Mammalian genome evolution: new clues from comparisons of eutherians, marsupials and monotremes.

1. Comparisons of chromosomes and gene maps of different mammals are yielding a big picture of the evolution of mammalian genome form and function. It has been particularly instructive to compare gene arrangements on the sex chromosomes between the three major groups of mammals. Eutheria (so-called placental mammals). Metatheria (marsupials) and Prototheria (monotremes), which diverged 150 and 170 Myr BP respectively. 2. A region amounting to 3% of the haploid genome is located on the X chromosome in all three groups, implying that this region must have been part of the original X in a common ancestor. This region comprises the long arm of the human X. 3. A region represented by the short arm of the human X is common to the X in all eutherians, but is autosomal in marsupials and monotremes; thus it was not a part of the original X, and must have been acquired by the X early in the eutherian radiation. 4. This recently acquired region was probably translocated to a pseudoautosomal region shared by the eutherian X and Y. Thus it was originally paired and exempt from X chromosome inactivation; stepwise deletion of this region from the Y and recruitment of the newly unpaired region of the X into the inactivation system could account for some of the peculiarities of this region of the human X. 5. The sex-determining gene TDF must lie on the Y in all mammals in which the Y is male determining. The autosomal location of the candidate gene ZFY in marsupials and monotremes eliminates it from consideration. The recently described candidate gene SRY has yet to pass the "marsupial test".     

Independent Evolution of Transcriptional Inactivation on Sex Chromosomes in Birds and Mammals

X chromosome inactivation in eutherian mammals has been thought to be tightly controlled, as expected from a mechanism that compensates for the different dosage of X-borne genes in XX females and XY males. However, many X genes escape inactivation in humans, inactivation of the X in marsupials is partial, and the unrelated sex chromosomes of monotreme mammals have incomplete and gene-specific inactivation of X-linked genes. The bird ZW sex chromosome system represents a third independently evolved amniote sex chromosome system with dosage compensation, albeit partial and gene-specific, via an unknown mechanism (i.e. upregulation of the single Z in females, down regulation of one or both Zs in males, or a combination). We used RNA-fluorescent in situ hybridization (RNA-FISH) to demonstrate, on individual fibroblast cells, inactivation of 11 genes on the chicken Z and 28 genes on the X chromosomes of platypus. Each gene displayed a reproducible frequency of 1Z/1X-active and 2Z/2X-active cells in the homogametic sex. Our results indicate that the probability of inactivation is controlled on a gene-by-gene basis (or small domains) on the chicken Z and platypus X chromosomes. This regulatory mechanism must have been exapted independently to the non-homologous sex chromosomes in birds and mammals in response to an over-expressed Z or X in the homogametic sex, highlighting the universal importance that (at least partial) silencing plays in the evolution on amniote dosage compensation and, therefore, the differentiation of sex chromosomes.     

Cytogenetic evidence for the absence of an inactivated X chromosome in a human female (XX) breast carcinoma cell line

Summary Barr body staining procedures were applied directly to the chamber slide cultures of female amniotic cells, WI38 fibroblasts, normal female kidney cells, and a human breast carcinoma cell line, Elco. A high frequency of Barr bodies was found in all the normal female control cells; however, no Barr bodies were observed in the Elco cells. By trypsin G-banding analysis, two normal X chromosomes were identified in all Elco cells. The late DNA replication pattern of the cell line was then studied with the terminal BrdU pulse method. Both X chromosomes in the Elco cell line were found to be euchromatic with a characteristic R-banding pattern; no late-replicating X chromosome was observed. The absence of both a Barr body and a late-replicating heterocyclic X chromosome provides strong cytogenetic evidence that an inactivated X chromosome is absent in the human breast carcinoma cells bearing two X chromosomes.     

Chromosomal instability in Afrotheria: fragile sites,evolutionary breakpoints and phylogenetic inference from genome sequence assemblies

Background  

Extant placental mammals are divided into four major clades (Laurasiatheria, Supraprimates, Xenarthra and Afrotheria). Given that Afrotheria is generally thought to root the eutherian tree in phylogenetic analysis of large nuclear gene data sets, the study of the organization of the genomes of afrotherian species provides new insights into the dynamics of mammalian chromosomal evolution. Here we test if there are chromosomal bands with a high tendency to break and reorganize in Afrotheria, and by analyzing the expression of aphidicolin-induced common fragile sites in three afrotherian species, whether these are coincidental with recognized evolutionary breakpoints.     

Haldane's rule in marsupials: what happens when both sexes are functionally hemizygous?

