The cell cycle checkpoint kinase CHK2 mediates DNA damage-induced stabilization of TTK/hMps1

Abstract

Cell cycle progression is monitored constantly to ensure faithful passage of genetic codes and genome stability. We have demonstrated previously that, upon DNA damage, TTK/hMps1 activates the checkpoint kinase CHK2 by phosphorylating CHK2 at Thr68. However, it remains to be determined whether and how TTK/hMps1 responds to DNA damage. In this report, we present evidence that TTK/hMps1 can be induced by DNA damage in normal human fibroblasts. Interestingly, the induction depends on CHK2 because CHK2-targeting small interfering RNA or a CHK2 inhibitor abolishes the increase. Such induction is mediated through phosphorylation of TTK/hMps1 at Thr288 by CHK2 and requires the CHK2 SQ/TQ cluster domain/forkhead-associated domain. In cells, TTK/ hMps1 phosphorylation at Thr288 is induced by DNA damage and forms nuclear foci, which colocalize partially with c-H2AX. Reexpression of TTK/hMps1 T288A mutant in TTK/hMps1-knockdown cells causes a defect in G 2 /M arrest, suggesting that phosphorylation at this site participates in the proper checkpoint execution. Our study uncovered a regulatory loop between TTK/hMps1 and CHK2 whereby DNA damage-activated CHK2 may facilitate the stabilization of TTK/hMps1, therefore maintaining the checkpoint control.

Identification and Selective Degradation of Neopeptide-Containing Truncated Mutant Proteins in the Tumors with High Microsatellite Instability

“…The physiologic roles of TTK, TCF7L2, and MARCKS have been studied in several cancers, and these proteins have significant relevance to cancer progression (41)(42)(43)(44). We originally expected that significant amounts of mutant TTK might be expressed because this mutant protein has minor changes and a readthrough stop codon.…”

UV-C irradiation delays mitotic progression by recruiting Mps1 to kinetochores

“…Mps1 binds to CHK2 and causes intermolecular phosphorylation, which subsequently facilitates the integrity of the G 2 checkpoint. 48,49 Interestingly, a fraction of Mps1 phosphorylated at threonine 288 accumulates in the DNA damage foci in response to X-irradiation during interphase. 49 Apart from the G 2 checkpoint, it has also been suggested that Mps1 is involved in the G 1 tetraploid checkpoint by association with p53.…”

“…48,49 Interestingly, a fraction of Mps1 phosphorylated at threonine 288 accumulates in the DNA damage foci in response to X-irradiation during interphase. 49 Apart from the G 2 checkpoint, it has also been suggested that Mps1 is involved in the G 1 tetraploid checkpoint by association with p53. 50 However, despite these connections, whether the DNA damage signal can activate the SAC remains controversial.…”

A Negative Feedback of the HIF-1α Pathway via Interferon-Stimulated Gene 15 and ISGylation

“…The beads were washed 5 times with 0.8 mL of sonication buffer containing 1% Triton X-100 and once with 0.8 mL of coimmunoprecipitation buffer supplemented with a 1Â protease inhibitor cocktail, 1 mmol/L DTT, and 1 mmol/L PMSF. The beads were then reacted with a 300 mL mixture containing the IVTT lysates, BSA (2 mg/mL), 1Â protease inhibitor cocktail, 1 mmol/L DTT, and 1 mmol/L PMSF at 4 C for 1 hour with rotation, washed 3 times with coimmunoprecipitation buffer, boiled for 7 minutes in 1Â SDS sample buffer, and then subjected to immunoblotting analysis (37).…”