Evolution of the <Emphasis Type="Italic">DAZ</Emphasis> gene and the <Emphasis Type="Italic">AZFc</Emphasis> region on primate Y chromosomes

Background  

The Azoospermia Factor c (AZFc) region of the human Y chromosome is a unique product of segmental duplication. It consists almost entirely of very long amplicons, represented by different colors, and is frequently deleted in subfertile men. Most of the AZFc amplicons have high sequence similarity with autosomal segments, indicating recent duplication and transposition to the Y chromosome. The Deleted in Azoospermia (DAZ) gene within the red-amplicon arose from an ancestral autosomal DAZ-like (DAZL) gene. It varies significantly between different men regarding to its copy number and the numbers of RNA recognition motif and DAZ repeat it encodes. We used Southern analyses to study the evolution of DAZ and AZFc amplicons on the Y chromosomes of primates.     

Evolution of X-Degenerate Y Chromosome Genes in Greater Apes: Conservation of Gene Content in Human and Gorilla,But Not Chimpanzee

Compared with the X chromosome, the mammalian Y chromosome is considerably diminished in size and has lost most of its ancestral genes during evolution. Interestingly, for the X-degenerate region on the Y chromosome, human has retained all 16 genes, while chimpanzee has lost 4 of the 16 genes since the divergence of the two species. To uncover the evolutionary forces governing ape Y chromosome degeneration, we determined the complete sequences of the coding exons and splice sites for 16 gorilla Y chromosome genes of the X-degenerate region. We discovered that all studied reading frames and splice sites were intact, and thus, this genomic region experienced no gene loss in the gorilla lineage. Higher nucleotide divergence was observed in the chimpanzee than the human lineage, particularly for genes with disruptive mutations, suggesting a lack of functional constraints for these genes in chimpanzee. Surprisingly, our results indicate that the human and gorilla orthologues of the genes disrupted in chimpanzee evolve under relaxed functional constraints and might not be essential. Taking mating patterns and effective population sizes of ape species into account, we conclude that genetic hitchhiking associated with positive selection due to sperm competition might explain the rapid decline in the Y chromosome gene number in chimpanzee. As we found no evidence of positive selection acting on the X-degenerate genes, such selection likely targets other genes on the chimpanzee Y chromosome. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Sex chromosome-to-autosome transposition events counter Y-chromosome gene loss in mammals

BackgroundAlthough the mammalian X and Y chromosomes evolved from a single pair of autosomes, they are highly differentiated: the Y chromosome is dramatically smaller than the X and has lost most of its genes. The surviving genes are a specialized set with extraordinary evolutionary longevity. Most mammalian lineages have experienced delayed, or relatively recent, loss of at least one conserved Y-linked gene. An extreme example of this phenomenon is in the Japanese spiny rat, where the Y chromosome has disappeared altogether. In this species, many Y-linked genes were rescued by transposition to new genomic locations, but until our work presented here, this has been considered an isolated case.ResultsWe describe eight cases of genes that have relocated to autosomes in mammalian lineages where the corresponding Y-linked gene has been lost. These gene transpositions originated from either the X or Y chromosomes, and are observed in diverse mammalian lineages: occurring at least once in marsupials, apes, and cattle, and at least twice in rodents and marmoset. For two genes - EIF1AX/Y and RPS4X/Y - transposition to autosomes occurred independently in three distinct lineages.ConclusionsRescue of Y-linked gene loss through transposition to autosomes has previously been reported for a single isolated rodent species. However, our findings indicate that this compensatory mechanism is widespread among mammalian species. Thus, Y-linked gene loss emerges as an additional driver of gene transposition from the sex chromosomes, a phenomenon thought to be driven primarily by meiotic sex chromosome inactivation.

Electronic supplementary material

The online version of this article (doi:10.1186/s13059-015-0667-4) contains supplementary material, which is available to authorized users.     

