Characterization of the repressor function of the nuclear orphan receptor retinoid receptor-related testis-associated receptor/germ cell nuclear factor.
Retinoid receptor-related testis-associated receptor (RTR)/germ cell nuclear factor is a nuclear orphan receptor that plays an important role in the control of gene expression during early embryonic development and gametogenesis. It has been shown to repress transcriptional activation. In this study, we further characterize this repressor function. We demonstrate that RTR can suppress the transcriptional activation induced by the estrogen receptor related-receptor alpha1 through its response element. The latter is at least in part due to competition for binding to the same response element. In addition, RTR inhibits basal transcriptional activation, indicating that it functions as an active repressor. Mammalian two-hybrid analyses showed that RTR interacts with the co-repressor nuclear co-repressor (N-CoR) but is unable to interact with the co-repressor SMRT or RIP140. Pull-down analyses with glutathione S-transferase-RTR fusion protein demonstrated that RTR physically interacts with N-CoR in vitro, suggesting a potential role for N-CoR in the transcriptional repression by RTR. To identify the regions in RTR essential for the binding of RTR to N-CoR, the effect of various deletion and point mutations on this interaction was examined. This analysis revealed that this interaction requires the hinge domain, helix 3 as well as the helix 12 region of RTR. The residues Ser(246)-Tyr(247) in the hinge domain, Lys(318) in helix 3, and Lys(489)-Thr(490) in helix 12 are identified as being critical in this interaction. Our results demonstrate that RTR can function as an active transcriptional repressor and that this repression can be mediated through interactions with the co-repressor N-CoR. We show that this interaction exhibits several characteristics unique to RTR. Through its repressor function, RTR can suppress the induction of transcriptional activation by other nuclear receptors. These repressor activities may provide important mechanisms by which RTR regulates gene expression during development and gametogenesis.
Yan
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Cell Biology Section, Laboratory of Pulmonary Pathobiology, NIEHS, National Institutes of Health, Research Triangle Park, North Carolina 27709, USA.
Jetten
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D048868:Adaptor Proteins, Signal Transducing; D000595:Amino Acid Sequence; D000818:Animals; D016466:CHO Cells; D002467:Cell Nucleus / Q000378:metabolism; D006224:Cricetinae; D004268:DNA-Binding Proteins / Q000737:chemistry* / Q000378:metabolism*; D004591:Electrophoresis, Polyacrylamide Gel; D017353:Gene Deletion; D018507:Gene Expression Regulation, Developmental; D005982:Glutathione Transferase / Q000378:metabolism; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009687:Nuclear Proteins / Q000378:metabolism; D056971:Nuclear Receptor Co-Repressor 1; D056985:Nuclear Receptor Co-Repressor 2; D000078723:Nuclear Receptor Interacting Protein 1; D057136:Nuclear Receptor Subfamily 6, Group A, Member 1; D010957:Plasmids / Q000378:metabolism; D017354:Point Mutation; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D018160:Receptors, Cytoplasmic and Nuclear / Q000737:chemistry* / Q000378:metabolism*; D011960:Receptors, Estrogen / Q000378:metabolism; D018168:Receptors, Retinoic Acid / Q000737:chemistry* / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D012097:Repressor Proteins / Q000378:metabolism; D020218:Response Elements; D017386:Sequence Homology, Amino Acid; D014158:Transcription, Genetic; D015533:Transcriptional Activation; D020798:Two-Hybrid System Techniques; D000097983:ERRalpha Estrogen-Related Receptor
D048868:Adaptor Proteins, Signal Transducing; D004268:DNA-Binding Proteins; D005982:Glutathione Transferase; D009687:Nuclear Proteins; D056971:Nuclear Receptor Co-Repressor 1; D056985:Nuclear Receptor Co-Repressor 2; D000078723:Nuclear Receptor Interacting Protein 1; D057136:Nuclear Receptor Subfamily 6, Group A, Member 1; D018160:Receptors, Cytoplasmic and Nuclear; D011960:Receptors, Estrogen; D018168:Receptors, Retinoic Acid; D011993:Recombinant Fusion Proteins; D012097:Repressor Proteins; D000097983:ERRalpha Estrogen-Related Receptor