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The journal and the authors apologise for an error in the above titled article published in this journal (vol 144, pp 433–445). The authors inadvertently presented duplicate sperm images for XY and XESxrbO mouse testes of Fig. 6 (bottom panels). This error does not change the findings of the paper, as this figure does not give a quantitative breakdown of the proportions of different shapes., (© 2020 Society for Reproduction and Fertility)

Following publication of the original article [1], the following error was reported: The actin control panel in Fig. 3 of this paper is reproduced from Fig. 7 of Touré et al, 2004 [2] by kind permission of the Genetics Society of America. Touré et al, 2004 used Northern blotting to show that the Y-linked genes Ssty1 and Ssty2 have reduced expression in a range of mouse genotypes with deletions on the Y chromosome long arm. This paper shows that two novel genes, Sly and Asty are also present on mouse Yq and have reduced expression in these deleted genotypes. A further companion paper was published in Human Molecular Genetics (Ellis et al, 2005 [3]) showing that X-linked genes are upregulated in the various deleted genotypes. Since two of the genotypes concerned are sterile and very hard to generate, all the Northern blot experiments in these papers were performed on a single membrane that was stripped and re-probed with a range of different X- and Y-linked genes. The same beta-actin loading control image thus necessarily applies to all the data presented, and was shown in all three papers. We regret that this was not mentioned appropriately in the Methods and figure legends at the time of publication.

During spermatogenesis, germ cells that fail to synapse their chromosomes or fail to undergo meiotic sex chromosome inactivation (MSCI) are eliminated via apoptosis during mid-pachytene. Previous work showed that Y-linked genes Zfy1 and Zfy2 act as 'executioners' for this checkpoint, and that wrongful expression of either gene during pachytene triggers germ cell death. Here, we show that in mice, Zfy genes are also necessary for efficient MSCI and the sex chromosomes are not correctly silenced in Zfy-deficient spermatocytes. This unexpectedly reveals a triple role for Zfy at the mid-pachytene checkpoint in which Zfy genes first promote MSCI, then monitor its progress (since if MSCI is achieved, Zfy genes will be silenced), and finally execute cells with MSCI failure. This potentially constitutes a negative feedback loop governing this critical checkpoint mechanism., (© The Author 2016. Published by Oxford University Press.)

In animals with heteromorphic sex chromosomes, all sex differences originate from the sex chromosomes, which are the only factors that are consistently different in male and female zygotes. In mammals, the imbalance in Y gene expression, specifically the presence vs. absence of Sry , initiates the differentiation of testes in males, setting up lifelong sex differences in the level of gonadal hormones, which in turn cause many sex differences in the phenotype of non-gonadal tissues. The inherent imbalance in the expression of X and Y genes, or in the epigenetic impact of X and Y chromosomes, also has the potential to contribute directly to the sexual differentiation of non-gonadal cells. Here, we review the research strategies to identify the X and Y genes or chromosomal regions that cause direct, sexually differentiating effects on non-gonadal cells. Some mouse models are useful for separating the effects of sex chromosomes from those of gonadal hormones. Once direct "sex chromosome effects" are detected in these models, further studies are required to narrow down the list of candidate X and/or Y genes and then to identify the sexually differentiating genes themselves. Logical approaches to the search for these genes are reviewed here.

In a male mouse, meiosis markers of processed DNA double strand breaks (DSBs) such as DMC1 and RAD51 are regularly seen in the non-PAR region of the X chromosome; these disappear late in prophase prior to entry into the first meiotic metaphase. Marker evidence for DSBs occurring in the non-PAR region of the Y chromosome is limited. Nevertheless, historically it has been documented that recombination can occur within the mouse Y short arm (Yp) when an additional Yp segment is attached distal to the X and/or the Y pseudoautosomal region (PAR). A number of recombinants identified among offsprings involved unequal exchanges involving repeated DNA segments; however, equal exchanges will have frequently been missed because of the paucity of markers to differentiate between the two Yp segments. Here, we discuss this historical data and present extensive additional data obtained for two mouse models with Yp additions to the X PAR. PCR genotyping enabled identification of a wider range of potential recombinants; the proportions of Yp exchanges identified among the recombinants were 9.7 and 22.4 %. The frequency of these exchanges suggests that the Yp segment attached to the X PAR is subject to the elevated level of recombinational DSBs that characterizes the PAR.

A previous study indicated that genetic information encoded on the mouse Y chromosome short arm (Yp) is required for efficient completion of the second meiotic division (that generates haploid round spermatids), restructuring of the sperm head, and development of the sperm tail. Using mouse models lacking a Y chromosome but with varying Yp gene complements provided by Yp chromosomal derivatives or transgenes, we recently identified the Y-encoded zinc finger transcription factors Zfy1 and Zfy2 as the Yp genes promoting the second meiotic division. Using the same mouse models we here show that Zfy2 (but not Zfy1) contributes to the restructuring of the sperm head and is required for the development of the sperm tail. The preferential involvement of Zfy2 is consistent with the presence of an additional strong spermatid-specific promotor that has been acquired by this gene. This is further supported by the fact that promotion of sperm morphogenesis is also seen in one of the two markedly Yp gene deficient models in which a Yp deletion has created a Zfy2/1 fusion gene that is driven by the strong Zfy2 spermatid-specific promotor, but encodes a protein almost identical to that encoded by Zfy1. Our results point to there being further genetic information on Yp that also has a role in restructuring the sperm head.

Mouse Zfy1 and Zfy2 encode zinc finger transcription factors that map to the short arm of the Y chromosome (Yp). They have previously been shown to promote meiotic quality control during pachytene (Zfy1 and Zfy2) and at the first meiotic metaphase (Zfy2). However, from these previous studies additional roles for genes encoded on Yp during meiotic progression were inferred. In order to identify these genes and investigate their function in later stages of meiosis, we created three models with diminishing Yp and Zfy gene complements (but lacking the Y-long-arm). Since the Y-long-arm mediates pairing and exchange with the X via their pseudoautosomal regions (PARs) we added a minute PAR-bearing X chromosome derivative to enable formation of a sex bivalent, thus avoiding Zfy2-mediated meiotic metaphase I (MI) checkpoint responses to the unpaired (univalent) X chromosome. Using these models we obtained definitive evidence that genetic information on Yp promotes meiosis II, and by transgene addition identified Zfy1 and Zfy2 as the genes responsible. Zfy2 was substantially more effective and proved to have a much more potent transactivation domain than Zfy1. We previously established that only Zfy2 is required for the robust apoptotic elimination of MI spermatocytes in response to a univalent X; the finding that both genes potentiate meiosis II led us to ask whether there was de novo Zfy1 and Zfy2 transcription in the interphase between meiosis I and meiosis II, and this proved to be the case. X-encoded Zfx was also expressed at this stage and Zfx over-expression also potentiated meiosis II. An interphase between the meiotic divisions is male-specific and we previously hypothesised that this allows meiosis II critical X and Y gene reactivation following sex chromosome silencing in meiotic prophase. The interphase transcription and meiosis II function of Zfx, Zfy1 and Zfy2 validate this hypothesis.

