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. Author manuscript; available in PMC: 2013 May 1.
Published in final edited form as: Cytoskeleton (Hoboken). 2012 Apr 3;69(5):303–311. doi: 10.1002/cm.21026

Fig. 8. Phosphorylation of S628 during mitosis.

Fig. 8

Cells expressing WT FLAG-MCAK were synchronized using an overnight thymidine block. Tetracycline was then added during the following steps to prevent further transcription of the cDNA. The cells were reversed from thymidine for 4 h followed by nocodazole treatment. When the majority of cells reached mitosis, mitotic cells were shaken off and released from the nocodazole block. Proteins were analyzed on Western blots with antibodies to FLAG and actin (as a loading control). Lane 1, cells remaining on the dish after the mitotic shake-off. Approximately 70% of these cells were in prophase. Lane 2, cells in nocodazole. Greater than 90% of the cells were in prometaphase. Lane 3, 30 min after reversal of the nocodazole block. The majority of the cells were in telophase or early G1. Note the presence of a slower migrating band during prometaphase indicating phosphorylation at S628, and the absence of MCAK in cells that have reached telophase.