Title Isolation and characterization of a novel DNA methyltransferase complex linking DNMT 3 B with components of the mitotic chromosome condensation machinery Permalink

Proper patterns of genome-wide DNA methylation, mediated by DNA methyltransferases DNMT1, -3A and -3B, are essential for embryonic development and genomic stability in mammalian cells. The de novo DNA methyltransferase DNMT3B is of particular interest because it is frequently overexpressed in tumor cells and is mutated in immunode®ciency, centromere instability and facial anomalies (ICF) syndrome. In order to gain a better understanding of DNMT3B, in terms of the targeting of its methylation activity and its role in genome stability, we biochemically puri®ed endogenous DNMT3B from HeLa cells. DNMT3B co-puri®es and interacts, both in vivo and in vitro, with several components of the condensin complex (hCAP-C, hCAP-E and hCAP-G) and KIF4A. Condensin mediates genome-wide chromosome condensation at the onset of mitosis and is critical for proper segregation of sister chromatids. KIF4A is proposed to be a motor protein carrying DNA as cargo. DNMT3B also interacts with histone deacetylase 1 (HDAC1), the co-repressor SIN3A and the ATP-dependent chromatin remodeling enzyme hSNF2H. Furthermore, DNMT3B co-localizes with condensin and KIF4A on condensed chromosomes throughout mitosis. These studies therefore reveal the ®rst direct link between the machineries regulating DNA methylation and mitotic chromosome condensation in mammalian cells. INTRODUCTION Chromatin structure, at both the local and whole-chromosome levels, is determined by many factors. One of the key regulators of chromatin structure is DNA methylation. Methylation of cytosine within the CpG dinucleotide in mammalian cells is an epigenetic modi®cation of the DNA that has been shown to be important for diverse processes such as transcriptional regulation, embryonic development, genomic imprinting and genome stability (1±4). DNA methylation patterns are established and maintained by three DNA methyltransferases (DNMTs): DNMT1, -3A and -3B (5,6). DNMT1 has traditionally been regarded as a maintenance methyltransferase specialized for copying DNA methylation patterns following DNA replication, whereas DNMT3A and DNMT3B are believed to be involved primarily with de novo methylation, particularly during embryonic development (3). DNA methylation is not randomly distributed throughout the mammalian genome. Rather, it tends to be concentrated in transposable elements (parasitic DNA), repeats and pericentromeric heterochromatin, and contributes to the transcriptionally repressed, highly condensed chromatin structure characteristic of these regions (7±9). DNA methylation is also believed to suppress mitotic recombination events involving highly repetitive regions (10±12). It remains unclear exactly how DNA methylation is `targeted' to select regions of the genome; however, emerging evidence indicates that other epigenetic modi®cations, particularly acetylation and methylation of the histone H3 and H4 N-terminal tails, determine which regions of the genome will be methylated. The core histone tails are extensively modi®ed by an ever-growing number of histone acetyltransferases and histone methylases (13±15). Establishment and maintenance of regional DNA methylation patterns is most likely accomplished by protein± *To whom correspondence should be addressed at present address: University of Florida, Department of Biochemistry and Molecular Biology, Box 100245, Gainesville, FL 32610, USA. Tel: +1 301 594 9509; Fax: +1 301 496 4951; Email: keithr@u ̄. edu 2716±2729 Nucleic Acids Research, 2004, Vol. 32, No. 9 DOI: 10.1093/nar/gkh589 Nucleic Acids Research, Vol. 32 No. 9 ã Oxford University Press 2004; all rights reserved Published online May 17, 2004