DNA methylation and normal chromosome behavior in neurospora depend on five components of a histone methyltransferase complex,DCDC

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. © 2010 lewis et al.