title: The stochastic silencing phenotype of Arabidopsis morc6 mutants reveals a role in efficient RNA-directed DNA methylation.
abstract: The RNA-directed DNA methylation (RdDM) pathway is of central importance to the initiation and maintenance of transcriptional gene silencing in plants. DNA methylation is directed to target sequences by a mechanism that involves production of small RNAs by RNA polymerase IV and long non-coding RNAs by RNA polymerase V. DNA methylation then leads to recruitment of histone-modifying enzymes, followed by establishment of a silenced chromatin state. Recently MORC6, a member of the microrchidia (MORC) family of adenosine triphosphatases (ATPases), has been shown to be involved in transcriptional gene silencing. However, reports differ regarding whether MORC6 is involved in RdDM itself or acts downstream of DNA methylation to enable formation of higher-order chromatin structure. Here we demonstrate that MORC6 is required for efficient RdDM at some target loci, and, using a GFP reporter system, we found that morc6 mutants show a stochastic silencing phenotype. By using cell sorting to separate silenced and unsilenced cells, we show that release of silencing at this locus is associated with a loss of DNA methylation. Thus our data support a view that MORC6 influences RdDM and that it is not acting downstream of DNA methylation. For some loci, efficient initiation or maintenance of DNA methylation may depend on the ability to form higher-order chromatin structure.
authors.affiliation: Department of Biology, University of York, YO10 5DD, York, UK.
authors.forename: Thomas R
authors.initials: TR
authors.lastname: Brabbs
authors.forename: Zhesi
authors.initials: Z
authors.lastname: He
authors.forename: Karen
authors.initials: K
authors.lastname: Hogg
authors.forename: Andrei
authors.initials: A
authors.lastname: Kamenski
authors.forename: Yi
authors.initials: Y
authors.lastname: Li
authors.forename: Konrad H
authors.initials: KH
authors.lastname: Paszkiewicz
authors.forename: Karen A
authors.initials: KA
authors.lastname: Moore
authors.forename: Peter
authors.initials: P
authors.lastname: O'Toole
authors.forename: Ian A
authors.initials: IA
authors.lastname: Graham
authors.forename: Louise
authors.initials: L
authors.lastname: Jones
mesh_terms: D000251:Adenosine Triphosphatases / Q000235:genetics / Q000378:metabolism / Q000502:physiology*; D017360:Arabidopsis / Q000235:genetics* / Q000378:metabolism; D029681:Arabidopsis Proteins / Q000235:genetics / Q000378:metabolism / Q000502:physiology*; D032461:Chromosomes, Plant / Q000378:metabolism; D019175:DNA Methylation; D018506:Gene Expression Regulation, Plant; D020868:Gene Silencing; D009154:Mutation; D010641:Phenotype; D011401:Promoter Regions, Genetic; D017422:Sequence Analysis, DNA; D013269:Stochastic Processes
chemical_list: D029681:Arabidopsis Proteins; D000251:Adenosine Triphosphatases; C573243:MORC6 protein, Arabidopsis
keywords: Arabidopsis thaliana; GFP; MORC; RdDM; chromatin; gene silencing