Outbred XY(Sry-) female mice that lack Sry due to the 11 kb deletion Sry(dl1Rlb) have very limited fertility. However, five lines of outbred XY(d) females with Y chromosome deletions Y(Del(Y)1Ct)-Y(Del(Y)5Ct) that deplete the Rbmy gene cluster and repress Sry transcription were found to be of good fertility. Here we tested our expectation that the difference in fertility between XO, XY(d-1) and XY(Sry-) females would be reflected in different degrees of oocyte depletion, but this was not the case. Transgenic addition of Yp genes to XO females implicated Zfy2 as being responsible for the deleterious Y chromosomal effect on fertility. Zfy2 transcript levels were reduced in ovaries of XY(d-1) compared with XY(Sry-) females in keeping with their differing fertility. In seeking the biological basis of the impaired fertility we found that XY(Sry-), XY(d-1) and XO,Zfy2 females produce equivalent numbers of 2-cell embryos. However, in XY(Sry-) and XO,Zfy2 females the majority of embryos arrested with 2-4 cells and almost no blastocysts were produced; by contrast, XY(d-1) females produced substantially more blastocysts but fewer than XO controls. As previously documented for C57BL/6 inbred XY females, outbred XY(Sry-) and XO,Zfy2 females showed frequent failure of the second meiotic division, although this did not prevent the first cleavage. Oocyte transcriptome analysis revealed major transcriptional changes resulting from the Zfy2 transgene addition. We conclude that Zfy2-induced transcriptional changes in oocytes are sufficient to explain the more severe fertility impairment of XY as compared with XO females.

We recently used three XO male mouse models with varying Y short-arm (Yp) gene complements, analysed at 30 days post partum, to demonstrate a Yp gene requirement for the apoptotic elimination of spermatocytes with a univalent X chromosome at the first meiotic metaphase. The three mouse models were i) XSxr(a)O in which the Yp-derived Tp(Y)1Ct(Sxr-a) sex reversal factor provides an almost complete Yp gene complement, ii) XSxr(b)O,Eif2s3y males in which Tp(Y)1Ct(Sxr-b) has a deletion completely or partially removing eight Yp genes - the Yp gene Eif2s3y has been added as a transgene to support spermatogonial proliferation, and iii) XOSry,Eif2s3y males in which the Sry transgene directs gonad development along the male pathway. In this study, we have used the same mouse models analysed at 6 weeks of age to investigate potential Yp gene involvement in spermiogenesis. We found that all three mouse models produce haploid and diploid spermatids and that the diploid spermatids showed frequent duplication of the developing acrosomal cap during the early stages. However, only in XSxr(a)O males did spermiogenesis continue to completion. Most strikingly, in XOSry,Eif2s3y males, spermatid development arrested at round spermatid step 7 so that no sperm head restructuring or tail development was observed. In contrast, in XSxr(b)O,Eif2s3y males, spermatids with substantial sperm head and tail morphogenesis could be easily found, although this was delayed compared with XSxr(a)O. We conclude that Sxr(a) (and therefore Yp) includes genetic information essential for sperm morphogenesis and that this is partially retained in Sxr(b).

Intragenomic conflicts arise when a genetic element favours its own transmission to the detriment of others. Conflicts over sex chromosome transmission are expected to have influenced genome structure, gene regulation, and speciation. In the mouse, the existence of an intragenomic conflict between X- and Y-linked multicopy genes has long been suggested but never demonstrated. The Y-encoded multicopy gene Sly has been shown to have a predominant role in the epigenetic repression of post meiotic sex chromatin (PMSC) and, as such, represses X and Y genes, among which are its X-linked homologs Slx and Slxl1. Here, we produced mice that are deficient for both Sly and Slx/Slxl1 and observed that Slx/Slxl1 has an opposite role to that of Sly, in that it stimulates XY gene expression in spermatids. Slx/Slxl1 deficiency rescues the sperm differentiation defects and near sterility caused by Sly deficiency and vice versa. Slx/Slxl1 deficiency also causes a sex ratio distortion towards the production of male offspring that is corrected by Sly deficiency. All in all, our data show that Slx/Slxl1 and Sly have antagonistic effects during sperm differentiation and are involved in a postmeiotic intragenomic conflict that causes segregation distortion and male sterility. This is undoubtedly what drove the massive gene amplification on the mouse X and Y chromosomes. It may also be at the basis of cases of F1 male hybrid sterility where the balance between Slx/Slxl1 and Sly copy number, and therefore expression, is disrupted. To the best of our knowledge, our work is the first demonstration of a competition occurring between X and Y related genes in mammals. It also provides a biological basis for the concept that intragenomic conflict is an important evolutionary force which impacts on gene expression, genome structure, and speciation., Competing Interests: The authors have declared that no competing interests exist.

Mammalian ZFY genes are located on the Y chromosome, and code putative transcription factors with 12-13 zinc fingers preceded by a large acidic (activating) domain. In mice, there are two genes, Zfy1 and Zfy2, which are expressed mainly in the testis. Their transcription increases in germ cells as they enter meiosis, both are silenced by meiotic sex chromosome inactivation (MSCI) during pachytene, and Zfy2 is strongly reactivated later in spermatids. Recently, we have shown that mouse Zfy2, but not Zfy1, is involved in triggering the apoptotic elimination of specific types of sex chromosomally aberrant spermatocytes. In humans, there is a single widely transcribed ZFY gene, and there is no evidence for a specific role in the testis. Here, we characterize ZFY transcription during spermatogenesis in mice and humans. In mice, we define a variety of Zfy transcripts, among which is a Zfy2 transcript that predominates in spermatids, and a Zfy1 transcript, lacking an exon encoding approximately half of the acidic domain, which predominates prior to MSCI. In humans, we have identified a major testis-specific ZFY transcript that encodes a protein with the same short acidic domain. This represents the first evidence that ZFY has a conserved function during human spermatogenesis. We further show that, in contrast to the full acidic domain, the short domain does not activate transcription in yeast, and we hypothesize that this explains the functional difference observed between Zfy1 and Zfy2 during mouse meiosis.

