Dosage regulation, and variation in gene expression and copy number of human Y chromosome ampliconic genes.

The Y chromosome harbors nine multi-copy ampliconic gene families expressed exclusively in testis. The gene copies within each family are >99% identical to each other, which poses a major challenge in evaluating their copy number. Recent studies demonstrated high variation in Y ampliconic gene copy number among humans. However, how this variation affects expression levels in human testis remains understudied. Here we developed a novel computational tool Ampliconic Copy Number Estimator (AmpliCoNE) that utilizes read sequencing depth information to estimate Y ampliconic gene copy number per family. We applied this tool to whole-genome sequencing data of 149 men with matched testis expression data whose samples are part of the Genotype-Tissue Expression (GTEx) project. We found that the Y ampliconic gene families with low copy number in humans were deleted or pseudogenized in non-human great apes, suggesting relaxation of functional constraints. Among the Y ampliconic gene families, higher copy number leads to higher expression. Within the Y ampliconic gene families, copy number does not influence gene expression, rather a high tolerance for variation in gene expression was observed in testis of presumably healthy men. No differences in gene expression levels were found among major Y haplogroups. Age positively correlated with expression levels of the HSFY and PRY gene families in the African subhaplogroup E1b, but not in the European subhaplogroups R1b and I1. We also found that expression of five Y ampliconic gene families is coordinated with that of their non-Y (i.e. X or autosomal) homologs. Indeed, five ampliconic gene families had consistently lower expression levels when compared to their non-Y homologs suggesting dosage regulation, while the HSFY family had higher expression levels than its X homolog and thus lacked dosage regulation.

Males as somatic investment in a parthenogenetic nematode.

We report the reproductive strategy of the nematode Mesorhabditis belari This species produces only 9% males, whose sperm is necessary to fertilize and activate the eggs. However, most of the fertilized eggs develop without using the sperm DNA and produce female individuals. Only in 9% of eggs is the male DNA utilized, producing sons. We found that mixing of parental genomes only gives rise to males because the Y-bearing sperm of males are much more competent than the X-bearing sperm for penetrating the eggs. In this previously unrecognized strategy, asexual females produce few sexual males whose genes never reenter the female pool. Here, production of males is of interest only if sons are more likely to mate with their sisters. Using game theory, we show that in this context, the production of 9% males by M. belari females is an evolutionary stable strategy.

Within-population Y-linked genetic variation for lifespan in Drosophila melanogaster.

The view that the Y chromosome is of little importance for phenotypic evolution stems from early studies of Drosophila melanogaster. This species' Y chromosome contains only 13 protein-coding genes, is almost entirely heterochromatic and is not necessary for male viability. Population genetic theory further suggests that non-neutral variation can only be maintained at the Y chromosome under special circumstances. Yet, recent studies suggest that the D. melanogaster Y chromosome trans-regulates hundreds to thousands of X and autosomal genes. This finding suggests that the Y chromosome may play a far more active role in adaptive evolution than has previously been assumed. To evaluate the potential for the Y chromosome to contribute to phenotypic evolution from standing genetic variation, we test for Y-linked variation in lifespan within a population of D. melanogaster. Assessing variation for lifespan provides a powerful test because lifespan (i) shows sexual dimorphism, which the Y is primarily predicted to contribute to, (ii) is influenced by many genes, which provides the Y with many potential regulatory targets and (iii) is sensitive to heterochromatin remodelling, a mechanism through which the Y chromosome is believed to regulate gene expression. Our results show a small but significant effect of the Y chromosome and thus suggest that the Y chromosome has the potential to respond to selection from standing genetic variation. Despite its small effect size, Y-linked variation may still be important, in particular when evolution of sexual dimorphism is genetically constrained elsewhere in the genome.

Roles of the Y chromosome genes in human cancers.

