CLASP2 and Plk1 interact and colocalize throughout mitosis. (A) Western blot analysis of mitotic HeLa cell extracts IP with an unspecific IgG (Control) and Plk1 antibody (Plk1 IP). The middle and rightmost panels show GFP IPs from mitotic HeLa cells transiently expressing GFP-CLASP2 full length, C terminal, and N terminal, respectively. Total protein extracts (Load), unbound proteins (Unb), and IPs were analyzed by Western blotting with the indicated antibodies. (B) Mitotic distribution of GFP-CLASP2α (green) transiently expressed in HeLa cells and immunostained for Plk1 (red), ACA (blue, right), and DNA counterstained with DAPI (blue, left). Far right panels show KT insets of selected z planes of the respective boxed regions. From top to bottom, the images are ordered as follows: Merge, GFP-CLASP2, Plk1, and ACA. Midbody colocalization during telophase is depicted in the merged figure. Bottom panels shows a nocodazole-treated cell to highlight the MT-independent CLASP2 and Plk1 KT colocalization. Bars: 5 µm and 1 µm (higher magnifications).