Nocodazole‐induced growth inhibition of human leukemia cells is linked to downregulation of CAK molecules

Abstract

A cytoskeleton is present in the cytoplasm of all living cells. The cytoskeleton is composed of three different types of fibers: microtubules, microfilaments, and intermediate. The microtubules play an important role in a variety of cellular processes including maintaining cell structure and pulling chromosomes apart during anaphase. Nocodazole is an inhibitor of microtubule polymerization. Inhibition of microtubule polymerization by Nocodazole often lead to arrest of cells in G2 or M phase. However, it is unclear if any cell cycle regulatory molecules are involved in Nocodazole‐mediated cell growth. In this study we focus on CDK activating kinase (CAK). The CAK is composed of three subunits, CDK7, cyclin H, and MAT1. The cyclin H is regulatory subunit of CDK7 whereas MAT1 determines the specificity of CDK7. Addition of Nocodazole significantly inhibited the growth of MV4‐11 and TF‐1a cells in culture. The inhibitions were dose‐dependent with maximal inhibition being observed at a concentration of 10uM of Nocodazole. Less than 1 μM had no significant inhibitory effect on the growth of the cells, measured by XTT proliferation assay. Paralleled to the changes in the cell proliferation, expression of CDK7 and cyclin H were marked decreased when Nocodazole were added to TF‐1a and MV4‐11 cells at concentrations of 5 and 10μM. Similarly, Nocodazole at concentration of 10μM downregulated the phosphorylation levels of CDK7 and the expression of RNA polymerase II in TF‐1a cells. However, Nocodazole had no effect on the expression of MAT1, suggesting that Nocodazole did not interfere with specificity of CDK7. We also found that Nocodazole had no effect on the expression of β‐actin, a non‐muscle cytoskeleton actin.Support or Funding InformationBarry University