Attention Deficit Hyperactivity Disorder (ADHD) is a common neurodevelopmental condition characterised by inattention, impulsivity and hyperactivity; it is frequently co-morbid with anxiety and conduct disorders, sleep perturbation and abnormal consummatory behaviours. Recent studies have implicated the neurosteroid-modulating enzyme steroid sulfatase (STS) as a modulator of ADHD-related endophenotypes. The effects of steroid sulfatase deficiency on homecage activity, feeding/drinking behaviours, anxiety-related behaviours (assayed in light-dark box and open field paradigms), social dominance and serum steroid hormone levels were determined by comparing 40,XY and 39,X(Y*)O mice. Subsequently, mice administered the steroid sulfatase inhibitor COUMATE acutely were compared to vehicle-treated mice on behavioural tasks sensitive to enzyme deficiency to dissociate between its developmental and ongoing effects. 39,X(Y*)O mice exhibited heightened reactivity to a novel environment, hyperactivity in the active phase, and increased water (but not food) consumption relative to 40,XY mice during a 24h period; the former group also demonstrated evidence for heightened emotional reactivity. There was no difference in social dominance between the 40,XY and 39,X(Y*)O mice. COUMATE administration had no effect on homecage activity, water consumption or anxiety measures in the open field. 39,X(Y*)O mice exhibited significantly lower dehydroepiandrosterone (DHEA) serum levels than 40,XY mice, but equivalent corticosterone levels. Together with previous data, the present results suggest that steroid sulfatase may influence core and associated ADHD behavioural endophenotypes via both developmental and ongoing mechanisms, and that the 39,X(Y*)O model may represent a useful tool for elucidating the neurobiological basis of these endophenotypes., (Copyright © 2011 Elsevier Ltd. All rights reserved.)

During male but not female mammalian meiosis, there is efficient apoptotic elimination of cells with unpaired (univalent) chromosomes at the first meiotic metaphase (MI) [1]. Apoptotic elimination of MI spermatocytes is seen in response to the univalent X chromosome of XSxr(a)O male mice [2], in which the X chromosome carries Sxr(a) [3, 4], the Y-chromosome-derived sex-reversal factor that includes the testis determinant Sry. Sxr(b) is an Sxr(a)-derived variant in which a deletion has removed six Y short-arm genes and created a Zfy2/Zfy1 fusion gene spanning the deletion breakpoint [4, 5]. XSxr(b)O males have spermatogonial arrest that can be overcome by the re-addition of Eif2s3y from the deletion as a transgene; however, XSxr(b)OEif2s3y transgenic males do not show the expected elimination of MI spermatocytes in response to the univalent [6]. Here we show that these XSxr(b)OEif2s3y males have an impaired apoptotic response with completion of the first meiotic division, but there is no second meiotic division. We then show that Zfy2 (but not the closely related Zfy1) is sufficient to reinstate the apoptotic response to the X univalent. These findings provide further insight into the basis for the much lower transmission of chromosomal errors originating at the first meiotic division in men than in women [7]., (Copyright © 2011 Elsevier Ltd. All rights reserved.)

The mammalian X and Y chromosomes share little homology and are largely unsynapsed during normal meiosis. This asynapsis triggers inactivation of X- and Y-linked genes, or meiotic sex chromosome inactivation (MSCI). Whether MSCI is essential for male meiosis is unclear. Pachytene arrest and apoptosis is observed in mouse mutants in which MSCI fails, e.g., Brca1(-/-), H2afx(-/-), Sycp1(-/-), and Msh5(-/-). However, these also harbor defects in synapsis and/or recombination and as such may activate a putative pachytene checkpoint. Here we present evidence that MSCI failure is sufficient to cause pachytene arrest. XYY males exhibit Y-Y synapsis and Y chromosomal escape from MSCI without accompanying synapsis/recombination defects. We find that XYY males, like synapsis/recombination mutants, display pachytene arrest and that this can be circumvented by preventing Y-Y synapsis and associated Y gene expression. Pachytene expression of individual Y genes inserted as transgenes on autosomes shows that expression of the Zfy 1/2 paralogs in XY males is sufficient to phenocopy the pachytene arrest phenotype; insertion of Zfy 1/2 on the X chromosome where they are subject to MSCI prevents this response. Our findings show that MSCI is essential for male meiosis and, as such, provide insight into the differential severity of meiotic mutations' effects on male and female meiosis., (Copyright © 2010 Elsevier Ltd. All rights reserved.)

The human and mouse sex chromosomes are enriched in multicopy genes required for postmeiotic differentiation of round spermatids into sperm. The gene Sly is present in multiple copies on the mouse Y chromosome and encodes a protein that is required for the epigenetic regulation of postmeiotic sex chromosome expression. The X chromosome carries two multicopy genes related to Sly: Slx and Slxl1. Here we investigate the role of Slx/Slxl1 using transgenically-delivered small interfering RNAs to disrupt their function. We show that Slx and Slxl1 are important for normal sperm differentiation and male fertility. Slx/Slxl1 deficiency leads to delay in spermatid elongation and sperm release. A high proportion of delayed spermatids are eliminated via apoptosis, with a consequent reduced sperm count. The remaining spermatozoa are abnormal with impaired motility and fertilizing abilities. Microarray analyses reveal that Slx/Slxl1 deficiency affects the metabolic processes occurring in the spermatid cytoplasm but does not lead to a global perturbation of sex chromosome expression; this is in contrast with the effect of Sly deficiency which leads to an up-regulation of X and Y chromosome genes. This difference may be due to the fact that SLX/SLXL1 are cytoplasmic while SLY is found in the nucleus and cytoplasm of spermatids.

Differences between males and females are normally attributed to developmental and hormonal differences between the sexes. Here, we demonstrate differences between males and females in gene silencing using a heterochromatin-sensitive reporter gene. Using "sex-reversal" mouse models with varying sex chromosome complements, we found that this differential gene silencing was determined by X chromosome complement, rather than sex. Genome-wide transcription profiling showed that the expression of hundreds of autosomal genes was also sensitive to sex chromosome complement. These genome-wide analyses also uncovered a role for Sry in modulating autosomal gene expression in a sex chromosome complement-specific manner. The identification of this additional layer in the establishment of sexual dimorphisms has implications for understanding sexual dimorphisms in physiology and disease., (Copyright © 2010 Elsevier Inc. All rights reserved.)

