Inheritance of an epigenetic mark: the CpG DNA methyltransferase 1 is required for de novo establishment of a complex pattern of non-CpG methylation.

Site-specific methylation of cytosines is a key epigenetic mark of vertebrate DNA. While a majority of the methylated residues are in the symmetrical (meC)pG:Gp(meC) configuration, a smaller, but significant fraction is found in the CpA, CpT and CpC asymmetric (non-CpG) dinucleotides. CpG methylation is reproducibly maintained by the activity of the DNA methyltransferase 1 (Dnmt1) on the newly replicated hemimethylated substrates (meC)pG:GpC. On the other hand, establishment and hereditary maintenance of non-CpG methylation patterns have not been analyzed in detail. We previously reported the occurrence of site- and allele-specific methylation at both CpG and non-CpG sites. Here we characterize a hereditary complex of non-CpG methylation, with the transgenerational maintenance of three distinct profiles in a constant ratio, associated with extensive CpG methylation. These observations raised the question of the signal leading to the maintenance of the pattern of asymmetric methylation. The complete non-CpG pattern was reinstated at each generation in spite of the fact that the majority of the sperm genomes contained either none or only one methylated non-CpG site. This observation led us to the hypothesis that the stable CpG patterns might act as blueprints for the maintenance of non-CpG DNA methylation. As predicted, non-CpG DNA methylation profiles were abrogated in a mutant lacking Dnmt1, the enzymes responsible for CpG methylation, but not in mutants defective for either Dnmt3a or Dnmt2.

Timing of entry of meiosis depends on a mark generated by DNA methyltransferase 3a in testis.

Reprogramming of DNA methylation is an essential part of gametogenesis, and a role of two members of the DNA methyltransferase (Dnmt) family, Dnmt3a and Dnmt3L, has been recognized. In an attempt to elucidate the role of Dnmt3a, we analyzed the progression of spermatogenesis in Dnmt3a (-/-) homozygotes during the first 3 weeks of post-natal development. The emerging picture was markedly different from that recently reported for the Dnmt3L protein. In the Dnmt3a (-/-) testis, at the expected time of entry into meiosis (11-13 dpp), the number of spermatocytes was greatly reduced. They progressively accumulated during the following days, but at a slower rate than in the wild type. Once started, however, the pachytene stage was apparently completed with normal chromosome pairing and formation of the sex vesicle, and spermatogenesis further progressed with the appearance and the expression of round spermatid specific markers. Interestingly and unlike Dnmt3L (-/-) spermatocytes, Dnmt3a (-/-) germ cells showed only a minor reduction in the methylation of interspersed repetitive elements and retroposons. The Dnmt3a might thus generate a mark important for the initiation of male meiosis that is distinct from that created by Dnmt3L.

A novel germ line-specific gene of the phosducin-like protein (PhLP) family. A meiotic function conserved from yeast to mice.

We identified a new member of the phosducin-like (PhLP) protein family that is predominantly, if not exclusively, expressed in male and female germ cells. In situ analysis on testis sections and analysis of purified spermatogenic cell fractions evidenced a stage-specific expression with high levels of RNA and protein in pachytene spermatocytes and round spermatids. Three mRNA species were detected, which correspond to different polyadenylation sites and vary in abundance during germ cell maturation. Only low levels of RNA were detected in whole ovary extracts, but expression of the protein became detectable within hours after hormonal induction of superovulation. The gene (Mgcphlp) is located on mouse chromosome 5 in the immediate vicinity of the Clock locus. The predicted amino acid sequence shows extensive similarities not only with the known mammalian PhLP proteins but also with the yeast phosducin-like protein Plp2, required for the production and growth of haploid cells. Expression of the murine protein was found to complement the defect of a yeast plp2 Delta mutant. We propose that MgcPhLP/Plp2 proteins exert a function in germ cell maturation that is conserved from yeast to mammals.