Journal: Scientific Reports

Article Title: Specific threonine-4 phosphorylation and function of RNA polymerase II CTD during M phase progression

doi: 10.1038/srep27401

Figure Lengend Snippet: ( a ) Co-staining of Plk1 (green) and Thr4-P (6D7; red) in HeLa cells. Representative image of metaphase chromosomes is shown. Line scans measured the relative localization of Plk1 and the Thr4-P-specific signals. Scale bars, 5 μm. ( b ) HeLa cells were synchronized with nocodazole (20 ng/ml) for 8 h and mitotic cells were collected using the shake off technique. Immunoprecipitation (IP) experiments with antibodies against Plk1 and CTD-Thr4-P (6D7) from extracts of shake off HeLa cells. Immunoprecipitates were analyzed by western blotting with the indicated antibodies. SN, supernatant; IgG, rabbit serum, isotype control. Asterisks, hash-tags and dots indicate bands for Plk1, Thr4-P and heavy chain, respectively. ( c ) Immunoprecipitated Pol IIA from HeLa cell extracts was used as a substrate for Plk1 and Plk3, and analyzed by Western blotting with mAbs specific for Thr4-P or non-modified CTD (1C7). WCE, whole cell extract.

Article Snippet: Specific kinase activity specified by the supplier (Proqinase, Freiburg) in pmol/μg × min were: Plk3 (131), Plk1 (152).

Techniques: Staining, Immunoprecipitation, Western Blot, Control, Modification