De Novo Origin of VCY2 from Autosome to Y-Transposed Amplicon

The formation of new genes is a primary driving force of evolution in all organisms. The de novo evolution of new genes from non-protein-coding genomic regions is emerging as an important additional mechanism for novel gene creation. Y chromosomes underlie sex determination in mammals and contain genes that are required for male-specific functions. In this study, a search was undertaken for Y chromosome de novo genes derived from non-protein-coding sequences. The Y chromosome orphan gene variable charge, Y-linked (VCY)2, is an autosome-derived gene that has sequence similarity to large autosomal fragments but lacks an autosomal protein-coding homolog. VCY2 locates in the amplicon containing long DNA fragments that were transposed from autosomes to the Y chromosome before the ape-monkey split. We confirmed that VCY2cannot be encoded by autosomes due to the presence of multiple disablers that disrupt the open reading frame, such as the absence of start or stop codons and the presence of premature stop codons. Similar observations have been made for homologs in the autosomes of the chimpanzee, gorilla, rhesus macaque, baboon and out-group marmoset, which suggests that there was a non-protein-coding ancestral VCY2 that was common to apes and monkeys that predated the transposition event. Furthermore, while protein-coding orthologs are absent, a putative non-protein-coding VCY2 with conserved disablers was identified in the rhesus macaque Y chromosome male-specific region. This finding implies that VCY2 might have not acquired its protein-coding ability before the ape-monkey split. VCY2 encodes a testis-specific expressed protein and is involved in the pathologic process of male infertility, and the acquisition of this gene might improve male fertility. This is the first evidence that de novo genes can be generated from transposed autosomal non-protein-coding segments, and this evidence provides novel insights into the evolutionary history of the Y chromosome.     

Regional localization of human M-BCR gene to chromosome 23 band q11 in the great apes

We hybridized a human M-BCR DNA probe to the chromosomes of chimpanzee (Pan troglodytes), gorilla (Gorilla gorilld) and orangutan (Pongo pygmaeus) by FISH-technique. The human M-BCR gene was localized to chromosome 23 band q11 (23q11), which is equivalent to the human chromosome 22 band q11 in all three species. The conservation of M-BCR gene in higher primates at the corresponding human chromosome locus provides phylogenetic clues concerning the evolution of genes.     

Y-Chromosome variation in hominids: intraspecific variation is limited to the polygamous chimpanzee

Background

We have previously demonstrated that the Y-specific ampliconic fertility genes DAZ (deleted in azoospermia) and CDY (chromodomain protein Y) varied with respect to copy number and position among chimpanzees (Pan troglodytes). In comparison, seven Y-chromosomal lineages of the bonobo (Pan paniscus), the chimpanzee''s closest living relative, showed no variation. We extend our earlier comparative investigation to include an analysis of the intraspecific variation of these genes in gorillas (Gorilla gorilla) and orangutans (Pongo pygmaeus), and examine the resulting patterns in the light of the species'' markedly different social and mating behaviors.

Methodology/Principal Findings

Fluorescence in situ hybridization analysis (FISH) of DAZ and CDY in 12 Y-chromosomal lineages of western lowland gorilla (G. gorilla gorilla) and a single lineage of the eastern lowland gorilla (G. beringei graueri) showed no variation among lineages. Similar findings were noted for the 10 Y-chromosomal lineages examined in the Bornean orangutan (Pongo pygmaeus), and 11 Y-chromosomal lineages of the Sumatran orangutan (P. abelii). We validated the contrasting DAZ and CDY patterns using quantitative real-time polymerase chain reaction (qPCR) in chimpanzee and bonobo.

Conclusion/Significance

High intraspecific variation in copy number and position of the DAZ and CDY genes is seen only in the chimpanzee. We hypothesize that this is best explained by sperm competition that results in the variant DAZ and CDY haplotypes detected in this species. In contrast, bonobos, gorillas and orangutans—species that are not subject to sperm competition—showed no intraspecific variation in DAZ and CDY suggesting that monoandry in gorillas, and preferential female mate choice in bonobos and orangutans, probably permitted the fixation of a single Y variant in each taxon. These data support the notion that the evolutionary history of a primate Y chromosome is not simply encrypted in its DNA sequences, but is also shaped by the social and behavioral circumstances under which the specific species has evolved.     