Male and female differ genetically by their respective sex chromosome composition, that is, XY as male and XX as female. Although both X and Y chromosomes evolved from the same ancestor pair of autosomes, the Y chromosome harbors male-specific genes, which play pivotal roles in male sex determination, germ cell differentiation, and masculinization of various tissues. Deletions or translocation of the sex-determining gene, SRY, from the Y chromosome causes disorders of sex development (previously termed as an intersex condition) with dysgenic gonads. Failure of gonadal development results not only in infertility, but also in increased risks of germ cell tumor (GCT), such as gonadoblastoma and various types of testicular GCT. Recent studies demonstrate that either loss of Y chromosome or ectopic expression of Y chromosome genes is closely associated with various male-biased diseases, including selected somatic cancers. These observations suggest that the Y-linked genes are involved in male health and diseases in more frequently than expected. Although only a small number of protein-coding genes are present in the male-specific region of Y chromosome, the impacts of Y chromosome genes on human diseases are still largely unknown, due to lack of in vivo models and differences between the Y chromosomes of human and rodents. In this review, we highlight the involvement of selected Y chromosome genes in cancer development in men.

Identification of sexually dimorphic genes in the neonatal mouse cortex and hippocampus.

The cerebral cortex and hippocampus are important for the control of cognitive functions and social behaviors, many of which are sexually dimorphic and tightly regulated by gonadal steroid hormones via activation of their respective nuclear receptors. As different levels of sex steroid hormones are present between the sexes during early development and their receptors act as transcription factors to regulate gene expression, we hypothesize that sexually dimorphic gene expression in the developing mouse cortex and hippocampus might result in sex differences in brain structures and neural circuits governing distinct behaviors between the sexes as adults. To test our hypothesis, we used gene expression microarrays to identify 90 candidate genes differentially expressed in the neonatal cortex/hippocampus between male and female mice, including 55 male-biased and 35 female-biased genes. Among these genes, sexually dimorphic expression of eight sex chromosome genes was confirmed by reverse transcription with quantitative PCR (RT-qPCR), including three located on the X chromosome (Xist, Eif2s3x, and Kdm6a), three on the Y chromosome (Ddx3y, Eif2s3y, and Kdm5d), and two in the pseudoautosomal region of the X and Y chromosomes (Erdr1 and Mid1). In addition, five autosomal genes (Cd151, Dab2, Klk8, Meg3, and Prkdc) were also validated for their sexually dimorphic expression in the neonatal mouse cortex/hippocampus. Gene Ontology annotation analysis suggests that many of these sexually dimorphic genes are involved in histone modifications, cell proliferation/death, androgen/estrogen signaling pathways, and synaptic organization, and these biological processes have been implicated in differential neural development, cognitive function, and neurological diseases between the sexes.

Sex control by Zfy siRNA in the mouse.

The objective of this work was to detect the influence of Y sperm forming of Mus musculus by silencing the Zfy gene during spermatogenesis. The recombination expression vectors pSilencer5.1/Zfy215 and pSilencer5.1/Zfy2102 were constructed. 64 male KunMing Mus were divided into four groups randomly and averagely. The two recombination expression vectors were injected into two groups, respectively, through testis. The other two groups were injected with the same volume of physiological saline and empty vector pSilencer5.1-H1 Retro, respectively. They were injected every ten days for a total of four injections. Seventeen days after the fourth injection, 8 male Mus of each group mated with 8 female Mus. The testis tissue of the other 8 male Mus of each group was collected, and the expression level of Zfy mRNA was determined by fluorescence quantitation real time PCR (qRT-PCR). The result showed that the expression of Zfy mRNA decreased significantly after injection of pSilencer5.1/Zfy2102 (P < 0.01), and that 72.3% of the offspring were female, a number significantly higher than in the control group (P < 0.01). In the pSilencer5.1/Zfy215 group, the expression of Zfy mRNA was significantly lower than in the control group (P < 0.05), but the female rate of offspring was not. It was concluded that the Zfy gene could play a role in the process of Y sperm formation.

Parent-of-origin effects on cardiac response to pressure overload in mice.

