Fig. 3
SKP2 promotes nuclear localization and transcriptional activity of YAP. a, b HEK293A cells were stably transfected with MYC-SKP2, cultured at medium (a) or high (b) density, and immunostained with a YAP-specific antibody (green). DAPI (blue) was used to stain DNA. Scale bar, 20 μm. c, d HEK293A cells expressing SKP2 shRNA (c) or gRNA (d) were immunostained with a YAP-specific antibody (red in c; green in d). DAPI (blue) was used to stain DNA. Scale bar, 20 μm. e HEK293A cells were stably infected with HA-FLAG-YAP and SKP2 shRNA and immunostained with an HA-specific antibody (red). DAPI (blue) was used to stain DNA. Scale bar, 20 μm. f YAP knockout HEK293A cells (generated by CRISPR-Cas9) were transfected with SFB-YAP (WT or 2KR) with or without MYC-SKP2, and immunostained with antibodies against YAP (red) and MYC (green). DAPI (blue) was used to stain DNA. Scale bar, 20 μm. g–i qPCR of ANKRD1, CYR61, and CTGF in HEK293A cells expressing MYC-SKP2 (g), SKP2 shRNA (h) or SKP2 gRNA (i, upper panel). Lower panel in i: immunoblotting of SKP2 and α-tubulin in three independent SKP2 knockout HEK293A clones generated by CRISPR-Cas9. n = 3 biological replicates. Error bars in g–i are s.e.m. Statistical significance was determined by a two-tailed, unpaired Student’s t-test. *P < 0.05; **P < 0.01; ***P < 0.001