Y Chromosomal Variation Tracks the Evolution of Mating Systems in Chimpanzee and Bonobo

The male-specific regions of the Y chromosome (MSY) of the human and the chimpanzee (Pan troglodytes) are fully sequenced. The most striking difference is the dramatic rearrangement of large parts of their respective MSYs. These non-recombining regions include ampliconic gene families that are known to be important for male reproduction,and are consequently under significant selective pressure. However, whether the published Y-chromosomal pattern of ampliconic fertility genes is invariable within P. troglodytes is an open but fundamental question pertinent to discussions of the evolutionary fate of the Y chromosome in different primate mating systems. To solve this question we applied fluorescence in situ hybridisation (FISH) of testis-specific expressed ampliconic fertility genes to metaphase Y chromosomes of 17 chimpanzees derived from 11 wild-born males and 16 bonobos representing seven wild-born males. We show that of eleven P. troglodytes Y-chromosomal lines, ten Y-chromosomal variants were detected based on the number and arrangement of the ampliconic fertility genes DAZ (deleted in azoospermia) and CDY (chromodomain protein Y)—a so-far never-described variation of a species'' Y chromosome. In marked contrast, no variation was evident among seven Y-chromosomal lines of the bonobo, P. paniscus, the chimpanzee''s closest living relative. Although, loss of variation of the Y chromosome in the bonobo by a founder effect or genetic drift cannot be excluded, these contrasting patterns might be explained in the context of the species'' markedly different social and mating behaviour. In chimpanzees, multiple males copulate with a receptive female during a short period of visible anogenital swelling, and this may place significant selection on fertility genes. In bonobos, however, female mate choice may make sperm competition redundant (leading to monomorphism of fertility genes), since ovulation in this species is concealed by the prolonged anogenital swelling, and because female bonobos can occupy high-ranking positions in the group and are thus able to determine mate choice more freely.     

Recent duplication and positive selection of the <Emphasis Type="Italic">GAGE</Emphasis> gene family

We report that the GAGE gene family of human Cancer/testis antigen (CTA) genes is likely to be in an early stage of its evolution. Members of this gene family are tandemly arranged on the X chromosome only in human, chimpanzee and macaque genomes and share a very high similarity. Phylogenetic trees show that the GAGE gene family began to duplicate after the split of human and chimpanzee. The estimated ages of the duplication events range from 4 million years ago to the present. The Ka/Ks values between the duplicates are significantly greater than 1, indicating that the mutation rate is higher in coding regions than non-coding regions of the genes, which suggests that the GAGE gene family is under positive selection. These findings indicate that the GAGE gene family may be a newly formed gene family undergoing rapid functional evolution. Yang Liu and Qiyun Zhu contributed equally to this work.     

AZFc somatic microdeletions and copy number polymorphism of the DAZ genes in human males exposed to natural background radiation

Ionizing radiations are known to induce tumors, chromosomal lesions and minisatellite length variations, yet no correlation has been demonstrated between radiation exposure and indels or copy number polymorphism (CNP) of the genes. We studied the impact of natural background radiation (NBR) on the human Y chromosome owing to its haploid status and clonal inheritance. We analyzed the AZFc region using the DNA from blood and semen of 100 males living near the coastal peninsula in Kerala (India), exposed to NBR along with other 50 normal fertile males. STS mapping of AZFc region showed random microdeletions without conclusive gr/gr or b1/b3 phenotypes. Using a highly specific novel Taqman assay based on sY587 sequence, we detected four copies of the DAZ genes in normal males and 4–16 in those exposed to NBR. Amongst NBR exposed males with multiples copies of the DAZ genes, 75% showed varying FISH signals for DAZ genes with cosmid 18E8 whereas 30% showed mosaicism in terms of presence/absence of the signals in 6–8% cells and unexpected number of signals in 9–12% interphase nuclei. Startlingly, all germline samples studied were found to be free from AZFc microdeletions and CNP of the DAZ genes. Since the DAZ genes are heavily implicated with the germ cell development, the cells with DAZ deletion/duplication are unlikely to survive. Alternatively, an innate mechanism may be operative to protect the germline from the effects of NBR.     

