SUMO modification enhances p66-mediated transcriptional repression of the Mi-2/NuRD complex

Human p66α and p66α are two potent transcriptional repressors that interact with the methyl-CpG-binding domain proteins MBD2 and MBD3. An analysis of the molecular mechanisms mediating repression resulted in the identification of two major repression domains in p66α and one in p66β. Both p66α and p66β are SUMO-modified in vivo: p66α at two sites (Lys-30 and Lys-487) and p66β at one site (Lys-33). Expression of SUMO1 enhanced the transcriptional repression activity of Gal-p66α and Gal-p66β. Mutation of the SUMO modification sites or using a SUMO1 mutant or a dominant negative Ubc9 ligase resulted in a significant decrease of the transcriptional repression of p66α and p66β. The Mi-2/NuRD components MBD3, RbAp46, RbAp48, and HDAC1 were found to bind to both p66α and p66β in vivo. Most of the interactions were not affected by the SUMO site mutations in p66α or p66β, with two exceptions. HDAC1 binding to p66α was lost in the case of a p66αK30R mutant, and RbAp46 binding was reduced in the case of a p66βK33R mutant. These results suggest that interactions within the Mi-2/NuRD complex as well as optimal repression are mediated by SUMOylation.