The genetic imprinting of individual loci or whole chromosomes, as in imprinted X-chromosome inactivation in mammals 1,2 , is established and reset during gametogenesis; defects in this process in the parent can result in disease in the offspring 3 . We describe a sperm-specific chromatin-based imprinting of the X chromosome in the nematode Caenorhabditis elegans that is restricted to histone H3 modifications. The epigenetic imprint is established during spermatogenesis and its stability in the offspring is affected by the presence of a pairing partner during meiosis in the parental germ line. We observed that DNA lacking a pairing partner during meiosis, the normal situation for the X chromosome in males, is targeted for methylation of histone H3 at Lys9 (H3-Lys9) and can be silenced. Targeting unpaired DNA for silencing during meiosis, a potential hallmark of genome defense, could therefore have a conserved role in imprinted X-chromosome inactivation and, ultimately, in sex chromosome evolution.The mechanisms by which imprinting is established during gametogenesis are not understood, but all such epigenetic phenomena clearly involve DNA methylation or histone modifications 4-7 . Methylation of H3-Lys9 is a highly conserved modification that is enriched in facultative and constitutive heterochromatin 8 . Methylation of histone H3 Lys4 (H3-Lys4), an opposing mark to methylation of H3-Lys9, is even more highly conserved and is enriched in transcriptionally competent or active DNA in all eukaryotes that have been examined 8 . Cytosine methylation, which is not found in all organisms, requires histone methylation in some species 9 , suggesting that histone modification may be the underlying mechanism guiding epigenetic processes. Indeed, methylation of histone H3 may provide an epigenetic mark during gametogenic imprinting of individual loci 10,11 . Histone modifications established during gametogenesis might therefore create a heritable epigenetic code that controls imprinted transcriptional competency in the offspring.The X chromosome in early adult germ cells of C. elegans lacks histone modifications associated with transcriptional competence 12 . The X chromosome is silenced in immature gametes of both sexes but shows sex-specific regulation during later gametic stages. In XX hermaphrodites, histones on the X chromosome become highly modified during early oogenesis, coincident with a burst of X-linked oocyte-specific gene expression 12 . In contrast, the unpaired X chromosome in (XO) males never accumulates 'activating' modifications during spermatogenesis but becomes enriched in methylated H3-Lys9 and condenses prematurely in pachytene nuclei 13 . This is markedly similar to what is observed for the XY body in mammalian spermatogenesis 13 . The X chromosome is also depleted of genes encoding sperm-enriched transcripts in C. elegans 12,14 . XX crossprogeny (produced from fertilization of a hermaphrodite by a male) therefore inherit sex chromosomes from each parent that differ substantially both in the spectrum ...

In C. elegans, mRNA production is initially repressed in the embryonic germline by a protein unique to C. elegans germ cells, PIE-1. PIE-1 is degraded upon the birth of the germ cell precursors, Z2 and Z3. We have identified a chromatin-based mechanism that succeeds PIE-1 repression in these cells. A subset of nucleosomal histone modifications, methylated lysine 4 on histone H3 (H3meK4) and acetylated lysine 8 on histone H4 (H4acetylK8), are globally lost and the DNA appears more condensed. This coincides with PIE-1 degradation and requires that germline identity is not disrupted. Drosophila pole cell chromatin also lacks H3meK4, indicating that a unique chromatin architecture is a conserved feature of embryonic germ cells. Regulation of the germline-specific chromatin architecture requires functional nanos activity in both organisms. These results indicate that genome-wide repression via a nanos-regulated, germ cell-specific chromatin organization is a conserved feature of germline maintenance during embryogenesis.

The DNA in eukaryotes is wrapped around a histone octamer core, together comprising the main subunit of chromatin, the nucleosome. Modifications of the nucleosomal histones in the genome correlate with the ability or inability of chromatin to form higher order structures, that in turn influence gene activity. The genome in primordial germ cells in early C. elegans germ cells carries a unique pattern of histone modifications that correlate with transcriptional repression in these cells, and aspects of this chromatin regulation are conserved in Drosophila. Loss of repression causes sterility in the adults, suggesting that chromatin-based repression is essential for germ line maintenance. The post-embryonic germ line also exhibits unique and dynamic aspects of chromatin regulation, with chromosome-wide regulation particularly evident on the X chromosome. Several properties of X-specific chromatin assembly are also sex-specific. These properties appear to be responding to the meiotic pairing status of the X chromosome, rather than the sex of the germ cells. Finally, gamete-specific chromatin regulation during gametogenesis impacts on X chromatin assembly in the offspring, leading to an apparent sperm-imprinted X inactivation in the early embryo. Other potential roles for germline-specific modes of chromatin assembly in genome regulation and protection are discussed.

Germline maintenance in the nematode C. elegans requires global repressive mechanisms that involve chromatin organization. During meiosis, the X chromosome in both sexes exhibits a striking reduction of histone modifications that correlate with transcriptional activation when compared with the genome as a whole. The histone modification spectrum on the X chromosome corresponds with a lack of transcriptional competence, as measured by reporter transgene arrays. The X chromosome in XO males is structurally analogous to the sex body in mammals, contains a histone modification associated with heterochromatin in other species and is inactivated throughout meiosis. The synapsed X chromosomes in hermaphrodites also appear to be silenced in early meiosis, but genes on the X chromosome are detectably expressed at later stages of oocyte meiosis. Silencing of the sex chromosome during early meiosis is a conserved feature throughout the nematode phylum, and is not limited to hermaphroditic species.