Expression pattern of proteins that bind to the ultraviolet‐responsive element (TGACAACA) in human keratinocytes

Abstract

We previously identified an ultraviolet (UV)-responsive element (URE; TGACAACA) that plays a role in both transcription and replication of polyoma sequences. Mouse polyclonal antibodies were raised against affinity-purified URE-bound proteins to characterize their expression patterns. These antibodies specifically recognized two of four URE-bound proteins, of 40 and 68 kDa. The 68-kDa protein was constitutively expressed in human keratinocytes, while the expression of the 40-kDa protein was induced by UV irradiation. Of the two, the 68-kDa protein bound to the URE with greater affinity than the 40-kDa protein, as determined by southwestern analysis. The expression of the 40-kDa protein increased as early as 1 h after UV irradiation of both rat fibroblasts and human keratinocytes and correlated with increased binding to the URE in an electrophoretic mobility shift assay. Other types of damage, as well as heat shock and serum stimulation, also induced the expression of this protein, suggesting that it may play a role in cellular response to stress or damage. The 40-kDa protein was expressed at the highest levels in the S phase of the cell cycle and was induced by aphidicolin, suggesting that it has a role in DNA replication. All together, these results suggest that exposure of human keratinocytes to damage- and stress-inducing agents modulates the expression of proteins that may play a role in regulating cellular response to DNA damage.