 (Green), and Pgk1 (Red) and Lamp2 (Green) in each of the six miRNAs knockdown H4SV cell lines. DAPI staining is shown in blue. The representative images are shown (D, F), and the results are quantified from three experiments (E, G). H DNA FISH monitoring X chromosome content in the six miRNA KD H4SV cell lines is quantified from three experiments. I qRT-PCR analysis monitoring miRNA levels in the whole-cell, nuclear, and cytoplasmic fractions from H4SV cells. Data were analyzed from three experiments. Data is expressed as mean ± SD (C, E, G, H, I). Scale bars: 5 µm (D, F).
