TY - JOUR
T1 - p53 N-terminus-targeted protein kinase activity is stimulated in response to wild type p53 and DNA damage
AU - Knippschild, Uwe
AU - Milne, Diane M.
AU - Campbell, Linda
AU - Meek, David
PY - 1996/10/3
Y1 - 1996/10/3
N2 - The p53 tumour suppressor protein plays a central role in the cellular defence against agents which cause genetic damage. The activity of p53 is regulated at different levels and is subject to multi-site phosphorylation by a variety of different protein kinases. In this paper we have characterised p53 N-terminus-targeted protein kinase (p53NK) activities, present in a range of cell lines, following fractionation of cellular lysates by ion exchange chromatography on HiTrap Q and Mono Q resins. Three peaks of p53NK activity were observed following fractionation of HeLa cell lysates; these activities were each able to catalyse phosphorylation of up to three residues (serines 4, 6 and 9 in murine p53) within the N-terminus of the p53 protein. Similarly, multiple p53NK activities were detected in the MethAp53(ts) cell line (which expresses the valine 135 temperature-sensitive p53 protein). Strikingly, when these cells were shifted from 38 degrees C (the non-permissive temperature) to 28 degrees C, at which the p53 adopts a wild type conformation, a fivefold stimulation of kinase activity was detected. Moreover, when the DNA damage-inducing drugs etoposide or camptothecin were added to the cells, a further stimulation of kinase activity was observed following growth at 28 degrees C, but not 38 degrees C. These data are consistent with a regulatory model in which p53 is sensitive to stress or DNA damage through phosphorylation at its N-terminus.
AB - The p53 tumour suppressor protein plays a central role in the cellular defence against agents which cause genetic damage. The activity of p53 is regulated at different levels and is subject to multi-site phosphorylation by a variety of different protein kinases. In this paper we have characterised p53 N-terminus-targeted protein kinase (p53NK) activities, present in a range of cell lines, following fractionation of cellular lysates by ion exchange chromatography on HiTrap Q and Mono Q resins. Three peaks of p53NK activity were observed following fractionation of HeLa cell lysates; these activities were each able to catalyse phosphorylation of up to three residues (serines 4, 6 and 9 in murine p53) within the N-terminus of the p53 protein. Similarly, multiple p53NK activities were detected in the MethAp53(ts) cell line (which expresses the valine 135 temperature-sensitive p53 protein). Strikingly, when these cells were shifted from 38 degrees C (the non-permissive temperature) to 28 degrees C, at which the p53 adopts a wild type conformation, a fivefold stimulation of kinase activity was detected. Moreover, when the DNA damage-inducing drugs etoposide or camptothecin were added to the cells, a further stimulation of kinase activity was observed following growth at 28 degrees C, but not 38 degrees C. These data are consistent with a regulatory model in which p53 is sensitive to stress or DNA damage through phosphorylation at its N-terminus.
M3 - Article
C2 - 8875976
SN - 0950-9232
VL - 13
SP - 1387
EP - 1393
JO - Oncogene
JF - Oncogene
IS - 7
ER -