During meiotic prophase in male mammals, the heterologous X and Y chromosomes remain largely unsynapsed, and meiotic sex chromosome inactivation (MSCI) leads to formation of the transcriptionally silenced XY body. In birds, the heterogametic sex is female, carrying Z and W chromosomes (ZW), whereas males have the homogametic ZZ constitution. During chicken oogenesis, the heterologous ZW pair reaches a state of complete heterologous synapsis, and this might enable maintenance of transcription of Z- and W chromosomal genes during meiotic prophase. Herein, we show that the ZW pair is transiently silenced, from early pachytene to early diplotene using immunocytochemistry and gene expression analyses. We propose that ZW inactivation is most likely achieved via spreading of heterochromatin from the W on the Z chromosome. Also, persistent meiotic DNA double-strand breaks (DSBs) may contribute to silencing of Z. Surprisingly, γH2AX, a marker of DSBs, and also the earliest histone modification that is associated with XY body formation in mammalian and marsupial spermatocytes, does not cover the ZW during the synapsed stage. However, when the ZW pair starts to desynapse, a second wave of γH2AX accumulates on the unsynapsed regions of Z, which also show a reappearance of the DSB repair protein RAD51. This indicates that repair of meiotic DSBs on the heterologous part of Z is postponed until late pachytene/diplotene, possibly to avoid recombination with regions on the heterologously synapsed W chromosome. Two days after entering diplotene, the Z looses γH2AX and shows reactivation. This is the first report of meiotic sex chromosome inactivation in a species with female heterogamety, providing evidence that this mechanism is not specific to spermatogenesis. It also indicates the presence of an evolutionary force that drives meiotic sex chromosome inactivation independent of the final achievement of synapsis.
References
[1]
Zickler D (2006) From early homologue recognition to synaptonemal complex formation. Chromosoma 115: 158–174.
[2]
Monesi V (1965) Differential rate of ribonucleic acid synthesis in the autosomes and sex chromosomes during male meiosis in the mouse. Chromosoma 17: 11–21.
[3]
Turner JM, Mahadevaiah SK, Ellis PJ, Mitchell MJ, Burgoyne PS (2006) Pachytene asynapsis drives meiotic sex chromosome inactivation and leads to substantial postmeiotic repression in spermatids. Dev Cell 10: 521–529.
[4]
Solovei I, Gaginskaya E, Hutchison N, Macgregor H (1993) Avian sex chromosomes in the lampbrush form: the ZW lampbrush bivalents from six species of bird. Chromosome Res 1: 153–166.
[5]
Solari AJ (1992) Equalization of Z and W axes in chicken and quail oocytes. Cytogenet Cell Genet 59: 52–56.
[6]
Moses MJ, Poorman PA (1981) Synaptosomal complex analysis of mouse chromosomal rearrangements. II. Synaptic adjustment in a tandem duplication. Chromosoma 81: 519–535.
[7]
Moses MJ, Poorman PA, Roderick TH, Davisson MT (1982) Synaptonemal complex analysis of mouse chromosomal rearrangements. IV. Synapsis and synaptic adjustment in two paracentric inversions. Chromosoma 84: 457–474.
[8]
Stefos K, Arrighi FE (1971) Heterochromatic nature of W chromosome in birds. Exp Cell Res 68: 228–231.
[9]
Becak ML, Becak W (1981) Behaviour of the ZW sex bivalent in the snake Bothrops jararaca. Chromosoma 83: 289–293.
[10]
Ohno S (1961) Sex chromosomes and microchromosomes of Gallus domesticus. Chromosoma 11: 484–498.
[11]
Solari AJ (1977) Ultrastructure of the synaptic autosomes and the ZW bivalent in chicken oocytes. Chromosoma 64: 155–165.
[12]
Jablonka E, Lamb MJ (1988) Meiotic pairing constraints and the activity of sex chromosomes. J Theor Biol 133: 23–36.
[13]
Baarends WM, Wassenaar E, van der Laan R, Hoogerbrugge J, Sleddens-Linkels E, et al. (2005) Silencing of unpaired chromatin and histone H2A ubiquitination in mammalian meiosis. Mol Cell Biol 25: 1041–1053.
[14]
Schimenti J (2005) Synapsis or silence. Nat Genet 37: 11–13.
[15]
Turner JM, Mahadevaiah SK, Fernandez-Capetillo O, Nussenzweig A, Xu X, et al. (2005) Silencing of unsynapsed meiotic chromosomes in the mouse. Nat Genet 37: 41–47.
[16]
Kelly WG, Aramayo R (2007) Meiotic silencing and the epigenetics of sex. Chromosome Res 15: 633–651.
[17]
Mahadevaiah SK, Turner JM, Baudat F, Rogakou EP, de Boer P, et al. (2001) Recombinational DNA double-strand breaks in mice precede synapsis. Nat Genet 27: 271–276.
