RETINOID-X RECEPTOR (RXR) DIFFERENTIALLY AUGMENTS THYROID-HORMONE RESPONSE IN CELL-LINES AS A FUNCTION OF THE RESPONSE ELEMENT AND ENDOGENOUS RXR CONTENT

Retinoid-X receptor (RXR) forms heterodimers with thyroid hormone rece ptor (TR) and significantly enhances binding to thyroid hormone respon se elements (TREs). Expression of RXR in a transient transfection assa y augments the T-3 response, but the influences of the specific cell l ine and TRE used have not been systematically studied. We determined R XR alpha and -beta augmentation of the TR alpha-mediated T-3 response in transient transfection assays of COS, JEG, and mouse embryonic stem (ES) cell lines for a series of eight wild-type thyroid hormone (T-3) and retinoic acid response elements (previously shown to bind TR). RX R augmented T-3-induced expression in COS and ES cells (1.5- to 4-fold greater expression with added RXR compared to TR alone), but had mini mal effect on augmentation of response in JEG cells. For most elements studied there was a proportional augmentation of basal and T-3-stimul ated expression. TREs from rat GH and laminin-B1, however, had relativ ely higher levels of T-3-induced expression as a result of RXR cotrans fection (T-3 induction ratios increased 2-fold or greater). Previous c haracterization of these elements demonstrates that they contain more than two hexameric binding domains, all of which can simultaneously bi nd TR. The influence of endogenous RXR expression in a cell line on RX R augmentation of the T-3 response was determined. RXR(alpha and -beta messenger RNA (mRNA) expression was quantitated by Northern blot in e ach cell line. COS and JEG cells expressed almost exclusively RXR alph a mRNA, although expression was almost a-fold higher in JEG compared t o COS cells (12 +/- 2.5 vs. 6.8 +/- 0.5 density units relative to acti n; mean +/- SE;P < 0.05). ES cells expressed only RXR beta mRNA, but a t a very low level (0.4 +/- 0.1). Nuclear extracts prepared from JEG a nd COS cells augmented TR binding proportional to the endogenous RXR m RNA expression, and the heterodimer band was supershifted by the addit ion of antibody to RXR alpha. Nuclear extracts from ES cells had no de tectable TR heterodimer binding to a range of response elements. RXR a ugmentation of the T-3 response differs among cell lines and is greate r in those with reduced endogenous RXR. Furthermore, the functional au gmentation of the T-3 response ratio by RXR is likely to require addit ional sequences contained in only a subset of elements in which RXR au gments TR binding.