Background: X monosomic mice (39,XO) have a remarkably mild phenotype when compared to women with Turner syndrome (45,XO). The generally accepted hypothesis to explain this discrepancy is that the number of genes on the mouse X chromosome which escape X inactivation, and thus are expressed at higher levels in females, is very small. However this hypothesis has never been tested and only a small number of genes have been assayed for their X-inactivation status in the mouse. We performed a global expression analysis in four somatic tissues (brain, liver, kidney and muscle) of adult 40,XX and 39,XO mice using the Illumina Mouse WG-6 v1_1 Expression BeadChip and an extensive validation by quantitative real time PCR, in order to identify which genes are expressed from both X chromosomes., Results: We identified several genes on the X chromosome which are overexpressed in XX females, including those previously reported as escaping X inactivation, as well as new candidates. However, the results obtained by microarray and qPCR were not fully concordant, illustrating the difficulty in ascertaining modest fold changes, such as those expected for genes escaping X inactivation. Remarkably, considerable variation was observed between tissues, suggesting that inactivation patterns may be tissue-dependent. Our analysis also exposed several autosomal genes involved in mitochondrial metabolism and in protein translation which are differentially expressed between XX and XO mice, revealing secondary transcriptional changes to the alteration in X chromosome dosage., Conclusions: Our results support the prediction that the mouse inactive X chromosome is largely silent, while providing a list of the genes potentially escaping X inactivation in rodents. Although the lower expression of X-linked genes in XO mice may not be relevant in the particular tissues/systems which are affected in human X chromosome monosomy, genes deregulated in XO mice are good candidates for further study in an involvement in Turner Syndrome phenotype.

Background: Mice with severe non-PAR Y chromosome long arm (NPYq) deficiencies are infertile in vivo and in vitro. We have previously shown that sperm from these males, although having grossly malformed heads, were able to fertilize oocytes via intracytoplasmic sperm injection (ICSI) and yield live offspring. However, in continuing ICSI trials we noted a reduced efficiency when cryopreserved sperm were used and with epididymal sperm as compared to testicular sperm. In the present study we tested if NPYq deficiency is associated with sperm DNA damage - a known cause of poor ICSI success., Results: We observed that epididymal sperm from mice with severe NPYq deficiency (that is, deletion of nine-tenths or the entire NPYq gene complement) are impaired in oocyte activation ability following ICSI and there is an increased incidence of oocyte arrest and paternal chromosome breaks. Comet assays revealed increased DNA damage in both epididymal and testicular sperm from these mice, with epididymal sperm more severely affected. In all mice the level of DNA damage was increased by freezing. Epididymal sperm from mice with severe NPYq deficiencies also suffered from impaired membrane integrity and abnormal chromatin condensation and suboptimal chromatin protamination. It is therefore likely that the increased DNA damage associated with NPYq deficiency is a consequence of disturbed chromatin remodeling., Conclusions: This study provides the first evidence of DNA damage in sperm from mice with NPYq deficiencies and indicates that NPYq-encoded gene/s may play a role in processes regulating chromatin remodeling and thus in maintaining DNA integrity in sperm.

Studies of mice with Y chromosome long arm deficiencies suggest that the male-specific region (MSYq) encodes information required for sperm differentiation and postmeiotic sex chromatin repression (PSCR). Several genes have been identified on MSYq, but because they are present in more than 40 copies each, their functions cannot be investigated using traditional gene targeting. Here, we generate transgenic mice producing small interfering RNAs that specifically target the transcripts of the MSYq-encoded multicopy gene Sly (Sycp3-like Y-linked). Microarray analyses performed on these Sly-deficient males and on MSYq-deficient males show a remarkable up-regulation of sex chromosome genes in spermatids. SLY protein colocalizes with the X and Y chromatin in spermatids of normal males, and Sly deficiency leads to defective repressive marks on the sex chromatin, such as reduced levels of the heterochromatin protein CBX1 and of histone H3 methylated at lysine 9. Sly-deficient mice, just like MSYq-deficient mice, have severe impairment of sperm differentiation and are near sterile. We propose that their spermiogenesis phenotype is a consequence of the change in spermatid gene expression following Sly deficiency. To our knowledge, this is the first successful targeted disruption of the function of a multicopy gene (or of any Y gene). It shows that SLY has a predominant role in PSCR, either via direct interaction with the spermatid sex chromatin or via interaction with sex chromatin protein partners. Sly deficiency is the major underlying cause of the spectrum of anomalies identified 17 y ago in MSYq-deficient males. Our results also suggest that the expansion of sex-linked spermatid-expressed genes in mouse is a consequence of the enhancement of PSCR that accompanies Sly amplification., Competing Interests: The authors have declared that no competing interests exist.

Background: Attention-deficit/hyperactivity disorder (ADHD) is a complex neurodevelopmental disorder characterized by deficits in attention, increased motor impulsivity, and hyperactivity. Preliminary work in mice and humans has suggested the X-linked gene STS (which encodes the enzyme steroid sulfatase) as a mediator of attentional functioning and as a candidate gene for ADHD., Methods: The effects of modulating the murine steroid sulfatase axis pharmacologically (through administration of the substrate dehydroepiandrosterone sulfate [DHEAS], 0-40 mg/kg, or acute inhibition of the enzyme by COUMATE, 10mg/kg) or genetically (through loss of the gene in 39,X(Y)*O mice) were assayed using the 5-choice serial reaction time task (5-CSRTT) a test of visuospatial attention and response control, and a locomotor activity paradigm., Results: DHEAS administration improved 5-CSRTT performance under attentionally demanding conditions, whereas steroid sulfatase inhibition impaired accuracy under the same conditions. Loss of Sts expression constitutively throughout development in 39,X(Y)*O mice resulted in deficits in 5-CSRTT performance at short stimulus durations and reduced anticipatory responding. Neither the pharmacologic nor the genetic manipulations affected basic locomotor activity., Conclusions: These data provide converging evidence indicating a role for steroid sulfatase in discrete aspects of attentional functioning and are suggestive of a role in motor impulsivity. The findings provide novel insights into the neurobiology of attention and strengthen the notion of STS as a candidate gene for the attentional component of ADHD.

Deletion analysis has established that genes on the Y chromosome are essential for normal sperm production in humans, mice, and Drosophila. In mice, long-arm deletions have an impact on spermiogenesis, with the most extensive deletions resulting in severe sperm head malformations and infertility. Intriguingly, smaller deletions are compatible with fertility but result in a distorted sex ratio in favor of females, and recently it was found that Y long-arm deletions are also associated with a marked upregulation of several X-encoded and Y-encoded spermatid-expressed genes. The mouse Y long arm encodes a number of distinct transcripts, each of which derives from multiple gene copies. Of these multicopy genes, the recently described Sly has been favored as the gene underlying the spermiogenic defects associated with Y long-arm deletions. To assess the candidacy of Sly, the expression of this gene was examined in the testis at the transcript and protein levels. Sly is transcribed after the first meiotic division in secondary spermatocytes and round spermatids and encodes two transcript variants, Sly_v1 and Sly_v2 (proteins referred to as SLY1 and SLY2). We raised an antibody against SLY1 which detected the protein in round and early elongating spermatids, where it is predominantly cytoplasmic. Yeast two-hybrid and coimmunoprecipitation studies demonstrated that SLY1 interacts with the acrosomal protein DKKL1, the histone acetyltransferase KAT5 (also known as TIP60), and the microtubule-associated protein APPBP2. Together, these data suggest SLY1 may be involved in multiple processes during spermiogenesis, including the control of gene expression and the development or function of the acrosome.

