Mechanism of ultraviolet B-induced cell cycle arrest in G2/M phase in immortalized skin keratinocytes with defective p53

The ultraviolet B (UVB) portion (280-320 nm) of solar radiation is consider ed to be a major etiologic factor in human skin cancer and is a known cause of extensive DNA damage. In this study, we observed that UVB exposure of i mmortalized epidermal keratinocytes (HaCat cells) harboring mutant p53 lead s to G(2)/M cell cycle arrest in both asynchronously growing and synchroniz ed cells in a dose dependent manner. Following UVB exposure (200 mJ/cm(2)), we observed a threefold increase in G(2)/M population at 6 h, which increa sed to sixfold. The observed G2/M arrest was associated with an increase in cyclin B level whereas cdc2 protein remained unchanged. However, we observ ed an accumulation of tyrosine 15 hyperphosphorylated cyclin B-cdc2 complex . In addition, we observed an increase in chk1 kinase and a decrease in cdc 25C protein levels. Chk1 phosphorylates cdc25C on serine 216 and inactivate s it whereas cdc25C dephosphorylates tyrosine 15 phosphate of cdc2 and acti vates the cdc2-cyclin B complex. Therefore, the increase in chk1 and the de crease in cdc25C both participate in inhibiting the G2/1M transition. Our d ata identifies two upstream targets leading to inhibition of cyclin B-cdc2 complexes, which explain the inhibition in cyclin B-associated cdcs kinase following UVB exposure. The inactive phosphorylated cdc2-cyclin B complex r emains sequestered in cytoplasm and may migrate to the nucleus following ac tivation. Our data also indicate that WB exerts unique effects in different types of skin keratinocytes having nonfunctional or mutant p53, (C) 2000 A cademic Press.