Loss of Retinoic Acid Receptors in Mouse Skin and Skin Tumors Is Associated with Activation of the rasHa Oncogene and High Risk for Premalignant Progression

Retinoic acid receptor transcripts (RARα and RARγ) are decreased in benign mouse epidermal tumors relative to normal skin and are almost absent in carcinomas. In this report, the expression of RARα and RARγ proteins was analyzed by immunoblotting in benign skin tumors induced by two different promotion protocols designed to yield tumors at low or high risk for malignant conversion. RARα was slightly reduced in papillomas promoted with 12- O -tetradecanoylphorbol-13-acetate (low risk) and markedly decreased or absent in papillomas promoted by mezerein (high risk). However, mezerein also caused substantial reduction of RARα in nontumorous skin. RARγ was not detected in tumors from either protocol and was greatly reduced in skin treated by either promoter. Both RARα and RARγ proteins were decreased in keratinocytes overexpressing an oncogenic v- rasHa gene, and RARα was underexpressed in a benign keratinocyte cell line carrying a mutated c- rasHa gene. Introduction of a recombinant RARα expression vector into benign keratinocyte tumor cells reduced the S-phase population and inhibited [3H]thymidine incorporation in response to retinoic acid. Furthermore, transactivation of B-RARE-tk-LUC by retinoic acid was markedly decreased in keratinocytes transduced with the v- rasHa oncogene (v- rasHa -keratinocytes). Blocking protein kinase C function in v- rasHa -keratinocytes with bryostatin restored RARα protein to near normal levels, reflecting the involvement of protein kinase C in RARα regulation. Both RARα and RARγ are down-regulated in cultured keratinocytes by 12- O -tetradecanoylphorbol-13-acetate, further implicating PKC in the regulation of retinoid receptors. Our data suggest that modulation of RARs could contribute to the neoplastic phenotype in mouse skin carcinogenesis and may be involved in the differential promoting activity of mezerein and 12- O -tetradecanoylphorbol-13-acetate, particularly for selecting tumors at high risk for malignant conversion.