Repression of Nrf1-Mediated Transactivation by MCRS2

Nrfl [p45 nuclear factor-erythroid 2 (p45 NF-E2)- related factor 1], a member of the CNC-bZIP (CNC basic region leucine zipper) family, is known to be a transcriptional activator via dimerization with distinct partners, such as Maf, FosB, c-Jun, JunD, etc. To find other proteins interacting with the CNC bZIP domain, the CNC-bZIP region of Nrfl was used as a bait in a yeast two-hybrid screening. MCRS2, a splicing variant of p78/MCRSl, was isolated as the Nrfl- interacting partner. The interaction between Nrfl and MCRS2 was confirmed in vitro by GST pull-down assays and in vivo by co-immunoprecipitation. The Nrfl-MCRS2 interaction domains were mapped to be the residues 354-447 of Nrfl and the residues 314-475 of MCRS2 respectively by yeast two- hybrid and GST pull- down assays. To further confirm the biological relevance, a reporter driven by CNC-bZIP protein binding sites was also shown to be repressed by MCRS2 in transient transfection assay. And a reporter gene activated by LexA-Nrfl was specifically repressed by MCRS2. Taken together, these results pointed to a new Nrfl-interacting protein MCRS2 that can mediate repression effect on Nrfl- mediated transcriptional activation. Until now, this is the first repressor protein on Nrfl transactivation activity and there is no other repressor (s) reported before.