Left ventricular (LV) hypertrophy (LVH) is an independent risk factor for cardiovascular mortality and is commonly caused by hypertension. In rodents, transverse aortic constriction (TAC) is a model regularly employed in mechanistic studies of the response of the LV to pressure overload. We previously reported that inbred strains of male mice manifest different cardiac responses to TAC, with C57BL/6J (B6) developing LV dilatation and impaired contractility and 129S1/SvImJ (129) males displaying concentric LVH. In the present study, we investigated sex and parent-of-origin effects on the response to TAC by comparing cardiac function, organ weights, expression of cardiac hypertrophy markers, and histology in female B6 and female 129 mice and in F1 progeny of reciprocal crosses between B6 and 129 mice (B6129F1 and 129B6F1). Five weeks after TAC, heart weight increased to the greatest extent in 129B6F1 mice and the least extent in 129 and B6129F1 mice. Female 129B6F1 and B6 mice were relatively protected from the increase in heart weight that occurs in their male counterparts with pressure overload. The response to TAC in 129 consomic mice bearing the B6 Y chromosome resembled that of 129 rather than 129B6F1 mice, indicating that the B6 Y chromosome does not account for the differences in the reciprocal cross. Our results suggest that susceptibility to LVH is more complex than simple Mendelian inheritance and that parental origin effects strongly impact the LV response to TAC in these commonly used inbred strains.

Changes in testosterone or temperature during the in vitro oocyte culture do not alter the sex ratio of bovine embryos.

High follicular testosterone levels have been associated with a skew in the sex ratio in favor of males following in vitro fertilization, whereas egg incubation temperature has been found to influence sex ratio in some reptiles. The incubation temperature interferes with the aromatase activity, resulting in a sex determination mechanism thought to be lost in mammals. In this work we aimed to test the effects of testosterone on sex ratio of bovine embryos produced in vitro and to determine whether effects of sex and temperature are effectively decoupled in mammals. Bovine oocytes were in vitro matured for 22 hr in TCM199, PVA, FSH and LH after a 22 hr meiotic arrest in TCM199, PVA and roscovitine 25 microM. Matured oocytes were in vitro fertilized and cultured up to Day 3, and embryos having three or more cells were sexed. In the first experiment, testosterone (0, 30, 300 and 1,500 nM), present both during meiotic inhibition and subsequent in vitro maturation (IVM), did not affect development rates or embryonic sex ratio. In the second experiment, increasing incubation temperatures (38, 39 or 40 degrees C) during meiotic inhibition and subsequent IVM, reduced embryo development, but did not change the sex ratio. Under our experimental conditions, testosterone does not promote a preferential selection of Y-chromosome bearing spermatozoa by the oocyte, and temperature and sex ratio seems to be decoupled in mammals.

Effects of modification of in vitro fertilization techniques on the sex ratio of the resultant bovine embryos.

The duration of sperm-oocyte co-incubation has been observed to affect the sex ratio of in vitro produced bovine embryos. The purpose of this study was to investigate some factors that may be responsible for the skewed sex ratio. The factors studied were selected combinations of the duration of co-incubation, the presence or absence of cumulus cells, and the level of hyaluronic acid (HA) in the culture medium. Experiment 1 examined the effect of selected combinations of different factors during the fertilization phase of in vitro oocyte culture. The factors were the nature of the sperm or its treatment, the duration of the sperm-oocyte co-incubation, and the level of hyaluronic acid in the culture medium. In experiment 2, the capacitation of frozen-thawed-Percoll-washed sperm (control), pre-incubated, and non-binding sperm was evaluated by the zona pellucida (ZP) binding assay and the hypo-osmotic swelling test (HOST). The purpose of experiment 3 was to determine the oocyte cleavage rate and sex ratio of the embryos (>5 cells) produced as a consequence of the 10 treatments used in experiment 1. In treatments 1-3 (experiments 1 and 3) COC were co-cultured with sperm for 1, 5 or 18 h. Polyspermic fertilization rose as the co-incubation period increased (1 h 6.5%, 5 h 15.9%, 18 h 41.8%; P<0.05), and the highest rate of normal fertilization was observed for 5h culture (73.4%; P<0.05). The sex ratio was significantly (P<0.05) skewed from the expected 50:50 towards males following 1 h (64.4%) and 5 h (67.3%) co-incubation, but was not affected by 18 h incubation (52.3%). In treatment 4, sperm was pre-incubated for 1h and cultured with COC for 5 h. Relative to control sperm, pre-incubation of sperm increased ZP binding (116 versus 180 per ZP; P<0.05) and decreased the proportion of HOST positive sperm (65.8-48.6%; P<0.05; experiment 2). Pre-incubation did not affect the rates of polyspermy, normal fertilization or the sex ratio of the embryos (experiments 1 and 3). The oocytes used in treatments 5-10 of experiments 1 and 3 were denuded prior to fertilization. Co-incubation of denuded oocytes for 1h (treatment 5) or 5h (treatment 6) resulted in levels of polyspermic fertilization similar to that for treatment 2 with significantly lower levels of normal fertilization (41.7% and 52.6%, respectively; P<0.05), and the 1h co-incubation significantly skewed (P<0.05) the proportion of male embryos to 70.0%. Denuded oocytes were fertilized for 5h with sperm unable to bind to cumulus cells (NB sperm) in treatment 7 or those that bound to cumulus cells (B) in treatment 8. These two treatments had similar rates of polyspermic, normal and non-fertilization. However, the B sperm caused the sex ratio of the embryos to be significantly skewed to males (63.9%; P<0.05). Fertilization of denuded oocytes in medium containing hyaluronic acid (0.1 mg/ml, treatment 9; 1.0 mg/ml treatment 10) significantly (P<0.05) reduced the incidence of polyspermic fertilization relative to treatments 2 and 6, and normal fertilization relative to treatment 2, but did not affect the sex ratio of the embryos. It was concluded that exposure of sperm to cumulus cells, either before fertilization of denuded oocytes or during the process of fertilization of complete COC, increased the proportion of male embryos produced by in vitro culture. It was hypothesized that this may be due to the capacitation state of the sperm, the cumulus-sperm interaction, and/or the ability of the sperm to bind to cumulus cells or oocytes.