During the process of speciation, diverging taxa often hybridize and produce offspring wherein the heterogametic sex (i.e., XY or ZW) is unfit (Haldane's rule). Dominance theory seeks to explain Haldane's rule in terms of the difference in X-linked dominance regimes experienced by the sexes. However, X inactivation in female mammals extends the effects of hemizygosity to both sexes. Here, we highlight where the assumptions of dominance theory are particularly problematic in marsupials, where X inactivation uniformly results in silencing the paternal X. We then present evidence of Haldane's rule for sterility but not for viability in marsupials, as well as the first violations of Haldane's rule for these traits among all mammals. Marsupials represent a large taxonomic group possessing heteromorphic sex chromosomes, where the dominance theory cannot explain Haldane's rule. In this light, we evaluate alternative explanations for the preponderance of male sterility in interspecific hybrids, including faster male evolution, X-Y interactions, and genomic conflict hypotheses.     

Chromatin configuration and epigenetic landscape at the sex chromosome bivalent during equine spermatogenesis

Pairing of the sex chromosomes during mammalian meiosis is characterized by the formation of a unique heterochromatin structure at the XY body. The mechanisms underlying the formation of this nuclear domain are reportedly highly conserved from marsupials to mammals. In this study, we demonstrate that in contrast to all eutherian species studied to date, partial synapsis of the heterologous sex chromosomes during pachytene stage in the horse is not associated with the formation of a typical macrochromatin domain at the XY body. While phosphorylated histone H2AX (γH2AX) and macroH2A1.2 are present as a diffuse signal over the entire macrochromatin domain in mouse pachytene spermatocytes, γH2AX, macroH2A1.2, and the cohesin subunit SMC3 are preferentially enriched at meiotic sex chromosome cores in equine spermatocytes. Moreover, although several histone modifications associated with this nuclear domain in the mouse such as H3K4me2 and ubH2A are conspicuously absent in the equine XY body, prominent RNA polymerase II foci persist at the sex chromosomes. Thus, the localization of key marker proteins and histone modifications associated with the XY body in the horse differs significantly from all other mammalian systems described. These results demonstrate that the epigenetic landscape and heterochromatinization of the equine XY body might be regulated by alternative mechanisms and that some features of XY body formation may be evolutionary divergent in the domestic horse. We propose equine spermatogenesis as a unique model system for the study of the regulatory networks leading to the epigenetic control of gene expression during XY body formation.     

哺乳动物性别分化的调控

哺乳动物性别分化调控的分子机制的研究特别是性别分化的层次调控、剂量补偿和性染色体进化这三个领域,已取得快速进展。已经发现Ｙ染色体性别决定区基因（ＳＲＹ）、Ｘ染色体ＤＳＳ－ＡＨＣ决定区基因１（ＤＡＸ－１）、甾类生成因子１基因（ＳＦ１）和Ｗｉｌｍｓ瘤抑制基因（ＷＴ－１）等与哺乳动物性别决定有关。ＳＲＹ启动睾丸分化,但胚胎发育成雄性的其余步骤由事丸分泌的激素控制。ＤＡＸ－１且编码一种女性特异功能的蛋白质,它在男性中被ＳＲＹ所抑制。ＳＦ－１和ＷＴ－１在ＳＲＹ开启之前作用于性腺和肾上腺发育的启动。哺乳动物通过随机失活雌性两条Ｘ染色体中的一条来使Ｘ连锁的基因在两性间的表达水平达到平衡（剂量补偿）。Ｘ染色体失活由Ｘ染色体失活中心（ＸＩＣ）控制。失活的Ｘ染色体专一转录基因（ＸＩＳＴ）是ＸＩＣ的强烈候选者,它可能参与Ｘ失活的启动。对有袋目和单孔目动物性染色体的研究为我们提供了其进化的信息。有证据支持性染色体起源于一对同源常染色体,而ＳＲＹ的祖先基因可能是ＳＯＸ－３。     

CSF2RA, ANT3, and STS are autosomal in marsupials: implications for the origin of the pseudoautosomal region of mammalian sex chromosomes

The X and Y Chromosomes (Chrs) of eutherian (``placental') mammals share a pseudo-autosomal region (PAR) that pairs and recombines at meiosis. In humans and other eutherians, the PAR contains several active genes and has also been thought to be critical for pairing and fertility. In order to explore the origin of the PAR, we cloned and mapped three human or mouse pseudoautosomal genes in marsupials, a group of mammals that diverged from eutherians about 130 (MYrBP). All three genes were autosomal in marsupials, and two co-localized with other human Xp genes on an autosome. This implies that the human PAR, like most of human Xp, represents a relic of an autosomal region added to both X and Y Chrs between 80 and 150 MYrBP. Received: 19 September 1997 / Accepted: 20 January 1998     

