Daniel W. Bellott

Mammalian Y chromosomes retain widely expressed dosage-sensitive regulators

The human X and Y chromosomes evolved from an ordinary pair of autosomes, but millions of years ago genetic decay ravaged the Y chromosome, and only three per cent of its ancestral genes survived. We reconstructed the evolution of the Y chromosome across eight mammals to identify biases in gene content and the selective pressures that preserved the surviving ancestral genes. Our findings indicate that survival was nonrandom, and in two cases, convergent across placental and marsupial mammals. We conclude that the gene content of the Y chromosome became specialized through selection to maintain the ancestral dosage of homologous X–Y gene pairs that function as broadly expressed regulators of transcription, translation and protein stability. We propose that beyond its roles in testis determination and spermatogenesis, the Y chromosome is essential for male viability, and has unappreciated roles in Turner’s syndrome and in phenotypic differences between the sexes in health and disease.

Convergent evolution of chicken Z and human X chromosomes by expansion and gene acquisition

In birds, as in mammals, one pair of chromosomes differs between the sexes. In birds, males are ZZ and females ZW. In mammals, males are XY and females XX. Like the mammalian XY pair, the avian ZW pair is believed to have evolved from autosomes, with most change occurring in the chromosomes found in only one sex—the W and Y chromosomes. By contrast, the sex chromosomes found in both sexes—the Z and X chromosomes—are assumed to have diverged little from their autosomal progenitors. Here we report findings that challenge this assumption for both the chicken Z chromosome and the human X chromosome. The chicken Z chromosome, which we sequenced essentially to completion, is less gene-dense than chicken autosomes but contains a massive tandem array containing hundreds of duplicated genes expressed in testes. A comprehensive comparison of the chicken Z chromosome with the finished sequence of the human X chromosome demonstrates that each evolved independently from different portions of the ancestral genome. Despite this independence, the chicken Z and human X chromosomes share features that distinguish them from autosomes: the acquisition and amplification of testis-expressed genes, and a low gene density resulting from an expansion of intergenic regions. These features were not present on the autosomes from which the Z and X chromosomes originated but were instead acquired during the evolution of Z and X as sex chromosomes. We conclude that the avian Z and mammalian X chromosomes followed convergent evolutionary trajectories, despite their evolving with opposite (female versus male) systems of heterogamety. More broadly, in birds and mammals, sex chromosome evolution involved not only gene loss in sex-specific chromosomes, but also marked expansion and gene acquisition in sex chromosomes common to males and females.

Avian W and mammalian Y chromosomes convergently retained dosage-sensitive regulators

After birds diverged from mammals, different ancestral autosomes evolved into sex chromosomes in each lineage. In birds, females are ZW and males are ZZ, but in mammals females are XX and males are XY. We sequenced the chicken W chromosome, compared its gene content with our reconstruction of the ancestral autosomes, and followed the evolutionary trajectory of ancestral W-linked genes across birds. Avian W chromosomes evolved in parallel with mammalian Y chromosomes, preserving ancestral genes through selection to maintain the dosage of broadly expressed regulators of key cellular processes. We propose that, like the human Y chromosome, the chicken W chromosome is essential for embryonic viability of the heterogametic sex. Unlike other sequenced sex chromosomes, the chicken W chromosome did not acquire and amplify genes specifically expressed in reproductive tissues. We speculate that the pressures that drive the acquisition of reproduction-related genes on sex chromosomes may be specific to the male germ line.

Reconstructing the Evolution of Vertebrate Sex Chromosomes

Sex chromosomes and their evolution have captivated researchers since their discovery. For more than 100 years, the dominant model of sex chromosome evolution has held that differentiated sex chromosomes, such as the X and Y chromosomes of mammals or the Z and W chromosomes of birds, evolved from ordinary autosomes, primarily through the degeneration of the sex-specific Y or W chromosome. At the same time, the sex chromosomes shared between sexes, the X and Z chromosomes, are expected to remain essentially untouched. This model was based on limited cytogenetic and genetic data. Only in the last decade, with the advent of genomics, has the complete sequence of any sex chromosome pair become available. High-quality finished sequences of the human and chimpanzee Y chromosomes, as well as the human X chromosome, have revealed sequence features unanticipated by the traditional model of sex chromosome evolution. Large, highly identical, tandem and inverted arrays of testis-expressed genes are major sources of innovation in gene content on sex-specific chromosomes as well as sex-shared chromosomes. Accounting for the emergence of these ampliconic structures presents a challenge for future studies of sex chromosome evolution.

