EVI1 Is a Transcriptional Repressor Able To Recruit SUV39H1 Histone Methylation Activity.

Abstract

The Evi1 (Ecotropic viral integration site 1) gene is located on chromosome 3q26 and encodes a highly conserved nuclear protein that belongs to the Kruppel family of transcription factors, containing two Cys2-His2 repeat zinc finger motifs domains. EVI1 is not expressed in normal hematopoietic tissues, but can be found overexpressed in acute myeloid leukemia/myelodysplastic syndrome (AML/MDS) cells as a consequence of chromosome 3 rearrangements. In vitro and in vivo studies have shown that a number of biological properties can be attributed to the EVI1 protein that might contribute to leukemogenesis: deregulation of cell proliferation, inhibition of TGF-ß signaling pathway, and suppression of stress induced apoptosis. We and others reported that EVI1 acts as a transcriptional repressor through its interaction with a subset of co-repressor proteins. More recently, we showed that EVI1 directly interacts with RUNX1 and GATA1 transcription factors. In both cases this interaction alters the ability of the transcription factors to bind their recognition sites on the DNA, leading to the deregulation of their target genes and to the impairment of the normal hematopoietic development. Interestingly, EVI1-mediated transcriptional silencing in association with HDAC proteins can be only partially rescued by HDAC inhibitors, suggesting that additional chromosomal modification might occur for gene silencing. Methylation by histone methyltransferase proteins (HMTs) is one type of these modifications, and when it occurs on certain residues can lead both to repression or activation of gene expression. The SUV39H1 (Suppressor of Variegation 3–9 Homolog 1) gene encodes a histone 3 specific methyltransferase. SUV39H1 selectively methylates lysine 9 of the amino terminus of the histone 3. Tri-methylation of histone H3 is important for recruiting heterochromatin protein 1 (HP1), thereby regulating gene expression, chromatin packaging and heterochromatin formation. Here we investigate the hypothesis that histone methylation may contribute to the transcriptional repressive potential of EVI1. We found that EVI1 physically interacts with the histone methyltransferase SUV39H1. The interaction between the two proteins requires the N-terminus of EVI1 where the proximal zinc finger domain is located; in particular, out of the seven motifs that compose this domain, the distal three motifs seem to be responsible for the binding between the two proteins. By pull-down assay with in vitro translated EVI1 this interaction appears to be direct. Futhermore, using in vitro histone methylation assay we show that EVI1 forms a complex with SUV39H1 that is able to methylate a recombinant H3, suggesting that the EVI1-SUV39H1 interaction does not affect the histone methyltransferase activity of SUV39H1. We suggest here that EVI1 interacts with SUV39H1 and that this binding could have a role in the epigenetic remodellig of the chromatin. According to this model, it is possible that EVI1 recruits histone methylation activity to selected genomic loci to silence tumor suppressor genes and repress differentiation factors. Because lysine methylation by SUV39H1 cannot take place on an already acetylated lysine, deacetylation activity associated with EVI1, through its interaction with HDACs, becomes a necessary preceeding step to SUV39H1 mediated histone methylation.