The meiotic behaviour of the XY pair in Lutreolina crassicaudata (Marsupialia: Didelphoidea)

The meiotic behaviour of the XY pair of the didelphid Lutreolina crassicaudata is analyzed by microspreading of spermatocytes for visualization of chromosomal axes and by three-dimensional reconstruction of spermatocyte nuclei from EM thin sections. The delay in pairing of sex chromosomes compared to autosomes and the absence of a synaptonemal complex between the axes of the X and Y chromosomes, already described for South American marsupials by three-dimensional reconstruction and for Australian species with synaptonemal complex microspreadings, is confirmed for this species. Sections demonstrate that at the diffuse stage and diplotene the dense plate occupies the region of the inner face of the nuclear envelope in contact with the XY body. Spreads show an structure similar in staining to the axes that becomes apparent simultaneously with the dense plate, called a balloon. The mechanism of XY pairing during meiotic prophase appears to be common to American and Australian marsupials as the same morphological pattern is found in all the species described. This mechanism is different from the way of pairing and segregation known for eutherian sex chromosomes.     

Analysis of male meiotic "sex body" proteins during XY female meiosis provides new insights into their functions

During male meiosis in mammals the X and Y chromosomes become condensed to form the sex body (XY body), which is the morphological manifestation of the process of meiotic sex chromosome inactivation (MSCI). An increasing number of sex body located proteins are being identified, but their functions in relation to MSCI are unclear. Here we demonstrate that assaying male sex body located proteins during XY female mouse meiosis, where MSCI does not take place, is one way in which to begin to discriminate between potential functions. We show that a newly identified protein, "Asynaptin" (ASY), detected in male meiosis exclusively in association with the X and Y chromatin of the sex body, is also expressed in pachytene oocytes of XY females where it coats the chromatin of the asynapsed X in the absence of MSCI. Furthermore, in pachytene oocytes of females carrying a reciprocal autosomal translocation, ASY associates with asynapsed autosomal chromatin. Thus the location of ASY to the sex body during male meiosis is likely to be a response to the asynapsis of the non-homologous regions [outside the pseudoautosomal region (PAR)] of the heteromorphic X-Y bivalent, rather than being related to MSCI. In contrast to ASY, the previously described sex body protein XY77 proved to be male sex body specific. Potential functions for MSCI and the sex body are discussed together with the possible roles of these two proteins.     

The tricky path to recombining X and Y chromosomes in meiosis

Sex chromosomes are the Achilles' heel of male meiosis in mammals. Mis-segregation of the X and Y chromosomes leads to sex chromosome aneuploidies, with clinical outcomes such as infertility and Klinefelter syndrome. Successful meiotic divisions require that all chromosomes find their homologous partner and achieve recombination and pairing. Sex chromosomes in males of many species have only a small region of homology (the pseudoautosomal region, PAR) that enables pairing. Until recently, little was known about the dynamics of recombination and pairing within mammalian X and Y PARs. Here, we review our recent findings on PAR behavior in mouse meiosis. We uncovered unexpected differences between autosomal chromosomes and the X-Y chromosome pair, namely that PAR recombination and pairing occurs later, and is under different genetic control. These findings imply that spermatocytes have evolved distinct strategies that ensure successful X-Y recombination and chromosome segregation.     

Weird mammals provide insights into the evolution of mammalian sex chromosomes and dosage compensation

The deep divergence of mammalian groups 166 and 190 million years ago (MYA) provide genetic variation to explore the evolution of DNA sequence, gene arrangement and regulation of gene expression in mammals. With encouragement from the founder of the field, Mary Lyon, techniques in cytogenetics and molecular biology were progressively adapted to characterize the sex chromosomes of kangaroos and other marsupials, platypus and echidna—and weird rodent species. Comparative gene mapping reveals the process of sex chromosome evolution from their inception 190 MYA (they are autosomal in platypus) to their inevitable end (the Y has disappeared in two rodent lineages). Our X and Y are relatively young, getting their start with the evolution of the sex-determining SRY gene, which triggered progressive degradation of the Y chromosome. Even more recently, sex chromosomes of placental mammals fused with an autosomal region which now makes up most of the Y. Exploration of gene activity patterns over four decades showed that dosage compensation via X-chromosome inactivation is unique to therian mammals, and that this whole chromosome control process is different in marsupials and absent in monotremes and reptiles, and birds. These differences can be exploited to deduce how mammalian sex chromosomes and epigenetic silencing evolved.     

