Dynamics of Dnmt1 interaction with the replication machinery and its role in postreplicative maintenance of DNA methylation.
Postreplicative maintenance of genomic methylation patterns was proposed to depend largely on the binding of DNA methyltransferase 1 (Dnmt1) to PCNA, a core component of the replication machinery. We investigated how the slow and discontinuous DNA methylation could be mechanistically linked with fast and processive DNA replication. Using photobleaching and quantitative live cell imaging we show that Dnmt1 binding to PCNA is highly dynamic. Activity measurements of a PCNA-binding-deficient mutant with an enzyme-trapping assay in living cells showed that this interaction accounts for a 2-fold increase in methylation efficiency. Expression of this mutant in mouse dnmt1-/- embryonic stem (ES) cells restored CpG island methylation. Thus association of Dnmt1 with the replication machinery enhances methylation efficiency, but is not strictly required for maintaining global methylation. The transient nature of this interaction accommodates the different kinetics of DNA replication and methylation while contributing to faithful propagation of epigenetic information.
Schermelleh
Lothar
L
Ludwig Maximilians University Munich, Department of Biology II, 82152 Martinsried, Germany.
Haemmer
Andrea
A
Spada
Fabio
F
Rösing
Nicole
N
Meilinger
Daniela
D
Rothbauer
Ulrich
U
Cardoso
M Cristina
MC
Leonhardt
Heinrich
H
D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D002460:Cell Line; D018899:CpG Islands; D000076123:DNA (Cytosine-5-)-Methyltransferase 1; D004248:DNA (Cytosine-5-)-Methyltransferases / Q000737:chemistry / Q000378:metabolism / Q000502:physiology*; D019175:DNA Methylation; D004261:DNA Replication; D053595:Embryonic Stem Cells / Q000378:metabolism; D036681:Fluorescence Recovery After Photobleaching; D006801:Humans; D051379:Mice; D018345:Mice, Knockout; D008969:Molecular Sequence Data; D017354:Point Mutation; D018809:Proliferating Cell Nuclear Antigen / Q000378:metabolism; D017434:Protein Structure, Tertiary; D016415:Sequence Alignment
D018809:Proliferating Cell Nuclear Antigen; D000076123:DNA (Cytosine-5-)-Methyltransferase 1; D004248:DNA (Cytosine-5-)-Methyltransferases; C000619380:DNMT1 protein, human; C000619381:Dnmt1 protein, mouse