Figure 4.

CDK12 interacts with PAK2 and positively correlate with each other. A. SNU-1 cell lysates and recombinant CDK12 and PAK2 proteins were immunoprecipitated with IgG or anti-CDK12/anti-PAK2 antibodies. Immunoprecipitated samples were subjected to western blotting with the indicated antibodies to detect the interaction of CDK12 and PAK2 ex vivo and in vitro. B. The domain architecture of human PAK2 (right) contains an N-terminal Cdc42/Rac interactive binding motif (CRIB) and kinase domain (aa249 to aa499. PAK2 can be cleaved into PAK2-p27 (aa2 to aa212) and PAK2-p34 (aa213 to aa524). (Left) HEK293T cells were transfected with MYC-tagged CDK12 and HA-tagged WT or N-terminally truncated PAK2 including p27, p34, and kinase domain. Immunoprecipitation was performed using anti-HA affinity gel followed by western blots using anti-MYC antibody. The PAK2-p27 binding to CDK12 is essential for this protein-protein interaction. C. The interaction between CDK12 (blue) and PAK2 (red) was predicted using a computational docking model. D. Representative images show that CDK12 (green) and PAK2 (red) are positively co-localized in nucleus (DAPI: blue) and cytoplasm shown by laser scanning confocal microscope through an immunofluorescence assay. E. The expression of PAK2 was examined by IHC staining with a gastric tumor array. The left panels show quantification of all samples and paired samples (n=72). The right panels show representative photographs of each group of paired samples. 100 × magnification. F. The IOD value statistics of CDK12 and PAK2 in human gastric tumor/adjacent tissues with IHC is shown in scatter diagram. CDK12 is positively correlated with PAK2 in human gastric tumors. The representative images are shown to the right from the same patient tissue. The R value calculated by Pearson formula. Data represent means ±SD. P<0.05: *, p<0.01: **, p<0.001: ***. Significance determined by two-tailed Student's t test.