The genes from the pseudoautosomal region 1 (PAR1) of the mammalian sex chromosomes: Synteny,phylogeny and selection

Sex chromosomes recombine restrictly in their homologous area, the pseudoautosomal region (PAR), represented by PAR1 and PAR2, which behave like an autosome in both pairing and recombination. The PAR1, common to most of the eutherian mammals, is located at the terminus of the sex chromosomes short arm and exhibit recombination rates ~20 times higher than the autosomes. Here, we assessed the interspecific evolutionary genomic dynamics of 15 genes of the PAR1 across 41 mammalian genera (representing six orders). The strong negative selection detected in most of the assessed groups reinforces the presence of evolutionary constraints, imposed by the important function of the PAR1 genes. Indeed, mutations in these genes are associated with various diseases in humans, including stature problems (Klinefelter Syndrome), leukemia and mental diseases. Yet, a few genes exhibiting positive selection (ω-value >1) were depicted in Rodentia (ASMT and ZBED1) and Primates (CRLF2 and CSF2RA). Rodents have the smallest described PAR1, while that of simian primates/humans underwent a 3 to 5 fold size reduction. The assessment of the PAR1 genes synteny revealed differences among the mammalian species, especially in the Rodentia order where chromosomic translocations from the sex chromosomes to the autosomes were observed. Such syntenic changes may be an evidence of the rapid evolution in rodents, as previous referred in other papers, also depicted by their increased branch lengths in the phylogenetic analyses. Concluding, we suggest that genome migration is an important factor influencing the evolution of mammals and may result in changes of the selective pressures operating on the genome.     

Meiotic sex chromosome inactivation in the marsupial Monodelphis domestica

In eutherian mammals, the X and Y chromosomes undergo meiotic sex chromosome inactivation (MSCI) during spermatogenesis in males. However, following fertilization, both the paternally (Xp) and maternally (Xm) inherited X chromosomes are active in the inner cell mass of the female blastocyst, and then random inactivation of one X chromosome occurs in each cell, leading to a mosaic pattern of X-chromosome activity in adult female tissues. In contrast, marsupial females show a nonrandom pattern of X chromosome activity, with repression of the Xp in all somatic tissues. Here, we show that MSCI also occurs during spermatogenesis in marsupials in a manner similar to, but more stable than that in eutherians. These findings support the suggestion that MSCI may have provided the basis for an early dosage compensation mechanism in mammals based solely on gametogenic events, and that random X-chromosome inactivation during embryogenesis may have evolved subsequently in eutherian mammals.     

Sex chromosome homology and incomplete, tissue-specific X-inactivation suggest that monotremes represent an intermediate stage of mammalian sex chromosome evolution

Female mammals have two X chromosomes and males have a single X and a smaller, male-determining Y chromosome. The dosage of X-linked gene products is equalized between the sexes by the genetic inactivation of one X chromosome in females. The characteristics of the mechanism of X-chromosome inactivation differ in eutherian and metatherian mammals, and it has been suggested that the metatherian system represents a more primitive stage. The present study of monotreme sex chromosomes and X-chromosome inactivation suggests that the prototherian mammals may represent an even more primitive stage. There is extensive G-band homology between the monotreme X and Y chromosomes, and differences in the patterns of replication of the two X chromosomes in females suggest that X inactivation is tissue specific and confined to the unpaired segment of the X. On the basis of these results, we propose a model for the differentiation of mammalian sex chromosomes and the evolution of the mechanism of X-chromosome inactivation. This model involves a gradual reduction of the Y chromosome and an accompanying gradual recruitment of (newly unpaired) X-linked loci under the control of a single inactivation center.     

Evolutionary dynamics of the human pseudoautosomal regions

Recombination between the X and Y human sex chromosomes is limited to the two pseudoautosomal regions (PARs) that present quite distinct evolutionary origins. Despite the crucial importance for male meiosis, genetic diversity patterns and evolutionary dynamics of these regions are poorly understood. In the present study, we analyzed and compared the genetic diversity of the PAR regions using publicly available genomic sequences encompassing both PAR1 and PAR2. Comparisons were performed through allele diversities, linkage disequilibrium status and recombination frequencies within and between X and Y chromosomes. In agreement with previous studies, we confirmed the role of PAR1 as a male-specific recombination hotspot, but also observed similar characteristic patterns of diversity in both regions although male recombination occurs at PAR2 to a much lower extent (at least one recombination event at PAR1 and in ≈1% in normal male meioses at PAR2). Furthermore, we demonstrate that both PARs harbor significantly different allele frequencies between X and Y chromosomes, which could support that recombination is not sufficient to homogenize the pseudoautosomal gene pool or is counterbalanced by other evolutionary forces. Nevertheless, the observed patterns of diversity are not entirely explainable by sexually antagonistic selection. A better understanding of such processes requires new data from intergenerational transmission studies of PARs, which would be decisive on the elucidation of PARs evolution and their role in male-driven heterosomal aneuploidies.