The mouse Y chromosome long arm (Yq) comprises approximately 70 Mb of repetitive, male-specific DNA together with a short (0.7-Mb) pseudoautosomal region (PAR). The repetitive non-PAR region (NPYq) encodes genes whose deficiency leads to subfertility and infertility, resulting from impaired spermiogenesis. In XSxr(a)Y*(X) mice, the only Y-specific material is provided by the Y chromosome short arm-derived sex reversal factor Sxr(a), which is attached to the X chromosome PAR; these males (NPYq- males) produce sperm with severely malformed heads and are infertile. In the present study, we investigated sperm function in these mice in the context of intracytoplasmic sperm injection (ICSI). Of 261 oocytes injected, 103 reached the 2-cell stage, and 46 developed to liveborn offspring. Using Xist RT-PCR genotyping as well as gamete and somatic cell karyotyping, all six predicted genotypes were identified among ICSI-derived progeny. The sex chromosome constitution of NPYq- males does not allow production of offspring with the same genotype, but one of the expected offspring genotypes is XY*(X)Sxr(a) (NPYq-(2)), which has the same Y gene complement as NPYq-. Analysis of NPYq-(2) males revealed they had normal-sized testes with ongoing spermatogenesis. Like NPYq- males, these males were infertile, and their sperm had malformed heads that nevertheless fertilized eggs via ICSI. In vitro fertilization (IVF), however, was unsuccessful. Overall, we demonstrated that a lack of NPYq-encoded genes does not interfere with the ability of sperm to fertilize oocytes via ICSI but does prevent fertilization via IVF. Thus, NPYq-encoded gene functions are not required after the sperm have entered the oocyte. The present work also led to development of a new mouse model lacking NPYq gene complement that will facilitate future studies of Y-encoded gene function.

During mammalian meiosis, synapsis of paternal and maternal chromosomes and the generation of DNA breaks are needed to allow reshuffling of parental genes. In mammals errors in synapsis are associated with a male-biased meiotic impairment, which has been attributed to a response to persisting DNA double-stranded breaks in the asynapsed chromosome segments. Recently it was discovered that the chromatin of asynapsed chromosome segments is transcriptionally silenced, providing new insights into the connection between asynapsis and meiotic impairment.

Chromosome synapsis during zygotene is a prerequisite for the timely homologous recombinational repair of meiotic DNA double-strand breaks (DSBs). Unrepaired DSBs are thought to trigger apoptosis during midpachytene of male meiosis if synapsis fails. An early pachytene response to asynapsis is meiotic silencing of unsynapsed chromatin (MSUC), which, in normal males, silences the X and Y chromosomes (meiotic sex chromosome inactivation [MSCI]). In this study, we show that MSUC occurs in Spo11-null mouse spermatocytes with extensive asynapsis but lacking meiotic DSBs. In contrast, three mutants (Dnmt3l, Msh5, and Dmc1) with high levels of asynapsis and numerous persistent unrepaired DSBs have a severely impaired MSUC response. We suggest that MSUC-related proteins, including the MSUC initiator BRCA1, are sequestered at unrepaired DSBs. All four mutants fail to silence the X and Y chromosomes (MSCI failure), which is sufficient to explain the midpachytene apoptosis. Apoptosis does not occur in mice with a single additional asynapsed chromosome with unrepaired meiotic DSBs and no disturbance of MSCI.

To shed light on the biological origins of sex differences in neural tube defects (NTDs), we examined Trp53-null C57BL/6 mouse embryos and neonates at 10.5 and 18.5 days post coitus (dpc) and at birth. We confirmed that female embryos show more NTDs than males. We also examined mice in which the testis-determining gene Sry is deleted from the Y chromosome but inserted onto an autosome as a transgene, producing XX and XY gonadal females and XX and XY gonadal males. At birth, Trp53 nullizygous mice were predominantly XY rather than XX, irrespective of gonadal type, showing that the sex difference in the lethal effect of Trp53 nullizygosity by postnatal day 1 is caused by differences in sex chromosome complement. At 10.5 dpc, the incidence of NTDs in Trp53-null progeny of XY* mice, among which the number of the X chromosomes varies independently of the presence or absence of a Y chromosome, was higher in mice with two copies of the X chromosome than in mice with a single copy. The presence of a Y chromosome had no protective effect, suggesting that sex differences in NTDs are caused by sex differences in the number of X chromosomes.

DNA double-strand breaks (DSBs) are extremely hazardous lesions for all DNA-bearing organisms and the mechanisms of DSB repair are highly conserved. In the eukaryotic mitotic cell cycle, DSBs are often present following DNA replication while, in meiosis, hundreds of DSBs are generated as a prelude to the reshuffling of the maternally and paternally derived genomes. In both cases, the DSBs are repaired by a process called homologous recombinational repair (HRR), which utilises an intact DNA molecule as the repair template. Mitotic and meiotic HRR are managed by 'checkpoints' that inhibit cell division until DSB repair is complete. Here we attempt to summarise the substantial recent progress in understanding the checkpoint management of HRR in mitosis (focussing mainly on mammals) and then go on to use this information as a framework for understanding the presumed checkpoint management of HRR in mammalian meiosis.

The mouse multi-copy X-linked gene Xlr-related, meiosis-regulated (Xmr/Slx) has previously been described as encoding a testis-specific nuclear protein expressed during male meiotic prophase, and during which it becomes concentrated in the inactive X and Y chromatin domain. These conclusions were based on Western blot and immunolocalization analysis using an antibody raised against a related lymphocyte protein, XLR; however, our recently published RNA in situ for Xmr revealed that transcripts are predominantly or exclusively postmeiotic, and this is supported by a growing body of microarray data. This led us to reanalyze the expression of Xmr, both at the RNA level by RT-PCR and by RNA fluorescence in situ hybridization, and at the protein level by using antibodies raised against XMR that do not recognize XLR. In agreement with our previous RNA in situ data, our further transcription analysis showed almost exclusive expression in spermatids, and Western blot and immunostaining with the XMR antibodies showed that the protein is cytoplasmic and restricted to spermatids. Furthermore, the previously used XLR antibody was shown not to cross-react with XMR, and it is suggested that the meiotically expressed nuclear protein recognized by this antibody is another member of the complex Xlr superfamily. As a result of these findings, the gene previously known as Xmr is now officially know as Slx, Sycp3-like, X-linked.

