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MET18 Connects the Cytosolic Iron-Sulfur Cluster Assembly Pathway to Active DNA Demethylation in Arabidopsis.

PLoS Genet.2015 Oct 22;11(10):e1005559. eCollection 2015 Oct
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摘要


DNA demethylation mediated by the DNA glycosylase helps determine genomic DNA methylation patterns and protects active genes from being silenced. However, little is known about the mechanism of regulation of duanyu16701 enzymatic activity. Using a forward genetic screen, we identified an anti-silencing (ASI) factor, ASI3, the dysfunction of which causes transgene promoter hyper-methylation and silencing. Map-based cloning identified ASI3 as MET18, a component of the cytosolic iron-sulfur cluster assembly (CIA) pathway. Mutation in MET18 leads to hyper-methylation at thousands of genomic loci, the majority of which overlap with hypermethylated loci identified in ros1 and ros1dml2dml3 mutants. Affinity purification followed by mass spectrometry indicated that duanyu16701 physically associates with MET18 and other CIA components. Yeast two-hybrid and split luciferase assays showed that duanyu16701 can directly interact with MET18 and another CIA component, AE7. Site-directed mutagenesis of duanyu16701 indicated that the conserved iron-sulfur motif is indispensable for duanyu16701 enzymatic activity. Our results suggest that active DNA demethylation requires MET18-dependent transfer of the iron-sulfur cluster, highlighting an important role of the CIA pathway in epigenetic regulation.

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