9-CIS AND ALL-TRANS-RETINOIC ACID INDUCE A SIMILAR PHENOTYPE IN HUMANTERATOCARCINOMA CELLS

Prior work has shown that all-trans retinoic acid (t-RA) treatment of the human teratocarcinoma (TC) cell line NTERA-2 clone D1 (abbreviated NT2/D1) induces a neuronal phenotype and other cell lineages. This st udy sought to explore the potential of 9-cis retinoic acid (9-cis RA) as a differentiation-inducing agent of this multipotent cell. Findings reported here show that 9-cis RA induced a phenotype similar to t-RA treatment of NT2/D1 cells. This similarity extended to their effects o n the nuclear receptors retinoic acid receptor-beta (RAR-beta) and ret inoid X receptor-alpha (RXR-alpha). Both retinoids prominently augment ed RAR-beta expression and transactivated a reporter plasmid containin g putative RAR response elements (RAREs) with direct repeats separated by five nucleotides (DR5). Both retinoids had no appreciable effect o n RXR-alpha expression and both minimally transactivated a reporter pl asmid containing putative RXR response elements (RXREs) with direct re peats separated by one nucleotide (DR1). These studies suggest that 9- cis RA and t-RA activate common events during retinoid-mediated NT2/Dl differentiation. This hypothesis was supported by the finding that NT 2/D1 cells rendered refractory to t-RA (NT2/D1-R1) were also resistant to 9-cis RA. To discover alterations that could confer retinoid-refra ctoriness, retinoid receptor expression was examined in NT2/D1-R1 cell s. In contrast to NT2/D1, the NT2/D1-R1 cell was found to have reduced RXR-alpha expression at the level of total cellular RNA. These studie s establish the effectiveness of 9-cis RA as a differentiation agent o f human TC cells and demonstrate that retinoids with different nuclear receptor affinities can induce similar phenotypes in NT2/D1 cells. In addition, the findings in the retinoid resistant NT2/D1-R1 cell impli cate a role for specific retinoid receptors in this human TC different iation program.