Background: The loss of all, or part of an X chromosome, in Turner syndrome (TS, 45,XO) results in deficits in attentional functioning., Methods: Using a 39,XO mouse model, we tested the hypothesis that X-monosomy and/or parental origin of the single X chromosome may influence visuospatial attentional functioning in a 5-choice serial reaction time task (5-CSRTT)., Results: Under attentionally demanding conditions 39,XO mice displayed impaired discriminative response accuracy and slowed correct reaction times relative to 40,XX mice; these deficits were alleviated in a version of the task with reduced attentional demands. Parental origin of the X did not affect performance of the 5-CSRTT. In contrast, the attentional phenotype was rescued in 40,XY*X mice possessing a single maternally inherited X chromosome and a small Y*X chromosome that comprises a complete pseudoautosomal region (PAR), and a small X-specific segment., Conclusions: Our findings are consistent with an X-monosomy effect on attention and suggest the existence of X-linked gene(s) that escape X-inactivation, are present on the small Y*X chromosome and impact on attentional functioning; the strongest candidate gene is Sts, encoding steroid sulfatase. The data inform the TS literature and indicate novel genetic mechanisms that may be of general significance to the neurobiology of attention.

The product of the Scmh1 gene, a mammalian homolog of Drosophila Sex comb on midleg, is a constituent of the mammalian Polycomb repressive complexes 1 (Prc1). We have identified Scmh1 as an indispensable component of the Prc1. During progression through pachytene, Scmh1 was shown to be excluded from the XY body at late pachytene, together with other Prc1 components such as Phc1, Phc2, Rnf110 (Pcgf2), Bmi1 and Cbx2. We have identified the role of Scmh1 in mediating the survival of late pachytene spermatocytes. Apoptotic elimination of Scmh1(-/-) spermatocytes is accompanied by the preceding failure of several specific chromatin modifications at the XY body, whereas synapsis of homologous autosomes is not affected. It is therefore suggested that Scmh1 is involved in regulating the sequential changes in chromatin modifications at the XY chromatin domain of the pachytene spermatocytes. Restoration of defects in Scmh1(-/-) spermatocytes by Phc2 mutation indicates that Scmh1 exerts its molecular functions via its interaction with Prc1. Therefore, for the first time, we are able to indicate a functional involvement of Prc1 during the meiotic prophase of male germ cells and a regulatory role of Scmh1 for Prc1, which involves sex chromosomes.

Unlabelled: In mouse and man, Y chromosome deletions are frequently associated with spermatogenic defects. XY(Tdy)(m1)qdelSry males have an extensive Yq deletion that almost completely abolishes the expression of two gene families, Ssty and Sly, located within the male-specific region of the mouse Y long arm. These males exhibit severe sperm defects and sterility. XY(RIII)qdel males have a smaller interstitial Yq deletion, removing approximately two thirds of Ssty/Sly gene copies, and display an increased incidence of mild sperm head anomalies with impairment of fertility and an intriguing distortion in the sex ratio of offspring in favor of females. Here we used intracytoplasmic sperm injection (ICSI) to investigate the functional capacity of sperm from these Yq deletion males. Any selection related to the ability of sperm to fertilize in vitro is removed by ICSI, and we obtained two generations of live offspring from the infertile males. Genotyping of ICSI-derived offspring revealed that the Y(Tdym1)qdel deletion does not interfere with production of Y chromosome-bearing gametes, as judged from the frequency of Y chromosome transmission to the offspring. ICSI results for XY(RIII)qdel males also indicate that there is no deficiency of Y sperm production in this genotype, although the data show an excess of females following in vitro fertilization and natural mating. Our findings suggest that 1) Yq deletions in mice do not bias the primary sex ratio and 2) Y(RIII)qdel spermatozoa have poorer fertilizing ability than their X-bearing counterparts. Thus, a normal complement of the Ssty and/or Sly gene families on mouse Yq appears necessary for normal sperm function., Summary: ICSI was successfully used to reproduce infertile mice with Yq deletions, and the analysis of sperm function in obtained offspring demonstrated that gene families located within the deletion interval are necessary for normal sperm function.

Transcriptional silencing of the sex chromosomes during male meiosis (MSCI) is conserved among organisms with limited sex chromosome synapsis, including mammals. Since the 1990s the prevailing view has been that MSCI in mammals is transient, with sex chromosome reactivation occurring as cells exit meiosis. Recently, we found that any chromosome region unsynapsed during pachytene of male and female mouse meiosis is subject to transcriptional silencing (MSUC), and we hypothesized that MSCI is an inevitable consequence of this more general meiotic silencing mechanism. Here, we provide direct evidence that asynapsis does indeed drive MSCI. We also show that a substantial degree of transcriptional repression of the sex chromosomes is retained postmeiotically, and we provide evidence that this postmeiotic repression is a downstream consequence of MSCI/MSUC. While this postmeiotic repression occurs after the loss of MSUC-related proteins at the end of prophase, other histone modifications associated with transcriptional repression have by then become established.

Across human cultures and mammalian species, sex differences can be found in the expression of aggression and parental nurturing behaviors: males are typically more aggressive and less parental than females. These sex differences are primarily attributed to steroid hormone differences during development and/or adulthood, especially the higher levels of androgens experienced by males, which are caused ultimately by the presence of the testis-determining gene Sry on the Y chromosome. The potential for sex differences arising from the different complements of sex-linked genes in male and female cells has received little research attention. To directly test the hypothesis that social behaviors are influenced by differences in sex chromosome complement other than Sry, we used a transgenic mouse model in which gonadal sex and sex chromosome complement are uncoupled. We find that latency to exhibit aggression and one form of parental behavior, pup retrieval, can be influenced by both gonadal sex and sex chromosome complement. For both behaviors, females but not males with XX sex chromosomes differ from XY. We also measured vasopressin immunoreactivity in the lateral septum, which was higher in gonadal males than females, but also differed according to sex chromosome complement. These results imply that a gene(s) on the sex chromosomes (other than Sry) affects sex differences in brain and behavior. Identifying the specific X and/or Y genes involved will increase our understanding of normal and abnormal aggression and parental behavior, including behavioral abnormalities associated with mental illness.

Imprinted genes are monoallelically expressed in a parent-of-origin-dependent manner and can affect brain and behavioural phenotypes. The X chromosome is enriched for genes affecting neurodevelopment and is donated asymmetrically to male and female progeny. Hence, X-linked imprinted genes could potentially influence sexually dimorphic neurobiology. Consequently, investigations into such loci may provide new insights into the biological basis of behavioural differences between the sexes and into why men and women show different vulnerabilities to certain mental disorders. In this review, we summarise recent advances in our knowledge of X-linked imprinted genes and the brain substrates that they may act upon. In addition, we suggest strategies for identifying novel X-linked imprinted genes and their downstream effects and discuss evolutionary theories regarding the origin and maintenance of X-linked imprinting., (2005 Wiley Periodicals, Inc.)