[18]
Padmore R, Cao L, Kleckner N (1991) Temporal comparison of recombination and synaptonemal complex formation during meiosis in S. cerevisiae. Cell 66: 1239–1256.
[19]
Moens PB, Chen DJ, Shen Z, Kolas N, Tarsounas M, et al. (1997) Rad51 immunocytology in rat and mouse spermatocytes and oocytes. Chromosoma 106: 207–215.
[20]
Ashley T, Plug AW, Xu J, Solari AJ, Reddy G, et al. (1995) Dynamic changes in Rad51 distribution on chromatin during meiosis in male and female vertebrates. Chromosoma 104: 19–28.
[21]
Moens PB, Kolas NK, Tarsounas M, Marcon E, Cohen PE, et al. (2002) The time course and chromosomal localization of recombination-related proteins at meiosis in the mouse are compatible with models that can resolve the early DNA-DNA interactions without reciprocal recombination. J Cell Sci 115: 1611–1622.
[22]
Schoenmakers S, Wassenaar E, van Cappellen WA, Derijck AA, de Boer P, et al. (2008) Increased frequency of asynapsis and associated meiotic silencing of heterologous chromatin in the presence of irradiation-induced extra DNA double strand breaks. Dev Biol 317: 270–281.
[23]
Peters AH, Plug AW, van Vugt MJ, de Boer P (1997) A drying-down technique for the spreading of mammalian meiocytes from the male and female germline. Chromosome Res 5: 66–68.
[24]
Essers J, Hendriks RW, Wesoly J, Beerens CE, Smit B, et al. (2002) Analysis of mouse Rad54 expression and its implications for homologous recombination. DNA Repair (Amst) 1: 779–793.
[25]
Costa Y, Cooke HJ (2007) Dissecting the mammalian synaptonemal complex using targeted mutations. Chromosome Res 15: 579–589.
[26]
Nakayama J, Rice JC, Strahl BD, Allis CD, Grewal SI (2001) Role of histone H3 lysine 9 methylation in epigenetic control of heterochromatin assembly. Science 292: 110–113.
[27]
Hughes GC (1963) The Population of Germ Cells in the Developing Female Chick. J Embryol Exp Morphol 11: 513–536.
Turner JM (2007) Meiotic sex chromosome inactivation. Development 134: 1823–1831.
[30]
Fernandez-Capetillo O, Mahadevaiah SK, Celeste A, Romanienko PJ, Camerini-Otero RD, et al. (2003) H2AX is required for chromatin remodeling and inactivation of sex chromosomes in male mouse meiosis. Dev Cell 4: 497–508.
[31]
de Napoles M, Mermoud JE, Wakao R, Tang YA, Endoh M, et al. (2004) Polycomb group proteins Ring1A/B link ubiquitylation of histone H2A to heritable gene silencing and X inactivation. Dev Cell 7: 663–676.
[32]
Fang J, Chen T, Chadwick B, Li E, Zhang Y (2004) Ring1b-mediated H2A ubiquitination associates with inactive X chromosomes and is involved in initiation of X inactivation. J Biol Chem 279: 52812–52815.
[33]
Baarends WM, Hoogerbrugge JW, Roest HP, Ooms M, Vreeburg J, et al. (1999) Histone ubiquitination and chromatin remodeling in mouse spermatogenesis. Dev Biol 207: 322–333.
[34]
Plath K, Fang J, Mlynarczyk-Evans SK, Cao R, Worringer KA, et al. (2003) Role of histone H3 lysine 27 methylation in X inactivation. Science 300: 131–135.
[35]
Namekawa SH, Park PJ, Zhang LF, Shima JE, McCarrey JR, et al. (2006) Postmeiotic sex chromatin in the male germline of mice. Curr Biol 16: 660–667.
[36]
Namekawa SH, VandeBerg JL, McCarrey JR, Lee JT (2007) Sex chromosome silencing in the marsupial male germ line. Proc Natl Acad Sci U S A 104: 9730–9735.
[37]
Bisoni L, Batlle-Morera L, Bird AP, Suzuki M, McQueen HA (2005) Female-specific hyperacetylation of histone H4 in the chicken Z chromosome. Chromosome Res 13: 205–214.
[38]
Solari AJ (1977) Ultrastructure of the synaptic autosomes and the ZW bivalent in chicken oocytes. Chromosoma 64: 155–165.
[39]
Franco MJ, Sciurano RB, Solari AJ (2007) Protein immunolocalization supports the presence of identical mechanisms of XY body formation in eutherians and marsupials. Chromosome Res 15: 815–824.
[40]
Mizuno S, Kunita R, Nakabayashi O, Kuroda Y, Arai N, et al. (2002) Z and W chromosomes of chickens: studies on their gene functions in sex determination and sex differentiation. Cytogenet Genome Res 99: 236–244.
[41]
O'Neill M, Binder M, Smith C, Andrews J, Reed K, et al. (2000) ASW: a gene with conserved avian W-linkage and female specific expression in chick embryonic gonad. Dev Genes Evol 210: 243–249.
