Phosphorylation of the p53 tumor suppressor protein is likely to play an important role in regulating its activity. Serine 15 phosphorylation of p53 leads to a stabilization of p53 by reducing its interaction with murine double minute 2, a negative regulatory partner. Recently, p53 was reported to be activated and phosphorylated at serine 15 following UV radiation. However, the signaling pathway that mediates UV-induced phosphorylation is less well characterized. Here, we provide evidence that UVB-induced phosphorylation of p53 at serine 15 is mediated directly by ERKs and p38 kinase. We find that in a mouse JB6 epidermal cell line, ERKs and p38 kinase form a complex with p53 following UVB radiation. Inhibition of ERKs or p38 kinase activity by the use of a dominant negative mutant of ERK2 or p38 kinase or their respective specific inhibitor, PD98059 or SB202190, results in abrogation of UVB-induced phosphorylation of p53 at serine 15. Strikingly, incubation of UVB-activated ERKs or p38 kinase immunoprecipitated complex with exogenous p53 shows serine 15 phosphorylation of both exogenous and co-precipitated endogenous p53 protein. Additionally, active recombinant ERK1/2 and p38 kinase but not JNKs are also able to phosphorylate p53 at serine 15 in vitro. Furthermore, pretreatment of cells with PD98059 or SB202190 blocks p53-dependent transcription activity but increases the level of p53 co-precipitated murine double minute. These results strongly suggest that both ERKs and p38 kinase have a direct role in UVB-induced phosphorylation of p53 at serine 15 in vivo.

UV-induced signal transduction may be involved in tumor promotion and induction of apoptosis. The role of protein kinase C (PKC) in UVB-induced signal transduction is not well understood. This study showed that UVB markedly induced translocation of membrane-associated PKC⑀ and PKC␦, but not PKC␣, from cytosol to membrane. Dominant negative mutant (DNM) PKC⑀ or PKC␦ inhibited UVB-induced translocation of PKC⑀ and PKC␦, respectively. UVB-induced activation of extracellular signal-regulated protein kinases (Erks) and c-Jun NH 2 -terminal kinases (JNKs) was strongly inhibited by DNM PKC⑀ and PKC␦, whereas the DNM of PKC␣ was less effective on the UVB-induced phosphorylation of Erks and JNKs. Among the PKC inhibitors used only rottlerin, a selective inhibitor of PKC␦, markedly inhibited the UVB-induced activation of Erks and JNKs, but not p38 kinases. Safingol, a selective inhibitor for PKC␣, did not show any inhibitory effect on UVB-induced mitogen-activated protein kinase activation. GF109203X is a stronger inhibitor of classical PKC than novel PKC. Lower concentrations of GF109203X (<10 M) had no effect on UVB-induced activation of Erks or JNKs. However, at higher concentrations (over 20 M), GF109203X inhibited UVB-induced activation of JNKs, Erks, and even p38 kinases. Meanwhile, rottlerin and GF109203X markedly inhibited UVB-induced apoptosis of JB6 cells, whereas safingol had little inhibitory effect. DNM-Erk2 cells and PD98059, a selective inhibitor for mitogen-activated protein kinase/extracellular signal-regulated kinase 1 that directly activates Erks, inhibited UVBinduced apoptosis. DNM-JNK1 cells also blocked UVBinduced apoptosis, whereas SB202190, a specific inhibitor for p38 kinases, did not produce the inhibitory effect. These data demonstrate that PKC␦ and PKC⑀, but not PKC␣, mediate UVB-induced signal transduction and apoptosis in JB6 cells through activation of Erks and JNKs.UV radiation from the sun is the major environmental factor responsible for a high incidence of nonmelanoma skin cancer (1-4). The electromagnetic spectrum of UV can be divided into three parts: UVA (320 -400 nm), UVB (290 -320 nm), and UVC (100 -290 nm) (2). In animal experiments, both UVB and UVC can act as complete carcinogens, whereas UVA can only act as a tumor promoter (2-6). Because UVC light does not penetrate the atmosphere (2), UVB radiation is believed to be responsible for most of the carcinogenic effects of sun exposure (2, 7-9). Irradiation by UVB or UVC is known to damage DNA and could cause gene mutations such as p53 or ras mutations (10 -12). Because of the importance of these genes in growth and differentiation, a DNA-damaging effect has been proposed as the mechanism of UV-induced initiation (2-9). The mechanism behind the tumor-promoting ability of UV, however, is not well understood.A number of reports have established that UVC and UVB induce certain gene expression (13-16). These "UV responses" activate several signal transduction pathways and transcription factors (14, 17). There are two transcription facto...

Ultraviolet light A (UVA) plays an important role in the etiology of human skin cancer, and UVA-induced signal transduction has a critical role in UVA-induced skin carcinogenesis. S6K co-immunoprecipitated with PI 3-kinase and possibly PDK1. The complex possibly possessed a partial basal level of phosphorylation, but not at MAPK sites, which was available for its activation by MAPKs in vitro. Thus, these results suggest that activation of MAPKs, like PI 3-kinase/mTOR, may be involved in UVA-induced phosphorylation and activation of p70 S6K .