Complex Evolution of a Y-Chromosomal Double Homeobox 4 (DUX4)-Related Gene Family in Hominoids

The human Y chromosome carries four human Y-chromosomal euchromatin/heterochromatin transition regions, all of which are characterized by the presence of interchromosomal segmental duplications. The Yq11.1/Yq11.21 transition region harbours a peculiar segment composed of an imperfectly organized tandem-repeat structure encoding four members of the double homeobox (DUX) gene family. By comparative fluorescence in situ hybridization (FISH) analysis we have documented the primary appearance of Y-chromosomal DUX genes (DUXY) on the gibbon Y chromosome. The major amplification and dispersal of DUXY paralogs occurred after the gibbon and hominid lineages had diverged. Orthologous DUXY loci of human and chimpanzee show a highly similar structural organization. Sequence alignment survey, phylogenetic reconstruction and recombination detection analyses of human and chimpanzee DUXY genes revealed the existence of all copies in a common ancestor. Comparative analysis of the circumjacent beta-satellites indicated that DUXY genes and beta-satellites evolved in concert. However, evolutionary forces acting on DUXY genes may have induced amino acid sequence differences in the orthologous chimpanzee and human DUXY open reading frames (ORFs). The acquisition of complete ORFs in human copies might relate to evolutionary advantageous functions indicating neo-functionalization. We propose an evolutionary scenario in which an ancestral tandem array DUX gene cassette transposed to the hominoid Y chromosome followed by lineage-specific chromosomal rearrangements paved the way for a species-specific evolution of the Y-chromosomal members of a large highly diverged homeobox gene family.     

Evolutionary breakpoint analysis on Y chromosomes of higher primates provides insight into human Y evolution

Comparative FISH mapping of PAC clones covering almost 3 Mb of the human AZFa region in Yq11.21 to metaphases of human and great apes unravels breakpoints that were involved in species-specific Y chromosome evolution. An astonishing clustering of evolutionary breakpoints was detected in the very proximal region on the long arm of the human Y chromosome in Yq11.21. These breakpoints were involved in deletions, one specific for the human and another for the orang-utan Y chromosome, in a duplicative translocation/transposition specific for bonobo and chimpanzee Y chromosomes and in a pericentric inversion specific for the gorilla Y chromosome. In addition, our comparative results allow the deduction of a model for the human Y chromosome evolution.     

Evolutionary divergence of human chromosome 9 as revealed by the position of the ABL protooncogene in higher primates

Attempts to solve the fundamental questions regarding the descent of man are dogged by superstitions and unexamined orthodoxies. The origin of humans, established a decade ago based upon cytological analysis of ape chromosomes, continues to be called into question. Although molecular methods have provided a framework for tracing the paths of human evolution, conclusive evidence remains elusive. We have used a single ABL gene probe derived from human chromosome 9 to assess the direction of change in the equivalent ape chromosomes. This approach has resulted in a few surprises which again challenge the prevailing view of early primate evolution based solely on chromosome banding patterns. The ABL protooncogene is present on human chromosome 9 at band q34. Similar DNA sequences presumed to represent an ABL gene, are present on chromosome 11 in chimpanzee (Pan troglodytes) but at a different relative location, indicating that the mechanism of the origin of human chromosome 9 is far more complex than has previously been suggested. Nevertheless, in gorilla (Gorilla gorilla) and orangutan (Pongo pygmaeus), the equivalent to human chromosome band 9 q34 is apparently located on chromosome 13 at a putative telomeric position and no discernible differences could be established. Despite the presence of the ABL protooncogene on human equivalent ape chromosomes, molecular systematics will continue to generate enigmas in the evolutionary context until the entire genome is sequenced.     

