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PKC isoforms interact with and phosphorylate DNMT1
Geneviève Lavoie1, Pierre-Olivier Estève, Nathalie Bibens Laulan
1Institut national de la recherche scientifique, INRS-Institut Armand-Frappier, Laval, QC, Canada.
BMC Biology
|May 31, 2011
Summary
Protein kinase C (PKC) isoforms specifically phosphorylate DNA methyltransferase 1 (DNMT1), particularly PKCζ. This interaction regulates genome-wide DNA methylation patterns and DNMT1 activity.
Area of Science:
- Epigenetics
- Molecular Biology
- Cell Signaling
Background:
- DNA methyltransferase 1 (DNMT1) is subject to phosphorylation, but the specific roles of protein kinase C (PKC) isoforms are unclear.
- The PKC family comprises over 12 isoforms with diverse characteristics.
Purpose of the Study:
- To investigate the specific PKC isoforms that phosphorylate DNMT1.
- To elucidate the functional consequences of DNMT1 phosphorylation by PKC isoforms, focusing on PKCζ.
Main Methods:
- In vitro phosphorylation assays using recombinant DNMT1 fragments.
- GST pull-down and co-immunoprecipitation to confirm protein interactions.
- Fluorescent microscopy for co-localization studies.
- Gene methylation analysis in HEK-293 cells overexpressing PKCζ and DNMT1.
Main Results:
- PKC isoforms α, βI, βII, δ, γ, η, ζ, and μ phosphorylated the N-terminal domain of DNMT1; PKCε did not.
- PKCζ directly interacted with and phosphorylated DNMT1, reducing its methyltransferase activity.
- Co-expression of PKCζ and DNMT1 decreased genome-wide DNA methylation levels.
Conclusions:
- DNMT1 phosphorylation by PKC is isoform-specific.
- PKCζ cooperates with DNMT1 to regulate DNA methylation patterns across the genome.
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