Rabbit Anti-Human CHEK1 (Phospho-Ser301) polyclonal antibody

Human lymphocytes gene expression is monitored before and after PHA stimulation over 72 h, using DNA microarray technology. Results are then compared with our previous bioinformatics predictions, which identified six leader genes of highest importance in human T lymphocytes cell cycle. Experimental data are strikingly compatible with bioinformatic predictions of the specific role and interaction of PCNA, CDC2, and CCNA2 at all phases of the cell cycle and of CHEK1 in regulating DNA repair and preservation. lt does not escape our notice that the conception and use of ad hoc arrays, based on a bioinformatics prediction which identifies the most important genes involved in a particular biological process, can really be an added value in cell biology and cancer research alternative to massive frequently misleading molecular genomics.

Cell cycle checkpoints ensure that eukaryotic cells do not enter mitosis after ionizing irradiation (IR). The G(2)-arrest after IR is the result of activation of multiple signalling pathways, the contributions of which vary with time after irradiation. We have studied the time evolution of the IR-induced G(2)-arrest in human B-lymphocyte cancer cell lines, as well as the molecular mechanisms responsible for the arrest. Cells that were in G(2) phase at the time of irradiation experienced a transient arrest that blocked entry into mitosis at 0-2 hours after IR (0.5 or 4 Gy). Activation of ATM and CHEK2 occurred at the same time as this early arrest and was, like the arrest, abrogated by the ATM-inhibitor KU-55933. A late, permanent and ATM-independent arrest (>= 6 hours after IR) of cells that were in G(2)/S/G(1) at the time of irradiation (4 Gy) was inactivated by caffeine. This late G(2)-arrest could not be explained by downregulation of genes with functions in G(2)/mitosis (e.g., PLK1, CCNB1/2), since the downregulation was transient and not accompanied by reduced protein levels. However, the persistent phosphorylation of CHEK1 after 4 Gy suggested a role for CHEK1 in the late arrest, consistent with the abrogation of the arrest in CHEK1-depleted cells. TP 53 was not necessary for the late G(2)-arrest, but mediated an intermediate arrest (2-10 hours after IR) independently of ATM and CHEK1. In conclusion, the IR-induced arrest in G(2) is mediated by ATM immediately after irradiation, with TP 53 for independent and transient back-up, while CHEK1 is necessary for the late arrest.