Abstract
Mirk/Dyrk1B is a serine/threonine kinase widely expressed in colon cancers. Serum starvation induced HD6 colon carcinoma cells to enter a quiescent G0 state, characterized by a 2N DNA content and a lower RNA content than G1 cells. Compared with cycling cells, quiescent cells exhibited 16-fold higher levels of the retinoblastoma protein p130/Rb2, which sequesters E2F4 to block entry into G1, 10-fold elevated levels of the CDK inhibitor p27kip1, and 10-fold higher levels of Mirk. However, depletion of Mirk did not prevent entry into G0, but enabled quiescent HD6, SW480, and colo320 colon carcinoma cells to acquire some biochemical characteristics of G1 cells, including increased levels of cyclin D1 and cyclin D3 because of slower turnover, increased activity of their CDK4/cyclin D complexes, and increased phosphorylation and decreased E2F4 sequestering ability of the CDK4 target, p130/Rb2. As a result, depletion of Mirk allowed some cells to escape quiescence and enabled cells released from quiescence to traverse G1 more quickly. The kinase activity of Mirk was increased by the chemotherapeutic drug 5-fluorouracil (5-FU). Treatment of p53 mutant colon cancer cells with 5-FU led to an elongated G1 in a Mirk-dependent manner, as G1 was shortened by ectopic overexpression of cyclin D1 mutated at the Mirk phosphorylation site (T288A), but not by wild-type cyclin D1. Mirk, through regulating cyclin D turnover, and the CDK inhibitor p27, as shown by depletion studies, functioned independently and additively to regulate the exit of tumor cells from quiescence.
Highlights
Solitary disseminated tumor cells that are negative for proliferation markers such as Ki67 are thought to be the source of tumor recurrence
Initial studies showed that most HD6 cells could be made to reversibly enter a quiescent state when the cells were cultured for 48 h in serum-free conditions in high glucose Dulbecco’s modified Eagle’s medium
Cultures of the serum-starved cells were analyzed in parallel to cultures of serum-starved cells released by a change to growth medium containing the chemotherapeutic drug 5-FU to initiate S phase checkpoints, and the mitotic inhibitor nocodazole to block any remaining cycling cells in G2ϩM, so only one cell cycle was allowed in the 24-h release period
Summary
The survival factors Bcl and BclXL were shown to mediate G0 quiescence in NIH3T3 cells and murine embryonic fibroblasts through increasing p27kip stability, which required phosphorylation of p27Ser-10 by Mirk [7]. These observations are consistent with Mirk function as a G0 kinase in nontransformed cells. Expression of leukemia-associated oncogenes in mouse hematopoietic stem cells induced DNA damage and activated a p21cip1-dependent response that led to reversible cell cycle arrest in a quiescent state and DNA repair.
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Free) 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a call
