Protein phosphatase type 1-dependent dephosphorylation of the retinoblastoma tumor suppressor protein in ultraviolet-irradiated human skin and keratinocytes.

This study provides evidence for the involvement of a type 1 protein serine/threonine phosphatase in the ultraviolet radiation-induced dephosphorylation of retinoblastoma tumor suppressor protein in human skin and cultured keratinocytes. The retinoblastoma gene product was localized to the nuclei and nucleoli of keratinocytes, and to the nuclei of basal and spinous layer cells of normal human epidermis. Western blot analysis of the retinoblastoma tumor suppressor protein antigen from keratinocytes and skin established the presence of the hypophosphorylated and hyperphosphorylated forms of retinoblastoma tumor suppressor protein. The exposure of keratinocytes and human skin to 200 J per cm2 of ultraviolet radiation, resulted in a rapid depletion in hyperphosphorylated retinoblastoma tumor suppressor protein, and the accumulation of growth inhibitory hypophosphorylated retinoblastoma tumor suppressor protein(105). In unirradiated and ultraviolet-irradiated keratinocytes retinoblastoma tumor suppressor protein was localized to the spindles of M-phase cells. In contrast, the exposure of keratinocytes to ultraviolet in the presence of 5 mM okadaic acid, resulted in an inhibition of retinoblastoma tumor suppressor protein translocation to the mitotic spindles of M-phase keratinocytes. In addition, the ultraviolet radiation-induced depletion in hyperphosphorylated retinoblastoma tumor suppressor protein, and accumulation of hypophosphorylated retinoblastoma tumor suppressor protein(105) was inhibited by 5 mM okadaic acid. Okadaic acid (0.5 nM), however, did not affect the ultraviolet radiation-induced dephosphorylation and depletion of hyperphosphorylation of the retinoblastoma tumor suppressor protein. Western blot analysis of ultraviolet-irradiated keratinocytes demonstrated that the hypophosphorylated growth inhibitory 105 kDa form of retinoblastoma tumor suppressor protein coimmunoprecipitated with the 38 kDa catalytic subunit of a type 1 protein serine/threonine phosphatase.