[42]
Smith CA (2007) Sex determination in birds: HINTs from the W sex chromosome? Sex Dev 1: 279–285.
[43]
Hori T, Asakawa S, Itoh Y, Shimizu N, Mizuno S (2000) Wpkci, encoding an altered form of PKCI, is conserved widely on the avian W chromosome and expressed in early female embryos: implication of its role in female sex determination. Mol Biol Cell 11: 3645–3660.
[44]
Mueller JL, Mahadevaiah SK, Park PJ, Warburton PE, Page DC, et al. (2008) The mouse X chromosome is enriched for multicopy testis genes showing postmeiotic expression. Nat Genet 40: 794–799.
[45]
Tres LL (1977) Extensive pairing of the XY bivalent in mouse spermatocytes as visualized by whole-mount electron microscopy. J Cell Sci 25: 1–15.
[46]
Wang PJ (2004) X chromosomes, retrogenes and their role in male reproduction. Trends Endocrinol Metab 15: 79–83.
[47]
Consortium ICGS (2004) Sequence and comparative analysis of the chicken genome provide unique perspectives on vertebrate evolution. Nature 432: 695–716.
[48]
Ellegren H, Hultin-Rosenberg L, Brunstrom B, Dencker L, Kultima K, et al. (2007) Faced with inequality: chicken do not have a general dosage compensation of sex-linked genes. BMC Biol 5: 40.
[49]
Khil PP, Oliver B, Camerini-Otero RD (2005) X for intersection: retrotransposition both on and off the X chromosome is more frequent. Trends Genet 21: 3–7.
[50]
Khil PP, Smirnova NA, Romanienko PJ, Camerini-Otero RD (2004) The mouse X chromosome is enriched for sex-biased genes not subject to selection by meiotic sex chromosome inactivation. Nat Genet 36: 642–646.
[51]
Mank JE, Ellegren H (2009) Sex-Linkage of Sexually Antagonistic Genes Is Predicted by Female, but Not Male, Effects in Birds. Evolution. doi: 10.1111/j.1558-5646.2009.00618.x.
[52]
Hamer G, Roepers-Gajadien HL, van Duyn-Goedhart A, Gademan IS, Kal HB, et al. (2003) DNA double-strand breaks and gamma-H2AX signaling in the testis. Biol Reprod 68: 628–634.
[53]
Mailand N, Bekker-Jensen S, Faustrup H, Melander F, Bartek J, et al. (2007) RNF8 ubiquitylates histones at DNA double-strand breaks and promotes assembly of repair proteins. Cell 131: 887–900.
[54]
O'Hagan HM, Mohammad HP, Baylin SB (2008) Double strand breaks can initiate gene silencing and SIRT1-dependent onset of DNA methylation in an exogenous promoter CpG island. PLoS Genet 4: e1000155.
[55]
Itoh Y, Melamed E, Yang X, Kampf K, Wang S, et al. (2007) Dosage compensation is less effective in birds than in mammals. J Biol 6: 2.
[56]
Melamed E, Arnold AP (2007) Regional differences in dosage compensation on the chicken Z chromosome. Genome Biol 8: R202.
[57]
Bowring FJ, Yeadon PJ, Stainer RG, Catcheside DE (2006) Chromosome pairing and meiotic recombination in Neurospora crassa spo11 mutants. Curr Genet 50: 115–123.
[58]
Cabrero J, Teruel M, Carmona FD, Jimenez R, Camacho JP (2007) Histone H3 lysine 9 acetylation pattern suggests that X and B chromosomes are silenced during entire male meiosis in a grasshopper. Cytogenet Genome Res 119: 135–142.
[59]
Bininda-Emonds OR, Cardillo M, Jones KE, MacPhee RD, Beck RM, et al. (2007) The delayed rise of present-day mammals. Nature 446: 507–512.
[60]
Kumar S, Hedges SB (1998) A molecular timescale for vertebrate evolution. Nature 392: 917–920.
[61]
Pereira SL, Baker AJ (2006) A mitogenomic timescale for birds detects variable phylogenetic rates of molecular evolution and refutes the standard molecular clock. Mol Biol Evol 23: 1731–1740.
[62]
Warren WC, Hillier LW, Marshall Graves JA, Birney E, Ponting CP, et al. (2008) Genome analysis of the platypus reveals unique signatures of evolution. Nature 453: 175–183.
[63]
Woodburne MO, Rich TH, Springer MS (2003) The evolution of tribospheny and the antiquity of mammalian clades. Mol Phylogenet Evol 28: 360–385.
[64]
van der Heijden GW, Derijck AA, Posfai E, Giele M, Pelczar P, et al. (2007) Chromosome-wide nucleosome replacement and H3.3 incorporation during mammalian meiotic sex chromosome inactivation. Nat Genet 39: 251–258.
