The Nuclear Receptor Corepressor N-CoR Regulates Differentiation: N-CoR Directly Interacts with MyoD

Abstract Classical ligand-activated nuclear receptors (e.g. thyroid hormone receptor, retinoic acid receptor), orphan nuclear receptors (e.g. Rev-erbAα/β), Mad/Max bHLH (basic helix loop helix)-LZ proteins, and oncoproteins, PLZF and LAZ3/BCL6, bind DNA and silence transcription by recruiting a repressor complex that contains N-CoR (nuclear receptor corepressor)/SMRT (silencing mediator of retinoic acid and thyroid hormone receptor), Sin3A/B, and HDAc-1/-2 proteins. The function of the corepressor, N-CoR, in the process of cellular differentiation and coupled phenotypic acquisition, has not been investigated. We examined the functional role of N-CoR in myogenesis (muscle differentiation), an ideal paradigm for the analysis of the determinative events that govern the cell’s decision to divide or differentiate. We observed that the mRNA encoding N-CoR was suppressed as proliferating myoblasts exited the cell cycle, and formed morphologically and biochemically differentiated myotubes. Exogenous expression of N-CoR (but not RIP13) in myogenic cells ablated 1) myogenic differentiation, 2) the expression of the myoD gene family that encode the myogenic specific bHLH proteins, and 3) the crucial cell cycle regulator, p21Waf-1/Cip-1 mRNA. Furthermore, N-CoR expression efficiently inhibits the myoD-mediated myogenic conversion of pluripotential C3H10T1/2 cells. We demonstrate that MyoD-mediated transactivation and activity are repressed by N-CoR. The mechanism involves direct interactions between MyoD and N-CoR; moreover, the interaction was dependent on the amino-terminal repression domain (RD1) of N-CoR and the bHLH region of MyoD. Trichostatin A treatment significantly stimulated the activity of MyoD by approximately 10-fold and inhibited the ability of N-CoR to repress MyoD-mediated transactivation, consistent with the involvement of the corepressor and the recruitment of a histone deacteylase activity in the process. This work demonstrates that the corepressor N-CoR is a key regulator of MyoD activity and mammalian differentiation, and that N-CoR has a multifaceted role in myogenesis.

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Response of SMRT (Silencing Mediator of Retinoic Acid and Thyroid Hormone Receptor) and N-CoR (Nuclear Receptor Corepressor) Corepressors to Mitogen-Activated Protein Kinase Kinase Kinase Cascades Is Determined by Alternative mRNA Splicing

Molecular Endocrinology ◽

10.1210/me.2007-0035 ◽

2007 ◽

Vol 21 (8) ◽

pp. 1924-1939 ◽

Cited By ~ 25

Author(s):

Brian A. Jonas ◽

Natalia Varlakhanova ◽

Fumihiko Hayakawa ◽

Michael Goodson ◽

Martin L. Privalsky

Keyword(s):

Retinoic Acid ◽

Thyroid Hormone ◽

Nuclear Receptor ◽

Hormone Receptor ◽

Thyroid Hormone Receptor ◽

Splice Variants ◽

Mrna Splicing ◽

Alternative Mrna Splicing ◽

Nuclear Receptor Corepressor ◽

Erk Kinase

Abstract The SMRT (silencing mediator of retinoic acid and thyroid hormone receptor) and N-CoR (nuclear receptor corepressor) corepressors are important mediators of transcriptional repression by nuclear hormone receptors. SMRT is regulated by MAPK kinase kinase (MAPKKK) cascades that induce its release from its receptor partners, its export from nucleus to cytoplasm, and derepression of target gene expression. Intriguingly, the otherwise closely related N-CoR is refractory to MAPKKK signaling under the same conditions. However, both SMRT and N-CoR are expressed as a series of alternatively spliced protein variants differing in structure and function. We have now characterized the impact of this alternative mRNA splicing on the corepressor response to MAPKKK signaling. Whereas the SMRTα, SMRTτ, and SMRTsp2 splice variants are released from their nuclear receptor partners in response to MAPKKK activation, the SMRTsp18 variant, which resembles N-CoR in its overall molecular architecture, is relatively refractory to this kinase-induced release. Alternative splicing of N-CoR, in contrast, had only minimal effects on the resistance of this corepressor to MAPKKK inhibition. Notably, all of the SMRT splice variants examined redistributed from nucleus to cytoplasm in response to MAPKKK cascade signaling, but none of the N-CoR splice variants did so. Different tiers of the MAPKKK cascade hierarchy contributed to these different aspects of corepressor regulation, with MAP/ERK kinase kinase 1 and MAP/ERK kinase 1 regulating subcellular redistribution and ERK2 regulating nuclear receptor-corepressor interaction. We conclude that cells can customize their transcriptional response to MAPKKK cascade signaling by selective expression of the SMRT or N-CoR locus, by selective utilization of a specific corepressor splice variant, and by selective exploitation of specific tiers of the MAPK cascade.

