COOPERATION BETWEEN RETINOIC ACID AND PHORBOL ESTERS ENHANCES HUMAN TERATOCARCINOMA DIFFERENTIATION

This study explored cooperation between the retinoic acid (RA) and pro tein kinase C (PKC) pathways during differentiation of the multipotent ial human teratocarcinoma (TC) cell line NTERA-2 clone D1 (abbreviated NT2/D1). We report here that, compared to RA treatment alone, RA comb ined with the PKC stimulator 12-O-tetradecanoylphorbol-13-acetate (TPA ) enhanced the regulated expression of the immunophenotypic differenti ation markers SSEA-3, a globo-series carbohydrate, and the ganglio-ser ies carbohydrate antigens GD2 and GD3. Northern analysis and transient transfection assays revealed that TPA co-treatment augmented the RA-i nduced expression and activation of the RA nuclear receptor-beta (RAR- beta), one early marker of RA response in NT2/Dl cells. This finding w as extended with transient co-transfection experiments using a PKC-alp ha expression vector which revealed that the PKC pathway can augment t he activation of RAR-beta by RA. These experiments establish PKC as a modulator of RAR-beta expression in NT2/Dl cells. Similarly, experimen ts showed that RA can modulate activation of the PKC-responsive AP-1 c omplex, a transcription factor rapidly activated by TPA. Northern anal ysis and transient transfection assays revealed that, compared to TPA treatment alone, RA and TPA augmented the expression and transcription al activity of AP-1 in NT2/D1 cells. In contrast, transient transfecti on assays revealed no cooperative effect between RA and TPA in HeLa ce lls, indicating that this effect in NT2/D1 cells is cell type-specific . In summary, these studies show that stimulation of the PKC second me ssenger pathway can modulate tumor differentiation and transcriptional activation of a retinoid receptor associated with RA response.