Deletions on the mouse Y-chromosome long arm (MSYq) lead to teratozoospermia and in severe cases to infertility. We find that the downstream transcriptional changes in the testis resulting from the loss of MSYq-encoded transcripts involve upregulation of multiple X- and Y-linked spermatid-expressed genes, but not related autosomal genes. Therefore, this indicates that in normal males, there is a specific repression of X and Y (gonosomal) transcription in post-meiotic cells, which depends on MSYq-encoded transcripts. Together with the known sex ratio skew in favour of females in the offspring of fertile MSYqdel males, this strongly suggests the existence of an intragenomic conflict between X- and Y-linked genes. Two potential antagonists in this conflict are the X-linked multicopy gene Xmr and its multicopy MSYq-linked relative Sly, which are upregulated and downregulated, respectively, in the testes of MSYqdel males. Xmr is also expressed during meiotic sex chromosome inactivation (MSCI), indicating a link between the MSCI and the MSYq-dependent gonosomal repression in spermatids. We therefore propose that this repression and MSCI itself are evolutionary adaptations to maintain a normal sex ratio in the face of X/Y antagonism.

Sex chromosome complement, by determining whether an ovary or testis develops, exerts indirect hormone-mediated effects on the development of sex-specific traits. However, this does not preclude more direct effects that are independent of gonadal hormones. To look for gonadal hormone-independent effects in sexually dimorphic immune responses, we used mice in which the testis determinant Sry has been moved from the Y chromosome to an autosome, thus allowing the production of mice that differ in sex chromosome complement while having the same gonadal type. This model permits comparison of XX and XY mice with ovaries or testes. These mice were immunized with an autoantigen, and draining lymph node cells were assessed for autoantigen-specific proliferative responses and cytokine production. Surprisingly, we found that the male complement of sex chromosomes (XY) was relatively stimulatory, whereas male sex hormones were inhibitory, for this immune response. This is the first experimental evidence of a compensatory yin-yang effect of sex chromosome complement and sex hormones on a biologic process.

Fundamentally different recombination defects cause apoptosis of mouse spermatocytes at the same stage in development, stage IV of the seminiferous epithelium cycle, equivalent to mid-pachynema in normal males. To understand the cellular response(s) that triggers apoptosis, we examined markers of spermatocyte development in mice with different recombination defects. In Spo11(-)(/)(-) mutants, which lack the double-strand breaks (DSBs) that initiate recombination, spermatocytes express markers of early to mid-pachynema, forming chromatin domains that contain sex body-associated proteins but that rarely encompass the sex chromosomes. Dmc1(-)(/)(-) spermatocytes, impaired in DSB repair, appear to arrest at or about late zygonema. Epistasis analysis reveals that this earlier arrest is a response to unrepaired DSBs, and cytological analysis implicates the BRCT-containing checkpoint protein TOPBP1. Atm(-)(/)(-) spermatocytes show similarities to Dmc1(-)(/)(-) spermatocytes, suggesting that ATM promotes meiotic DSB repair. Msh5(-)(/)(-) mutants display a set of characteristics distinct from these other mutants. Thus, despite equivalent stages of spermatocyte elimination, different recombination-defective mutants manifest distinct responses, providing insight into surveillance mechanisms in male meiosis.

Background: The male-specific region of the mouse Y chromosome long arm (MSYq) is comprised largely of repeated DNA, including multiple copies of the spermatid-expressed Ssty gene family. Large deletions of MSYq are associated with sperm head defects for which Ssty deficiency has been presumed to be responsible., Results: In a search for further candidate genes associated with these defects we analyzed changes in the testis transcriptome resulting from MSYq deletions, using testis cDNA microarrays. This approach, aided by accumulating mouse MSYq sequence information, identified transcripts derived from two further spermatid-expressed multicopy MSYq gene families; like Ssty, each of these new MSYq gene families has multicopy relatives on the X chromosome. The Sly family encodes a protein with homology to the chromatin-associated proteins XLR and XMR that are encoded by the X chromosomal relatives. The second MSYq gene family was identified because the transcripts hybridized to a microarrayed X chromosome-encoded testis cDNA. The X loci ('Astx') encoding this cDNA had 92-94% sequence identity to over 100 putative Y loci ('Asty') across exons and introns; only low level Asty transcription was detected. More strongly transcribed recombinant loci were identified that included Asty exons 2-4 preceded by Ssty1 exons 1, 2 and part of exon 3. Transcription from the Ssty1 promotor generated spermatid-specific transcripts that, in addition to the variable inclusion of Ssty1 and Asty exons, included additional exons because of the serendipitous presence of splice sites further downstream., Conclusion: We identified further MSYq-encoded transcripts expressed in spermatids and deriving from multicopy Y genes, deficiency of which may underlie the defects in sperm development associated with MSYq deletions.

In Neurospora, DNA unpaired in meiosis both is silenced and induces silencing of all DNA homologous to it. This process, called meiotic silencing by unpaired DNA, is thought to protect the host genome from invasion by transposable elements. We now show that silencing of unpaired (unsynapsed) chromosome regions also takes place in the mouse during both male and female meiosis. The tumor suppressor protein BRCA1 is implicated in this silencing, mirroring its role in the meiotic silencing of the X and Y chromosomes in normal male meiosis. These findings impact on the interpretation of the relationship between synaptic errors and sterility in mammals and extend our understanding of the biology of Brca1.

In mammalian spermatogenesis, the X and Y chromosomes are transcriptionally silenced during the pachytene stage of meiotic prophase (meiotic sex chromosome inactivation, MSCI), forming a condensed chromatin domain termed the sex or XY body. The nucleosomal core histone H2AX is phosphorylated within the XY chromatin domain just prior to MSCI, and it has been hypothesized that this triggers the chromatin condensation and transcriptional repression. Here, we show that the kinase ATR localizes to XY chromatin at the onset of MSCI and that this localization is disrupted in mice with a mutant form of the tumor suppressor protein BRCA1. In the mutant pachytene cells, ATR is usually present at nonsex chromosomal sites, where it colocalizes with aberrant sites of H2AX phosphorylation; in these cells, there is MSCI failure. In rare pachytene cells, ATR does locate to XY chromatin, H2AX is then phosphorylated, a sex body forms, and MSCI ensues. These observations highlight an important role for BRCA1 in recruiting the kinase ATR to XY chromatin at the onset of MSCI and provide compelling evidence that it is ATR that phosphorylates H2AX and triggers MSCI.

