The transcriptional cofactor MCAF1/ATF7IP is involved in histone gene expression and cellular senescence.

Cellular senescence is post-mitotic or oncogene-induced events combined with nuclear remodeling. MCAF1 (also known as hAM or ATF7IP), a transcriptional cofactor that is overexpressed in various cancers, functions in gene activation or repression, depending on interacting partners. In this study, we found that MCAF1 localizes to PML nuclear bodies in human fibroblasts and non-cancerous cells. Interestingly, depletion of MCAF1 in fibroblasts induced premature senescence that was characterized by cell cycle arrest, SA-ß-gal activity, and senescence-associated heterochromatic foci (SAHF) formation. Under this condition, core histones and the linker histone H1 significantly decreased at both mRNA and protein levels, resulting in reduced nucleosome formation. Consistently, in activated Ras-induced senescent fibroblasts, the accumulation of MCAF1 in PML bodies was enhanced via the binding of this protein to SUMO molecules, suggesting that sequestration of MCAF1 to PML bodies promotes cellular senescence. Collectively, these results reveal that MCAF1 is an essential regulator of cellular senescence.

The role of methyl-binding proteins in chromatin organization and epigenome maintenance.

Methylated DNA can be specifically recognized by a set of proteins called methyl-CpG-binding proteins (MBPs), which belong to three different structural families in mammals: the MBD family, the Kaiso and Kaiso-like proteins and the SRA domain proteins. A current view is that, once bound to methylated DNA, MBPs translate the DNA methylation signal into appropriate functional states, through interactions with diverse partners. However, if some of the biological functions of MBPs have been widely described--notably transcriptional repression--others are poorly understood, and more generally the extent of MBP activities remains unclear. Here we propose to discuss the role of MBPs in two crucial nuclear events: chromatin organization and epigenome maintenance. Finally, important challenges for future research as well as for biomedical applications in pathologies such as cancers--in which DNA methylation patterns are widely altered--will be mentioned.

The human proteins MBD5 and MBD6 associate with heterochromatin but they do not bind methylated DNA.

BACKGROUND: MBD5 and MBD6 are two uncharacterized mammalian proteins that contain a putative Methyl-Binding Domain (MBD). In the proteins MBD1, MBD2, MBD4, and MeCP2, this domain allows the specific recognition of DNA containing methylated cytosine; as a consequence, the proteins serve as interpreters of DNA methylation, an essential epigenetic mark. It is unknown whether MBD5 or MBD6 also bind methylated DNA; this question has interest for basic research, but also practical consequences for human health, as MBD5 deletions are the likely cause of certain cases of mental retardation. PRINCIPAL FINDINGS: Here we report the first functional characterization of MBD5 and MBD6. We have observed that the proteins colocalize with heterochromatin in cultured cells, and that this localization requires the integrity of their MBD. However, heterochromatic localization is maintained in cells with severely decreased levels of DNA methylation. In vitro, neither MBD5 nor MBD6 binds any of the methylated sequences DNA that were tested. CONCLUSIONS: Our data suggest that MBD5 and MBD6 are unlikely to be methyl-binding proteins, yet they may contribute to the formation or function of heterochromatin. One isoform of MBD5 is highly expressed in oocytes, which suggests a possible role in epigenetic reprogramming after fertilization.

Sequence-specific recognition of methylated DNA by human zinc-finger proteins.

DNA methylation is an essential epigenetic mark. Three classes of mammalian proteins recognize methylated DNA: MBD proteins, SRA proteins and the zinc-finger proteins Kaiso, ZBTB4 and ZBTB38. The last three proteins can bind either methylated DNA or unmethylated consensus sequences; how this is achieved is largely unclear. Here, we report that the human zinc-finger proteins Kaiso, ZBTB4 and ZBTB38 can bind methylated DNA in a sequence-specific manner, and that they may use a mode of binding common to other zinc-finger proteins. This suggests that many other sequence-specific methyl binding proteins may exist.

Many paths to one goal? The proteins that recognize methylated DNA in eukaryotes.

