Effects of chromatin structure on the enzymatic and DNA binding functions of DNA methyltransferases DNMT1 and Dnmt3a in vitro.
DNA methylation is an epigenetic modification of the genome critical for numerous processes, including transcriptional repression and maintenance of chromatin structure. Recent studies have revealed connections between DNA methylation and other epigenetic modifications such as ATP-dependent chromatin remodeling. It remains unclear, however, exactly how chromatin and epigenetic chromatin modifications affect the biological properties of the DNA methyltransferases (DNMT1, DNMT3A, and DNMT3B). Using a highly purified system and the 5S rDNA gene as free DNA or assembled into a mononucleosome, we have compared the effects of chromatin structure on DNMT1 and Dnmt3a. The catalytic efficiency for both enzymes decreased on the mononucleosome, approximately 8-fold for DNMT1 and 17-fold for Dnmt3a. DNMT1 and Dnmt3a bound to DNA and mononucleosomal substrates in gel shift experiments with approximately equal affinity and in a cooperative manner. We also show that DNMT1 interacts with hSNF2H chromatin remodeling enzyme and that DNMT1 binds mononucleosomes with higher affinity in the presence of hSNF2H. These findings raise interesting implications about the interactions of mammalian DNA methyltransferases with chromatin and provide the first evidence that a chromatin remodeling enzyme can alter the biological properties of a DNMT.
Epigenetic Gene Regulation and Cancer Section, LRBGE/NCI/NIH, Bldg. 41, Rm. C306, 41 Library Dr., Bethesda, MD 20892, USA. keithr@ufl.edu
Andrea K
AK
Robertson
Theresa M
TM
Geiman
Umesh T
UT
Sankpal
Gordon L
GL
Hager
Keith D
KD
Robertson
D001665:Binding Sites; D002843:Chromatin / Q000737:chemistry* / Q000648:ultrastructure*; D004247:DNA / Q000737:chemistry*; D000076123:DNA (Cytosine-5-)-Methyltransferase 1; D004248:DNA (Cytosine-5-)-Methyltransferases / Q000737:chemistry*; D019175:DNA Methylation; D004268:DNA-Binding Proteins / Q000737:chemistry; D004789:Enzyme Activation; D007700:Kinetics; D009707:Nucleosomes / Q000737:chemistry; D013329:Structure-Activity Relationship
D002843:Chromatin; D004268:DNA-Binding Proteins; D009707:Nucleosomes; D004247:DNA; D000076123:DNA (Cytosine-5-)-Methyltransferase 1; D004248:DNA (Cytosine-5-)-Methyltransferases; C000619380:DNMT1 protein, human; C112218:TRDMT1 protein, human