A widely employed germ cell marker is an ancient disordered protein with reproductive functions in diverse eukaryotes

The advent of sexual reproduction and the evolution of a dedicated germline in multicellular organisms are critical landmarks in eukaryotic evolution. We report an ancient family of GCNA (germ cell nuclear antigen) proteins that arose in the earliest eukaryotes, and feature a rapidly evolving intrinsically disordered region (IDR). Phylogenetic analysis reveals that GCNA proteins emerged before the major eukaryotic lineages diverged; GCNA predates the origin of a dedicated germline by a billion years. Gcna gene expression is enriched in reproductive cells across eukarya – either just prior to or during meiosis in single-celled eukaryotes, and in stem cells and germ cells of diverse multicellular animals. Studies of Gcna-mutant C. elegans and mice indicate that GCNA has functioned in reproduction for at least 600 million years. Homology to IDR-containing proteins implicated in DNA damage repair suggests that GCNA proteins may protect the genomic integrity of cells carrying a heritable genome. DOI: http://dx.doi.org/10.7554/eLife.19993.001

Conserved microRNA targeting reveals preexisting gene dosage sensitivities that shaped amniote sex chromosome evolution

Mammalian X and Y Chromosomes evolved from an ordinary autosomal pair. Genetic decay of the Y led to X Chromosome inactivation (XCI) in females, but some Y-linked genes were retained during the course of sex chromosome evolution, and many X-linked genes did not become subject to XCI. We reconstructed gene-by-gene dosage sensitivities on the ancestral autosomes through phylogenetic analysis of microRNA (miRNA) target sites and compared these preexisting characteristics to the current status of Y-linked and X-linked genes in mammals. Preexisting heterogeneities in dosage sensitivity, manifesting as differences in the extent of miRNA-mediated repression, predicted either the retention of a Y homolog or the acquisition of XCI following Y gene decay. Analogous heterogeneities among avian Z-linked genes predicted either the retention of a W homolog or gene-specific dosage compensation following W gene decay. Genome-wide analyses of human copy number variation indicate that these heterogeneities consisted of sensitivity to both increases and decreases in dosage. We propose a model of XY/ZW evolution incorporating such preexisting dosage sensitivities in determining the evolutionary fates of individual genes. Our findings thus provide a more complete view of the role of dosage sensitivity in shaping the mammalian and avian sex chromosomes and reveal an important role for post-transcriptional regulatory sequences (miRNA target sites) in sex chromosome evolution.

Reconstructing sex chromosome evolution

The mammalian X and Y chromosomes evolved from an ordinary pair of autosomes that existed in a reptilian ancestor that probably relied on temperature-dependent sex determination, as in crocodiles today. Independently and concurrently, the avian Z and W chromosomes (ZZ males, ZW females) evolved from a different pair of autosomes that was present in the same ancestor. Both the mammalian XY pair and the avian ZW pair have emerged with specialized and disproportionate roles in germ cell development. These germ cell specializations are best understood in the mammalian Y chromosomes and are only now being appreciated in the mammalian X chromosomes and the avian ZW pair. To reconstruct and better understand natures sex chromosome experiment, we have set out to comprehensively sequence and compare the sex chromosomes of four primates, two rodents, an ungulate, a marsupial and a bird. I will describe insights that have emerged from this ongoing effort.

