Fig. 5

HCK maintains the CDK6 level to promote cell proliferation. (a) CDK6 expression decreased in both human cell lines and BM LSCs after HCK deficiency, as determined by RT-PCR. (b) Immunoblotting analysis of CDK6 expression after HCK deficiency in LSCs, THP-1 and U937. (c) Quantification of CDK6 protein level by densitometric analysis. (d) shRNA targeting CDK6, followed by immunoblotting for CDK6. (e) The numbers of cells were counted at the indicated days after infection with shRNA targeting CDK6, HCK or scrambled shRNA. (f) Representative images of colonies formed by CDK6-knockdown THP-1 cells after 14 days. (g) The colony numbers were calculated in g. (h) Representative flow cytometric analysis of the cell cycle distribution in THP-1 cells targeted by shHCK or SCR was determined using a BrdU incorporation assay. (i) HCK-knockdown and CDK6 overexpression, followed by immunoblotting for HCK and CDK6. (j) The numbers of THP-1 cells were counted at the indicated days after infection with shRNA targeting HCK, with shRNA targeting HCK and plasmids encoding CDK6, or with scrambled shRNA. (k) The relative proliferation of THP-1 cells on the indicated days after infection with shRNA targeting HCK, with shRNA targeting HCK and plasmids encoding CDK6, or with scrambled shRNA by Cell Counting Kit-8 assays. (l) Representative images of colonies formed by THP-1 cells after infection with shRNA targeting HCK, with shRNA targeting HCK and plasmids encoding CDK6, or with scrambled shRNA. (m) The colony numbers were calculated in m. (n) The cell cycle distribution of THP-1 cells infected with shRNA targeting HCK, with shRNA targeting HCK and plasmids encoding CDK6, or with scrambled shRNA