Meiotic drive by the Y-linked D gene in Aedes aegypti (L.) (Diptera: Culicidae) is associated with disruption of spermiogenesis, leading to premature senescence of spermatozoa.

Y chromosome meiotic drive in the mosquito Aedes aegypti, due to the gene D (Distorter) in coupling with M (male determination) [the MD haplotype], is associated with spermiogenic disruption, leading to senescence, at a rate proportionate to male excess. Spermiogenesis was compared between 'Enhanced Mutant' males with a strongly female-depleted sex ratio (8.9% females), 'Mutant' males showing a lesser degree of distortion (38.3% females), and two controls with normal sex ratios (51.2% and 49.2% females). Sections of testes dissected from mature pupae and adults aged 0, 4, 8, 12 and 16 days were examined by transmission electron microscopy. A difference between Mutant and control spermiogenesis was apparent as early as the pupal stage when some Mutant spermatids showed extra tail elements (axonemes and/or mitochondrial derivatives). The same was true of Enhanced Mutant males but to a more extreme degree. Sperm senescence was evident in Enhanced Mutant testes from day 0 of adult life but in Mutant testes not until day 4. Progressive disorganisation was associated with many loose organelles, and disturbance of the anterior-posterior axis of gamete differentiation within the testis. Degenerative changes of a similar kind in the controls did not become apparent until day 8. These findings are discussed with respect to other characteristics of this meiotic drive system, in terms of a theory of inhibition of reduction division in spermatogenesis associated with fragmentation of the X chromosome, leading to the formation of a restitution nucleus as early as metaphase 1.

Sex chromosome-linked genes in plants.

Recent studies of plant sex chromosome-linked genes have revealed many interesting characteristics, although there are limited reports about heteromorphic sex chromosomes in flowering plants. Sex chromosome-linked genes in angiosperms have been characterized mainly in the dioecious plant Silene latifolia. Although all such genes were isolated from transcripts of male flower buds of S. latifolia, most seem to be housekeeping genes except for the petal- and stamen-specific MADS box gene on the Y chromosome (SlAP3Y) and the male reproductive organ-specific gene on the X chromosome (MROS3X). Recent evolutionary studies have revealed at least three evolutionary strata on the X chromosome that are related to stepwise loss of recombination between the sex chromosomes. Moreover, genetic maps showed conservation of gene organization on the X chromosome in the genus Silene and substantial pericentric inversion between the X and Y chromosomes of S. latifolia during evolution. A comparison between paralogs on the sex chromosomes revealed that introns of the Y-linked genes are longer than those of X-linked paralogs. Although analyses of sex chromosome-linked genes suggest that degeneration of the Y chromosome has occurred, the Y chromosome in flowering plants remains the largest in the male genome, unlike that of mammals. Accumulation of repetitive sequences and the entire chloroplast genome on the Y chromosome appear to have contributed to this large size. However, more detailed studies will be required to help explain the basis for the fact that heteromorphic sex chromosomes in angiosperms are large.