Recent work has indicated altered emotional functioning in Turner's syndrome (TS) subjects (45,XO). We examined the role of X-chromosome deficiency on fear reactivity in X-monosomic mice (39,XO), and found that they exhibited anxiogenic behaviour relative to normal females (40,XX). A molecular candidate for this effect is Steroid sulfatase (Sts) as this is located in the pseudoautosomal region (PAR) of the X-chromosome and consequently is normally biallelically expressed. In addition, the steroid sulfatase enzyme (STS) is putatively linked to fear reactivity by an effect on GABAA receptors via the action of neurosteroids. Real-time PCR demonstrated that levels of Sts mRNA were reduced by half in the brains of 39,XO mice compared with 40,XX, and that expression levels of a number of GABAA subunits previously shown to be important components of fear processing (Gabra3, Gabra1 and Gabrg2) were also altered. However, 40,XY*X mice, in which the Y*X is a small chromosome comprising of a complete PAR and a small non-PAR segment of the X-chromosome, exhibited the same pattern of fear reactivity behaviour as 39,XO animals, but equivalent expression levels of Sts, Gabra1, Gabra3 and Gabrg2 to 40,XX females. This showed that although Sts may cause alterations in GABAA subunit expression, these changes do not result in increased fear reactivity. This suggests an alternative X-chromosome gene, that escapes inactivation, is responsible for the differences in fear reactivity between 39,XO and 40,XX mice. These findings inform the TS data, and point to novel genetic mechanisms that may be of general significance to the neurobiology of fear., (Copyright 2004 Oxford University Press)

The aim of this study is to develop an overview of genetic events during spermatogenesis using a novel, specifically targeted gonadal gene set. Two subtracted cDNA libraries enriched for testis specific and germ cell specific genes were constructed, characterized and sequenced. The combined libraries contain >1905 different genes, the vast majority previously uncharacterized in testis. cDNA microarray analysis of the first wave of murine spermatogenesis and of selected germ cell-deficient models was used to correlate the expression of groups of genes with the appearance of defined germ cell types, suggesting their cellular expression patterns within the testis. Real-time RT-PCR and comparison to previously known expression patterns confirmed the array-derived transcription profiles of 65 different genes, thus establishing high confidence in the profiles of the uncharacterized genes investigated in this study. A total of 1748 out of 1905 genes showed significant change during the first spermatogenic wave, demonstrating the successful targeting of the libraries to this process. These findings highlight unknown genes likely to be important in germ cell production, and demonstrate the utility of these libraries in further studies. Transcriptional analysis of well-characterized mouse models of infertility will allow us to address the causes and progression of the pathology in related human infertility phenotypes.

The mouse Y chromosome carries 10 distinct genes or gene families that have open reading frames suggestive of retained functionality; it has been assumed that many of these function in spermatogenesis. However, we have recently shown that only two Y genes, the testis determinant Sry and the translation initiation factor Eif2s3y, are essential for spermatogenesis to proceed to the round spermatid stage. Thus, any further substantive mouse Y-gene functions in spermatogenesis are likely to be during sperm differentiation. The complex Ssty gene family present on the mouse Y long arm (Yq) has been implicated in sperm development, with partial Yq deletions that reduce Ssty expression resulting in impaired fertilization efficiency. Here we report the identification of a more extensive Yq deletion that abolishes Ssty expression and results in severe sperm defects and sterility. This result establishes that genetic information (Ssty?) essential for normal sperm differentiation and function is present on mouse Yq.

Histone macroH2A1.2 and the murine heterochromatin protein 1, HP1 beta, have both been implicated in meiotic sex chromosome inactivation (MSCI) and the formation of the XY-body in male meiosis. In order to get a closer insight into the function of histone macroH2A1.2 we have investigated the localisation of macroH2A1.2 in surface spread spermatocytes from normal male mice and in oocytes of XX and XYTdym1 mice. Oocytes of XYTdym1 mice have no XY-body or MSCI despite having an XY chromosome constitution, so the presence or absence of 'XY-body' proteins in association with the X and/or Y chromosome of these oocytes enables some discrimination between potential functions of XY-body located proteins. We demonstrate here that macroH2A1.2 localises to the X and Y chromatin of spermatocytes as they condense to form the XY-body but is not associated with the X and Y chromatin of XYTdym1 early pachytene oocytes. MacroH2A1.2 and HP1 beta co-localise to autosomal pericentromeric heterochromatin in spermatocytes. However, the two proteins show temporally and spatially distinct patterns of association to X and Y chromatin.

Multicopy Y-chromosomal genes in human and mouse have been postulated to play a role in spermatogenesis. The mouse Y long arm (Yq) carries hundreds of supposedly intronless copies of Ssty, for which no protein has hitherto been identified; mice lacking Yq are sterile with grossly abnormal sperm. We have now identified an Ssty-encoded protein (Ssty1) that is expressed in spermatids. The protein is absent from spermatids of mice that lack Yq, but is not reduced in mice with a two-thirds reduction of Ssty copies, implying that most do not produce this protein. Furthermore, no protein was produced by a strongly transcribed intronless Ssty transgene, raising doubts as to the protein-encoding potential of these intronless genes. We have now identified an intron-containing copy that is also present in multiple copies on Yq. One or more intron-containing copies are retained in the Ssty-deficient mice and may be the source of the Ssty1 protein.

In mammals and birds, the sex of the gonads is determined by genes on the sex chromosomes. For example, the mammalian Y-linked gene Sry causes testis differentiation. The testes then secrete testosterone, which acts on the brain (often after conversion to estradiol) to cause masculine patterns of development. If this were the only reason for sex differences in neural development, then XX and XY brain cells would have to be deemed otherwise equivalent. This equivalence is doubtful because of recent experimental results demonstrating that some XX and XY tissues, including the brain, are sexually dimorphic even when they develop in a similar endocrine environment. Although X and Y genes probably influence brain phenotype in a sex-specific manner, much more information is needed to identify the magnitude and character of these effects.

Mice of the XO genotype with a paternally derived X chromosome (XpO) have placental hyperplasia in late pregnancy, although in early pregnancy the ectoplacental cone, a placental precursor, is smaller in XpO mice than in their XX sibs. This early size deficiency of the ectoplacental cone is apparently a consequence of Xp imprinting, because XmO embryos (with a maternally derived X chromosome) are unaffected. In the present study we sought to establish whether XpO placental hyperplasia in late pregnancy is also a consequence of Xp imprinting. Placental weight data were first collected from litters that included XpO or XmO fetuses and XX controls. Comparison of XO placentae with XX placentae showed that XpO and XmO placentae are hyperplastic. This finding suggested that the hyperplasia might be an X dosage effect, and this hypothesis was supported by the finding that XY male fetuses from the same crosses also had larger placentae than their XX sibs. Further analysis of a range of sex-chromosome variant genotypes, including XmYSry-negative females and XXSry transgenic males, showed that mouse fetuses with one X chromosome consistently had larger placentae than littermates with two X chromosomes, independent of their gonadal/androgen status.