Evolutionary comparison of the reproductive genes, DAZL and BOULE, in primates with and without DAZ

Genes of the DAZ (Deleted in AZoospermia) gene family, DAZ, DAZL (DAZ-Like), and BOULE, encode closely related RNA-binding proteins that are required for fertility in numerous organisms, yet the genomes of different organisms possess different complements of DAZ family genes. Thus, invertebrates such as flies and worms contain just a single DAZ homolog, boule, while genomes of vertebrates, other than catarrhine primates (Old World monkeys and hominids), possess both Boule and Dazl genes. Finally, catarrhine primates possess BOULE, DAZL, and DAZ genes. Since the DAZ genes arose recently in evolution in the catarrhine lineage, we sought to examine how the sequences and expression of this gene family may have changed after the introduction of a new member, DAZ. Based on previous results, we hypothesized that the introduction of a new member of the DAZ gene family into catarrhines could reduce functional constraint on DAZL. Surprisingly, however, we found that platyrrhine DAZL demonstrated significantly more sequence divergence than catarrhine DAZL (p=0.0006 for nucleotide and p=0.05 for amino acid sequence); however, comparison of K _a/K _s ratios suggests that the DAZL and BOULE genes are under similar functional constraints regardless of lineage. Thus, our data are most consistent with the hypothesis that the introduction of DAZ did not affect the evolution of DAZL or BOULE, and that a higher neutral mutation rate in platyrrhines than in catarrhines, along with the greater tolerance of DAZL for variation relative to BOULE, may be the foundation for the observed differences in sequence divergence in this gene family.     

Novel gene acquisition on carnivore Y chromosomes

Despite its importance in harboring genes critical for spermatogenesis and male-specific functions, the Y chromosome has been largely excluded as a priority in recent mammalian genome sequencing projects. Only the human and chimpanzee Y chromosomes have been well characterized at the sequence level. This is primarily due to the presumed low overall gene content and highly repetitive nature of the Y chromosome and the ensuing difficulties using a shotgun sequence approach for assembly. Here we used direct cDNA selection to isolate and evaluate the extent of novel Y chromosome gene acquisition in the genome of the domestic cat, a species from a different mammalian superorder than human, chimpanzee, and mouse (currently being sequenced). We discovered four novel Y chromosome genes that do not have functional copies in the finished human male-specific region of the Y or on other mammalian Y chromosomes explored thus far. Two genes are derived from putative autosomal progenitors, and the other two have X chromosome homologs from different evolutionary strata. All four genes were shown to be multicopy and expressed predominantly or exclusively in testes, suggesting that their duplication and specialization for testis function were selected for because they enhance spermatogenesis. Two of these genes have testis-expressed, Y-borne copies in the dog genome as well. The absence of the four newly described genes on other characterized mammalian Y chromosomes demonstrates the gene novelty on this chromosome between mammalian orders, suggesting it harbors many lineage-specific genes that may go undetected by traditional comparative genomic approaches. Specific plans to identify the male-specific genes encoded in the Y chromosome of mammals should be a priority.     

Gene-Level,but Not Chromosome-Wide,Divergence between a Very Young House Fly Proto-Y Chromosome and Its Homologous Proto-X Chromosome

