LSH Cooperates with DNA Methyltransferases To Repress Transcription

Abstract

LSH, a protein related to the SNF2 family of chromatin-remodeling ATPases, is required for efficient DNA methylation in mammals. How LSH functions to support DNA methylation and whether it associates with a large protein complex containing DNA methyltransferase (DNMT) enzymes is currently unclear. Here we show that, unlike many other chromatin-remodeling ATPases, native LSH is present mostly as a monomeric protein in nuclear extracts of mammalian cells and cannot be detected in a large multisubunit complex. However, when targeted to a promoter of a reporter gene, LSH acts as an efficient transcriptional repressor. Using this as an assay to identify proteins that are required for LSH-mediated repression we found that LSH cooperates with the DNMTs DNMT1 and DNMT3B and with the histone deacetylases (HDACs) HDAC1 and HDAC2 to silence transcription. We show that transcriptional repression by LSH and interactions with HDACs are lost in DNMT1 and DNMT3B knockout cells but that the enzymatic activities of DNMTs are not required for LSH-mediated silencing. Our data suggest that LSH serves as a recruiting factor for DNMTs and HDACs to establish transcriptionally repressive chromatin which is perhaps further stabilized by DNA methylation at targeted loci.In vertebrate genomes, DNA methylation patterns are established during gametogenesis, embryo development, and cell differentiation by enzymes of the DNA cytosine methyltransferase family, which includes the maintenance DNA methyltransferase (DNMT) DNMT1 and the de novo methyltransferases DNMT3A and DNMT3B (1,24,45). DNMT1 binds to PCNA and functions primarily during S phase to restore fully methylated CpGs on hemimethylated daughter DNA strands generated during DNA replication (3, 4). DNMT3A and -3B are able to methylate unmethylated DNA and in mouse embryogenesis are required during gastrulation, when DNA methylation patterns are established in differentiating cell lineages of the embryo (24). Mice lacking DNMT proteins die early during embryogenesis and display aberrant expression of retrotransposons and various imprinted and nonimprinted genes (14,18,24). Genetic studies with plants and mammals have revealed that additional factors besides DNMTs are required for the establishment of DNA methylation patterns in vivo. Loss-of-function mutations in SNF2 family-related putative chromatin-remodeling ATPases such as the Arabidopsis thaliana DDM1 (decrease in DNA methylation 1) protein and its murine homolog Lsh (lymphoid-specific helicase) lead to dramatic hypomethylation of the genome in Arabidopsis thaliana and mice, respectively (5, 16). Unlike animals that are null for DNMTs, Lsh-deficient mice develop to term but die soon after birth with symptoms of renal failure (5). Interestingly, mice expressing a hypomorph allele of Lsh with targeted disruption of the SNF2 domain survive much longer and display modest hypomethylation of DNA and premature aging (39). Collectively, these studies indicate that the low levels of DNA methylation (ϳ30 to 35% of the wild-type level) in Lsh...

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Downstream targets of FOXM1: CEP55 and HELLS are cancer progression markers of head and neck squamous cell carcinoma

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Repeat elements and the Arabidopsis DNA methylation landscape

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