Exogenous expression of a dominant negative RORα1 vector in muscle cells impairs differentiation: RORα1 directly interacts with p300 and MyoD

ROR/RZR is an orphan nuclear receptor that has no known ligand in the ‘classical sense’. In the present study we demonstrate that RORα is constitutively expressed during the differentiation of proliferating myoblasts to post-mitotic multinucleated myotubes, that have acquired a contractile phenotype. Exogenous expression of dominant negative RORα1∆E mRNA in myogenic cells significantly reduces the endogenous expression of RORα1 mRNA, represses the accumulation and delays the activation of mRNAs encoding MyoD and myogenin [the muscle-specific basic helix– loop–helix (bHLH) proteins] and p21Waf-1/Cip-1 (a cdk inhibitor). Immunohistochemistry demonstrates that morpho-logical differentiation is delayed in cells expressing the ROR∆E transcript. Furthermore, the size and development of mutlinucleated myotubes is impaired. The E region of RORα1 interacts with p300, a cofactor that functions as a coactivator in nuclear receptor and MyoD-mediated transactivation. Consistent with the functional role of RORα1 in myogenesis, we observed that RORα1 directly interacts with the bHLH protein MyoD. This interaction was mediated by the N-terminal activation domain of the bHLH protein, MyoD, and the RORα1 DNA binding domain/C region. Furthermore, we demonstrated that p300, RORα1 and MyoD interact in a non-competitive manner. In conclusion, this study provides evidence for a biological role and positive influence of RORα1 in the cascade of events involved in the activation of myogenic-specific markers and cell cycle regulators and suggests that crosstalk between the retinoid-related orphan (ROR) nuclear receptors and the myogenic bHLH proteins has functional consequences for differentiation.