DNA methylation is an epigenetically inherited chemical modification that is associated with transcriptional silencing and is essential for mammalian development. The DNA methylation signal is read out by methyl-CpG binding proteins (MBPs) that specifically bind to methylated DNA. Three structurally divergent families of MBPs have been identified so far: the MBD family, the SRA family and a family of proteins with Zinc fingers. In this review, we describe how the distinct families of methyl-CpG binding proteins have evolved, how they each recognize and maintain the DNA methylation mark, and finally how they turn this mark into biological effect.

The Drosophila jumonji gene encodes a JmjC-containing nuclear protein that is required for metamorphosis.

Jumonji (Jmj) is a transcriptional repressor that plays important roles in the suppression of cell proliferation and development of various tissues in the mouse. To further clarify the roles of Jmj during development and gain insight into mechanisms of Jmj-mediated transcriptional regulation, we have taken advantage of Drosophila as a model organism. Drosophila Jmj (dJmj) shares high homology with mammalian Jmj in the JmjN, JmjC and AT-rich interaction domains, as well as in the N-terminal repression domain. dJmj localizes to hundreds of euchromatic sites but not to chromocenter heterochromatin on salivary gland polytene chromosomes. In addition, dJmj is excluded from regions stained with an antibody against Ser5-phosphorylated RNA polymerase II, suggesting a function of dJmj in transcriptionally inactive chromatin. Loss of djmj results in larval and pupal lethality with phenotypes similar to those observed in mutants of ecdysone-regulated genes, implying the involvement of dJmj in the repression of gene expression in the ecdysone pathway. Transgenic mouse Jmj mostly colocalizes with dJmj and partially rescues the phenotypes of djmj mutants, indicating that dJmj is a functional homolog of mammalian Jmj. Furthermore, mutation in djmj suppresses position effect variegation of the T(2;3)Sb(V) rearrangement. These findings suggest that dJmj controls expression of developmentally important genes through modification of chromatin into a transcriptionally silenced state.

Identification of ZNF200 as a novel binding partner of histone H3 methyltransferase G9a.

G9a belongs to the subfamily of histone H3 lysine 9 (H3-K9)-specific methyltransferases. On amino acid sequence alignment of human and Drosophila G9a, we found that the N-terminal region from amino acids 532-605 to be evolutionarily conserved and named this the G9a homology domain (GHD). Using the GHD of human G9a (hG9a) as a bait, we isolated cDNA encoding a zinc finger protein 200 (ZNF200), which contains five C(2)H(2)-type zinc finger domains in tandem arrays. Interaction between G9a and ZNF200 could be demonstrated by in vitro binding assays and immunoprecipitation experiments using cultured human HEK293 cell extracts. GST pull-down assays using deletion derivatives of ZNF200 revealed that the interaction is through a region encompassing three of the five zinc finger domains. Furthermore, ZNF200 appear to co-localize with G9a in the nucleoplasm of HEK293 cells as discrete speckles. These results demonstrate that ZNF200 is a novel binding partner of G9a.

Identification of a novel BTB-zinc finger transcriptional repressor, CIBZ, that interacts with CtBP corepressor.

The transcriptional corepressor C-terminal binding protein (CtBP) is thought to be involved in development and oncogenesis, but the regulation of its corepressor activity is largely unknown. We show here that a novel BTB-zinc finger protein, CIBZ (CtBP-interacting BTB zinc finger protein; a mouse ortholog of rat ZENON that was recently identified as an e-box/dyad binding protein), redistributes CtBP to pericentromeric foci from a diffuse nuclear localization in interphase cells. CIBZ physically associates with CtBP via a conserved CtBP binding motif, PLDLR. When heterologously targeted to DNA, CIBZ represses transcription via two independent repression domains, an N-terminal BTB domain and a PLDLR motif-containing RD2 region, in a histone deacetylase-independent and -dependent manner, respectively. Mutation in the PLDLR motif abolishes the CIBZ-CtBP interaction and transcriptional repression activity of RD2, but does not affect the repression activity of the BTB domain. Furthermore, this PLDLR-mutated CIBZ cannot target CtBP to pericentromeric foci, although it is localized to the pericentromeric foci itself. These results suggest that at least one repression mechanism mediated by CIBZ is recruitment of the CtBP/HDAC complex to pericentromeric foci, and that CIBZ may regulate pericentromeric targeting of CtBP.