X and Y chromosomes are usually derived from a pair of homologous autosomes, which then diverge from each other over time. Although Y-specific features have been characterized in sex chromosomes of various ages, the earliest stages of Y chromosome evolution remain elusive. In particular, we do not know whether early stages of Y chromosome evolution consist of changes to individual genes or happen via chromosome-scale divergence from the X. To address this question, we quantified divergence between young proto-X and proto-Y chromosomes in the house fly, Musca domestica. We compared proto-sex chromosome sequence and gene expression between genotypic (XY) and sex-reversed (XX) males. We find evidence for sequence divergence between genes on the proto-X and proto-Y, including five genes with mitochondrial functions. There is also an excess of genes with divergent expression between the proto-X and proto-Y, but the number of genes is small. This suggests that individual proto-Y genes, but not the entire proto-Y chromosome, have diverged from the proto-X. We identified one gene, encoding an axonemal dynein assembly factor (which functions in sperm motility), that has higher expression in XY males than XX males because of a disproportionate contribution of the proto-Y allele to gene expression. The upregulation of the proto-Y allele may be favored in males because of this gene’s function in spermatogenesis. The evolutionary divergence between proto-X and proto-Y copies of this gene, as well as the mitochondrial genes, is consistent with selection in males affecting the evolution of individual genes during early Y chromosome evolution.     

Evolution of homologous sequences on the human X and Y chromosomes, outside of the meiotic pairing segment.

A sequence isolated from the long arm of the human Y chromosome detects a highly homologous locus on the X. This homology extends over at least 50 kb of DNA and is postulated to be the result of a transposition event between the X and Y chromosomes during recent human evolution, since homologous sequences are shown to be present on the X chromosome alone in the chimpanzee and gorilla.     

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.     

Selection of DNA sequences from interval 6 of the human Y chromosome with homology to a Y chromosomal fertility gene sequence of Drosophila hydei

Summary An experimental approach towards the molecular analysis of the male fertility function, located in interval 6 of the human Y chromosome, is presented. This approach is not based on the knowledge of any gene product but on the assumption that the functional DNA structure of male fertility genes, evolutionary conserved with their position on the Y chromosome, may contain an evolutionary conserved frame structure or at least conserved sequence elements. We tested this hypothesis by using dhMiF1, a fertility gene sequence of the Y chromosome of Drosophila hydei, as a screening probe on a pool of cloned human Y-DNA sequences. We were able to select 10 human Y-DNA sequences of which 7 could be mapped to Y interval 6 (the pY6H sequence family). Since the only fertility gene of the human Y chromosome is mapped to the same Y interval, our working hypothesis seems to be strongly supported. Most interesting in this respect is the isolation of the Y-specific repetitive pY6H65 sequence. The pY6H65 locus extends to a length of at least 300 kb in Y interval 6 and has a locus-specific repetitive sequence organization, reminiscent of the functional DNA structure of Y chromosomal fertility genes of Drosophila. We identified the simple sequence family (CA)_n as one sequence element conserved between the Drosophila dhMiFi fertility gene sequence and the homologous human Y-DNA sequences.     

The chimpanzee GH locus: composition, organization, and evolution

In most mammals the growth hormone (GH) locus comprises a single gene expressed primarily in the anterior pituitary gland. However, in higher primates multiple duplications of the GH gene gave rise to a complex locus containing several genes. In man this locus comprises five genes, including GH-N (expressed in pituitary) and four genes expressed in the placenta, but in other species the number and organization of these genes vary. The situation in chimpanzee has been unclear, with suggestions of up to seven GH-like genes. We have re-examined the GH locus in chimpanzee and have deduced the complete sequence. The locus includes five genes apparently organized in a fashion similar to that in human, with two of these genes encoding GH-like proteins, and three encoding chorionic somatomammotropins/placental lactogens (CSHs/PLs). There are notable differences between the human and chimpanzee loci with regard to the expressed proteins, gene regulation, and gene conversion events. In particular, one human gene (hCSH-L) has changed substantially since the chimpanzee/human split, potentially becoming a pseudogene, while the corresponding chimpanzee gene (CSH-A1) has been conserved, giving a product almost identical to the adjacent CSH-A2. Chimpanzee appears to produce two CSHs, with potentially differing biological properties, whereas human produces a single CSH. The pattern of gene conversion in human has been quite different from that in chimpanzee. The region around the GH-N gene in chimpanzee is remarkably polymorphic, unlike the corresponding region in human. The results shed new light on the complex evolution of the GH locus in higher primates.