p66  and p66  of the Mi-2/NuRD complex mediate MBD2 and histone interaction

Abstract

The Mi-2/NuRD complex is a multi-subunit protein complex with enzymatic activities involving chromatin remodeling and histone deacetylation. Targeting of Mi-2/NuRD to methylated CpG sequences mediates gene repression. The function of p66α and of p66β within the multiple subunits has not been addressed. Here, we analyzed the in vivo function and binding of both p66-paralogs. Both factors function in synergy, since knocking-down p66α affects the repressive function of p66β and vice versa. Both proteins interact with MBD2 functionally and biochemically. Mutation of a single amino acid of p66α abolishes in vivo binding to MBD2 and interferes with MBD2-mediated repression. This loss of binding results in a diffuse nuclear localization in contrast to wild-type p66α that shows a speckled nuclear distribution. Furthermore, wild-type subnuclear distribution of p66α and p66β depends on the presence of MBD2. Both proteins interact with the tails of all octamer histones in vitro, and acetylation of histone tails interferes with p66 binding. The conserved region 2 of p66α is required for histone tail interaction as well as for wild-type subnuclear distribution. These results suggest a two-interaction forward feedback binding mode, with a stable chromatin association only after deacetylation of the histones has occurred.

The human Mi-2/NuRD complex and gene regulation

“…However, subsequent analysis has left little doubt that MBD2, like MBD3, is biochemically linked to the other subunits of Mi-2/NuRD. Several groups have since described the biochemical identification of both MBD2 and MBD3 containing forms of the complex (Feng and Zhang, 2001;Le Guezennec et al, 2006), and a direct interaction between MBD2 and p66, another component of the Mi-2/NuRD complex, has been documented (Brackertz et al, 2002(Brackertz et al, , 2006. These experiments have served to clarify the relationship between Mi-2/NuRD and the MBD family, indicating that MBD2 and MBD3 are alternative subunits.…”

“…As mentioned above, the p66 subunits interact with the MBD2 subunit and may be involved in interactions of the complex with methylated DNA (Feng and Zhang, 2001;Brackertz et al, 2006). Like the RbA p46/48 subunits, both p66 isoforms have the capacity to interact directly with core histones (Brackertz et al, 2006).…”

Time-resolved Global and Chromatin Proteomics during Herpes Simplex Virus Type 1 (HSV-1) Infection

“…S3), which show respectively monotonic (steady) or early (at 3 hpi) decrease in protein abundance in the cell after HSV-1 infection. Interestingly, within these two clusters, we detected a number of proteins involved in regulation of methylation of lysine at position 4 of histone H3, particularly tri-methylation (H3K4me3) (DPY30, WDR82 or OGT) (39 -41); histone H2A mono-ubiquitylation (SKP1) (42); or proteins facilitating transcriptional repression (CTBP2, GATAD2B, LOXL2) (43)(44)(45)(46)(47)(48). Furthermore, cluster 15 contained transcriptional regulator ATRX protein, which in complex with DAXX and histone H3.3 is important for heterochromatin silencing at multiple genomic regions (49).…”

The Chromatin Remodeling Factor Mi-2α Acts as a Novel Co-activator for Human c-Myb

“…Conflicting evidence exists regarding the composition of the NuRD complex (17), probably reflecting its heterogeneity as well as its tissue and target specificity. In short, the consensus NuRD complex consists of the core subunits: HDAC1 and -2, RbAp46 and -48, Mi-2␣ and/or -␤ (18 -20), one or more MTA proteins (MTA1, -2, or -3, or splice variants of these) (21), MBD2 or MBD3 (22), and often p66␣ and/or -␤ (23).…”