title: DNA methylation and normal chromosome behavior in Neurospora depend on five components of a histone methyltransferase complex, DCDC.
abstract: Methylation of DNA and of Lysine 9 on histone H3 (H3K9) is associated with gene silencing in many animals, plants, and fungi. In Neurospora crassa, methylation of H3K9 by DIM-5 directs cytosine methylation by recruiting a complex containing Heterochromatin Protein-1 (HP1) and the DIM-2 DNA methyltransferase. We report genetic, proteomic, and biochemical investigations into how DIM-5 is controlled. These studies revealed DCDC, a previously unknown protein complex including DIM-5, DIM-7, DIM-9, CUL4, and DDB1. Components of DCDC are required for H3K9me3, proper chromosome segregation, and DNA methylation. DCDC-defective strains, but not HP1-defective strains, are hypersensitive to MMS, revealing an HP1-independent function of H3K9 methylation. In addition to DDB1, DIM-7, and the WD40 domain protein DIM-9, other presumptive DCAFs (DDB1/CUL4 associated factors) co-purified with CUL4, suggesting that CUL4/DDB1 forms multiple complexes with distinct functions. This conclusion was supported by results of drug sensitivity tests. CUL4, DDB1, and DIM-9 are not required for localization of DIM-5 to incipient heterochromatin domains, indicating that recruitment of DIM-5 to chromatin is not sufficient to direct H3K9me3. DIM-7 is required for DIM-5 localization and mediates interaction of DIM-5 with DDB1/CUL4 through DIM-9. These data support a two-step mechanism for H3K9 methylation in Neurospora.
authors.affiliation: Institute of Molecular Biology, University of Oregon, Eugene, Oregon, United States of America.
authors.forename: Zachary A
authors.initials: ZA
authors.lastname: Lewis
authors.forename: Keyur K
authors.initials: KK
authors.lastname: Adhvaryu
authors.forename: Shinji
authors.initials: S
authors.lastname: Honda
authors.forename: Anthony L
authors.initials: AL
authors.lastname: Shiver
authors.forename: Marijn
authors.initials: M
authors.lastname: Knip
authors.forename: Ragna
authors.initials: R
authors.lastname: Sack
authors.forename: Eric U
authors.initials: EU
authors.lastname: Selker
mesh_terms: D020090:Chromosome Segregation / Q000235:genetics; D015825:Chromosomes, Fungal / Q000378:metabolism*; D019175:DNA Methylation; D005656:Fungal Proteins / Q000235:genetics / Q000378:metabolism; D020043:Genes, Essential / Q000235:genetics; D005800:Genes, Fungal / Q000235:genetics; D006570:Heterochromatin / Q000378:metabolism; D000076983:Histone Methyltransferases; D011495:Histone-Lysine N-Methyltransferase / Q000378:metabolism*; D006657:Histones / Q000378:metabolism; D008239:Lysine / Q000378:metabolism; D008954:Models, Biological; D009097:Multienzyme Complexes / Q000378:metabolism*; D009492:Neurospora crassa / Q000201:enzymology* / Q000235:genetics*; D011485:Protein Binding
chemical_list: D005656:Fungal Proteins; D006570:Heterochromatin; D006657:Histones; D009097:Multienzyme Complexes; D000076983:Histone Methyltransferases; D011495:Histone-Lysine N-Methyltransferase; D008239:Lysine