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A novel orphan receptor specific for a subset of thyroid hormone-responsive elements and its interaction with the retinoid/thyroid hormone receptor subfamily.

Molecular and Cellular Biology ◽

10.1128/mcb.14.10.7025 ◽

1994 ◽

Vol 14 (10) ◽

pp. 7025-7035 ◽

Cited By ~ 242

Author(s):

R Apfel ◽

D Benbrook ◽

E Lernhardt ◽

M A Ortiz ◽

G Salbert ◽

...

Keyword(s):

Amino Acid ◽

Thyroid Hormone ◽

Dna Binding ◽

Nuclear Receptors ◽

Nuclear Receptor ◽

Dna Sequences ◽

Hormone Receptor ◽

Thyroid Hormone Receptor ◽

Orphan Receptor ◽

Retinoid Receptor

The steroid/hormone nuclear receptor superfamily comprises several subfamilies of receptors that interact with overlapping DNA sequences and/or related ligands. The thyroid/retinoid hormone receptor subfamily has recently attracted much interest because of the complex network of its receptor interactions. The retinoid X receptors (RXRs), for instance, play a very central role in this subfamily, forming heterodimers with several receptors. Here we describe a novel member of this subfamily that interacts with RXR. Using a v-erbA probe, we obtained a cDNA which encodes a novel 445-amino-acid protein, RLD-1, that contains the characteristic domains of nuclear receptors. Northern (RNA) blot analysis showed that in mature rats, the receptor is highly expressed in spleen, pituitary, lung, liver, and fat. In addition, weaker expression is observed in several other tissues. Amino acid sequence alignment and DNA-binding data revealed that the DNA-binding domain of the new receptor is related to that of the thyroid/retinoid subgroup of nuclear receptors. RLD-1 preferentially binds as a heterodimer with RXR to a direct repeat of the half-site sequence 5'-G/AGGTCA-3', separated by four nucleotides (DR-4). Surprisingly, this binding is dependent to a high degree on the nature of the spacing nucleotides. None of the known nuclear receptor ligands activated RLD-1. In contrast, a DR-4-dependent constitutive transcriptional activation of a chloramphenicol acetyltransferase reporter gene by the RLD-1/RXR alpha heterodimer was observed. Our data suggest a highly specific role for this novel receptor within the network of gene regulation by the thyroid/retinoid receptor subfamily.

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Novel Mechanism of Nuclear Receptor Corepressor Interaction Dictated by Activation Function 2 Helix Determinants

Molecular and Cellular Biology ◽

10.1128/mcb.22.19.6831-6841.2002 ◽

2002 ◽

Vol 22 (19) ◽

pp. 6831-6841 ◽

Cited By ~ 69

Author(s):

Anna N. Moraitis ◽

Vincent Giguère ◽

Catherine C. Thompson

Keyword(s):

Retinoic Acid ◽

Thyroid Hormone ◽

Ligand Binding ◽

Nuclear Receptor ◽

Thyroid Hormone Receptor ◽

Activation Function ◽

Binding Domain ◽

Ligand Binding Domain ◽

Cerebellar Development ◽

Nuclear Receptor Corepressor

ABSTRACT Transcriptional regulation by nuclear receptors is controlled by the concerted action of coactivator and corepressor proteins. The product of the thyroid hormone-regulated mammalian gene hairless (Hr) was recently shown to function as a thyroid hormone receptor corepressor. Here we report that Hr acts as a potent repressor of transcriptional activation by RORα, an orphan nuclear receptor essential for cerebellar development. In contrast to other corepressor-nuclear receptor interactions, Hr binding to RORα is mediated by two LXXLL-containing motifs, a mechanism associated with coactivator interaction. Mutagenesis of conserved amino acids in the ligand binding domain indicates that RORα activity is ligand-dependent, suggesting that corepressor activity is maintained in the presence of ligand. Despite similar recognition helices shared with coactivators, Hr does not compete for the same molecular determinants at the surface of the RORα ligand binding domain, indicating that Hr-mediated repression is not simply through displacement of coactivators. Remarkably, the specificity of Hr corepressor action can be transferred to a retinoic acid receptor by exchanging the activation function 2 (AF-2) helix. Repression of the chimeric receptor is observed in the presence of retinoic acid, demonstrating that in this context, Hr is indeed a ligand-oblivious nuclear receptor corepressor. These results suggest a novel molecular mechanism for corepressor action and demonstrate that the AF-2 helix can play a dynamic role in controlling corepressor as well as coactivator interactions. The interaction of Hr with RORα provides direct evidence for the convergence of thyroid hormone and RORα-mediated pathways in cerebellar development.

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