IDENTIFICATION OF A REGULATORY FUNCTION FOR AN ORPHAN RECEPTOR IN MUSCLE - COUP-TF-II AFFECTS THE EXPRESSION OF THE MYOD GENE FAMILY DURINGMYOGENESIS

COUP-TF II is an 'orphan steroid receptor' that binds a wide variety o f AGGTCA repeats and represses thyroid hormone (T-3) and retinoid depe ndent trans-activation; however, very little is known of its functiona l and/or developmental role during mammalian cell differentiation. T-3 and retinoids have been demonstrated to promote terminal muscle diffe rentiation via activation of the muscle specific myoD gene family (myo D, myogenin, myf-5 and MRF-4). The myoD gene family can direct the fat e of mesodermal cell lineages, repress proliferation, activate differe ntiation and the contractile phenotype. Hence, we investigated the exp ression and functional role of COUP-TF II during muscle differentiatio n. Proliferating C2C12 myoblasts expressed COUP-TF II mRNA which was r epressed when cells were induced to differentiate into post-mitotic mu ltinucleated myotubes by serum withdrawal. Concomitant with the decrea se of COUP-TF II mRNA was the appearance of muscle specific mRNAs (e.g . myogenin, alpha-actin). We show that Escherichia coli expressed full length and truncated COUP-TF II bound in a sequence specific manner t o the T-3 response elements (TREs) in the myoD and myogenin regulatory HLH genes [Olson (1992) Dev. Biol. 154, 261-272]; and the TRE in the skeletal a-actin contractile protein gene. COUP-TF II diminished the h omodimeric binding of the thyroid hormone receptor and the heterodimer ic binding of thyroid hormone and retinoid X receptor complexes to the se TREs. Constitutive over-expression of COUP-TF II cDNA in mouse C2C1 2 myogenic cells suppressed the levels of myoD mRNA and blocked the in duction of myogenin mRNA, whereas constitutive expression of anti-sens e COUP-TF II cDNA significantly increased the steady state levels of m yoD mRNA and hyper-induced myogenin mRNA. These studies demonstrate fo r the first time (i) that COUP-TF II, functions as a physiologically r elevant antagonistic regulator of myogenesis via direct effects on the myoD gene family and (ii) direct evidence for the developmental role of COUP-TF II during mammalian cell differentiation.