The nuclear receptor mouse retinoid X receptor ␣ (mRXR␣) was shown to be constitutively phosphorylated in its NH 2 -terminal A/B region, which contains potential phosphorylation sites for proline-directed Ser/ Thr kinases. Mutants for each putative site were generated and overexpressed in transfected COS-1 cells. Constitutively phosphorylated residues identified by tryptic phosphopeptide mapping included serine 22 located in the A1 region that is specific to the RXR␣1 isoform. Overexpression and UV activation of the stressactivated kinases, c-Jun NH 2 -terminal kinases 1 and 2 (JNK1 and JNK2), hyperphosphorylated RXR␣, resulting in a marked decrease in its electrophoretic mobility. This inducible hyperphosphorylation involved three residues (serines 61 and 75 and threonine 87) in the B region of RXR␣ and one residue (serine 265) in the ligand binding domain (E region). Binding assays performed in vitro with purified recombinant proteins demonstrated that JNKs did not interact with RXR␣ but bound to its heterodimeric partners, retinoic acid receptors ␣ and ␥ (RAR␣ and RAR␥). Hyperphosphorylation by JNKs did not affect the transactivation properties of either RXR␣ homodimers or RXR␣/RAR␣ heterodimers in transfected cultured cells.Retinoids are derivatives of vitamin A that play key roles in a variety of biological processes ranging from pattern formation and organogenesis during embryogenesis to maintenance of homeostasis in the adult (1-4). Retinoids exert their pleiotropic effects through two classes of nuclear receptors acting as liganddependent transcriptional regulators, the retinoic acid receptors (RARs) 1 and the retinoid X receptors (RXRs) (4 -7). RARs are activated by both all-trans-retinoic acid (tRA) and 9-cisretinoic acid (9cRA), whereas RXRs are activated exclusively by 9cRA. There are three RAR isotypes and three RXR isotypes (␣, , and ␥), encoded by distinct genes, and for each isotype, there are at least two main isoforms, which differ in their NH 2 -terminal A regions and are generated by differential promoter usage and/or alternative splicing (5,6,8). Several lines of evidence support the conclusion that RAR/RXR heterodimers are the functional units transducing the retinoid signal in vivo (Refs. 4 -6, 9, and 10 and references therein). However, RXRs are also able to heterodimerize with other members of the nuclear receptor superfamily, such as the thyroid hormone receptors, the vitamin D 3 receptor, and the peroxisome proliferator activated receptors (11-15).As other members of the nuclear steroid/thyroid hormone receptor superfamily, RARs and RXRs exhibit a conserved modular structure with six variably conserved functional regions (A to F) ( Fig. 1 and Refs. 5 and 6). The amino-terminal A/B region of RARs and RXRs contains a ligand-independent transactivation function AF-1 (16, 17), while the highly conserved C region is included in the DNA-binding domain. The E region is more complex, since, in addition to the ligand-binding domain, it contains a dimerization surface and a ligand-dependent tra...