Horse Y chromosome assembly displays unique evolutionary features and putative stallion fertility genes

Dynamic evolutionary processes and complex structure make the Y chromosome among the most diverse and least understood regions in mammalian genomes. Here, we present an annotated assembly of the male specific region of the horse Y chromosome (eMSY), representing the first comprehensive Y assembly in odd-toed ungulates. The eMSY comprises single-copy, equine specific multi-copy, PAR transposed, and novel ampliconic sequence classes. The eMSY gene density approaches that of autosomes with the highest number of retained X–Y gametologs recorded in eutherians, in addition to novel Y-born and transposed genes. Horse, donkey and mule testis RNAseq reveals several candidate genes for stallion fertility. A novel testis-expressed XY ampliconic sequence class, ETSTY7, is shared with the parasite Parascaris genome, providing evidence for eukaryotic horizontal transfer and inter-chromosomal mobility. Our study highlights the dynamic nature of the Y and provides a reference sequence for improved understanding of equine male development and fertility.The rapidly evolving Y chromosome accumulates male-benefit genes but is often poorly characterized in many mammals. Here, the authors assemble the male specific region of the horse Y chromosome and investigate its evolution and function.

Cost-effective high-throughput single-haplotype iterative mapping and sequencing for complex genomic structures

The reference sequences of structurally complex regions can be obtained only through highly accurate clone-based approaches. We and others have successfully used single-haplotype iterative mapping and sequencing (SHIMS) 1.0 to assemble structurally complex regions across the sex chromosomes of several vertebrate species and to allow for targeted improvements to the reference sequences of human autosomes. However, SHIMS 1.0 is expensive and time consuming, requiring resources that only a genome center can provide. Here we introduce SHIMS 2.0, an improved SHIMS protocol that allows even a small laboratory to generate high-quality reference sequence from complex genomic regions. Using a streamlined and parallelized library-preparation protocol, and taking advantage of inexpensive high-throughput short-read-sequencing technologies, a small laboratory with both molecular biology and bioinformatics experience can sequence and assemble 192 large-insert bacterial artificial chromosome (BAC) or fosmid clones in 1 week. In SHIMS 2.0, in contrast to other pooling strategies, each clone is sequenced with a unique barcode, thus enabling clones containing nearly identical sequences to be multiplexed in a single sequencing run and assembled separately. Relative to SHIMS 1.0, SHIMS 2.0 decreases the required cost and time by two orders of magnitude while preserving high sequencing accuracy.

Preexisting heterogeneities in gene dosage sensitivity shaped sex chromosome evolution in mammals and birds

Mammalian X and Y chromosomes evolved from an ordinary autosomal pair. Genetic decay of the Y led to X chromosome inactivation (XCI) in females, but some Y-linked genes were retained during the course of sex chromosome evolution, and many X-linked genes did not become subject to XCI. We reconstructed gene-by-gene dosage sensitivities on the ancestral autosomes through phylogenetic analysis of microRNA (miRNA) target sites and compared these preexisting characteristics to the current status of Y-linked and X-linked genes in mammals. Preexisting heterogeneities in dosage sensitivity, manifesting as differences in the extent of miRNA-mediated repression, predicted either the retention of a Y homolog or the acquisition of XCI following Y gene decay. Analogous heterogeneities among avian Z-linked genes predicted the retention of a W homolog. Genome-wide analyses of human copy number variation indicate that these heterogeneities consisted of sensitivity to both increases and decreases in dosage. We propose a model of XY/ZW evolution incorporating such preexisting dosage sensitivities in determining the evolutionary fates of individual genes. Our findings thus provide a more complete view of the role of dosage sensitivity in shaping the mammalian and avian sex chromosomes, and reveal an important role for post-transcriptional regulatory sequences (miRNA target sites) in sex chromosome evolution.Mammalian X and Y chromosomes evolved from an ordinary autosomal pair; genetic decay decimated the Y, which in turn necessitated X chromosome inactivation (XCI). Genes of the ancestral autosomes are often assumed to have undertaken these transitions on uniform terms, but we hypothesized that they varied in their dosage constraints. We inferred such constraints from conservation of microRNA (miRNA)-mediated repression, validated by analysis of experimental data. X-linked genes with a surviving Y homolog have the most conserved miRNA target sites, followed by genes with no Y homolog and subject to XCI, and then genes with no Y homolog but escaping XCI; this heterogeneity existed on the ancestral autosomes. Similar results for avian Z-linked genes, with or without a W homolog, lead to a model of XY/ZW evolution incorporating preexisting dosage sensitivities of individual genes in determining their evolutionary fates, and ultimately shaping the mammalian and avian sex chromosomes.