Tariq Ezaz

Relationships between Vertebrate ZW and XY Sex Chromosome Systems

The peculiar cytology and unique evolution of sex chromosomes raise many fundamental questions. Why and how sex chromosomes evolved has been debated over a century since H.J. Muller suggested that sex chromosome pairs evolved ultimately from a pair of autosomes. This theory was adapted to explain variations in the snake ZW chromosome pair and later the mammal XY. S. Ohno pointed out similarities between the mammal X and the bird/reptile Z chromosomes forty years ago, but his speculation that they had a common evolutionary origin, or at least evolved from similar regions of the genome, has been undermined by comparative gene mapping, and it is accepted that mammal XY and reptile ZW systems evolved independently from a common ancestor. Here we review evidence for the alternative theory, that ZW<-->XY transitions occurred during evolution, citing examples from fish and amphibians, and probably reptiles. We discuss new work from comparative genomics and cytogenetics that leads to a reconsideration of Ohnos idea and advance a new hypothesis that the mammal XY system may have arisen directly from an ancient reptile ZW system.

The dragon lizard Pogona vitticeps has ZZ/ZW micro-sex chromosomes

The bearded dragon, Pogona vitticeps (Agamidae: Reptilia) is an agamid lizard endemic to Australia. Like crocodilians and many turtles, temperature-dependent sex determination (TSD) is common in agamid lizards, although many species have genotypic sex determination (GSD). P. vitticeps is reported to have GSD, but no detectable sex chromosomes. Here we used molecular cytogenetic and differential banding techniques to reveal sex chromosomes in this species. Comparative genomic hybridization (CGH), GTG- and C-banding identified a highly heterochromatic microchromosome specific to females, demonstrating female heterogamety (ZZ/ZW) in this species. We isolated the P. vitticeps W chromosome by microdissection, re-amplified the DNA and used it to paint the W. No unpaired bivalents were detected in male synaptonemal complexes at meiotic pachytene, confirming male homogamety. We conclude that P. vitticeps has differentiated previously unidentifable W and Z micro-sex chromosomes, the first to be demonstrated in an agamid lizard. Our finding implies that heterochromatinization of the heterogametic chromosome occurred during sex chromosome differentiation in this species, as is the case in some lizards and many snakes, as well as in birds and mammals. Many GSD reptiles with cryptic sex chromosomes may also prove to have micro-sex chromosomes. Reptile microchromosomes, long dismissed as non-functional minutiae and often omitted from karyotypes, therefore deserve closer scrutiny with new and more sensitive techniques.

Sex chromosome evolution in lizards: independent origins and rapid transitions.

Reptiles epitomize the variability of reproductive and sex determining modes and mechanisms among amniotes. These modes include gonochorism (separate sexes) and parthenogenesis, oviparity, viviparity, and ovoviviparity, genotypic sex determination (GSD) with male (XX/XY) and female (ZZ/ZW) heterogamety and temperature-dependent sex determination (TSD). Lizards (order Squamata, suborder Sauria) are particularly fascinating because the distribution of sex-determining mechanisms shows no clear phylogenetic segregation. This implies that there have been multiple transitions between TSD and GSD, and between XY and ZW sex chromosome systems. Approximately 1,000 species of lizards have been karyotyped and among those, fewer than 200 species have sex chromosomes, yet they display remarkable diversity in morphology and degree of degeneration. The high diversity of sex chromosomes as well as the presence of species with TSD, imply multiple and independent origins of sex chromosomes, and suggest that the mechanisms of sex determination are extremely labile in lizards. In this paper, we review the current state of knowledge of sex chromosomes in lizards and the distribution of sex determining mechanisms and sex chromosome forms within and among families. We establish for the first time an association between the occurrence of female heterogamety and TSD within lizard families, and propose mechanisms by which female heterogamety and TSD may have co-evolved. We suggest that lizard sex determination may be much more the result of an interplay between sex chromosomes and temperature than previously thought, such that the sex determination mode is influenced by the nature of heterogamety as well as temperature sensitivity and the stage of sex chromosome degeneration.

Sex reversal triggers the rapid transition from genetic to temperature-dependent sex

Sex determination in animals is amazingly plastic. Vertebrates display contrasting strategies ranging from complete genetic control of sex (genotypic sex determination) to environmentally determined sex (for example, temperature-dependent sex determination). Phylogenetic analyses suggest frequent evolutionary transitions between genotypic and temperature-dependent sex determination in environmentally sensitive lineages, including reptiles. These transitions are thought to involve a genotypic system becoming sensitive to temperature, with sex determined by gene–environment interactions. Most mechanistic models of transitions invoke a role for sex reversal. Sex reversal has not yet been demonstrated in nature for any amniote, although it occurs in fish and rarely in amphibians. Here we make the first report of reptile sex reversal in the wild, in the Australian bearded dragon (Pogona vitticeps), and use sex-reversed animals to experimentally induce a rapid transition from genotypic to temperature-dependent sex determination. Controlled mating of normal males to sex-reversed females produces viable and fertile offspring whose phenotypic sex is determined solely by temperature (temperature-dependent sex determination). The W sex chromosome is eliminated from this lineage in the first generation. The instantaneous creation of a lineage of ZZ temperature-sensitive animals reveals a novel, climate-induced pathway for the rapid transition between genetic and temperature-dependent sex determination, and adds to concern about adaptation to rapid global climate change.

Sex Chromosome Evolution in Amniotes: Applications for Bacterial Artificial Chromosome Libraries

Variability among sex chromosome pairs in amniotes denotes a dynamic history. Since amniotes diverged from a common ancestor, their sex chromosome pairs and, more broadly, sex-determining mechanisms have changed reversibly and frequently. These changes have been studied and characterized through the use of many tools and experimental approaches but perhaps most effectively through applications for bacterial artificial chromosome (BAC) libraries. Individual BAC clones carry 100–200 kb of sequence from one individual of a target species that can be isolated by screening, mapped onto karyotypes, and sequenced. With these techniques, researchers have identified differences and similarities in sex chromosome content and organization across amniotes and have addressed hypotheses regarding the frequency and direction of past changes. Here, we review studies of sex chromosome evolution in amniotes and the ways in which the field of research has been affected by the advent of BAC libraries.

Transitions Between Sex-Determining Systems in Reptiles and Amphibians

Important technological advances in genomics are driving a new understanding of the evolution of sex determination in vertebrates. In particular, comparative chromosome mapping in reptiles has shown an intriguing distribution of homology in sex chromosomes across reptile groups. When this new understanding is combined with the widespread distribution of genetic and temperature-dependent sex-determination mechanisms among reptiles, it is apparent that transitions between modes have occurred many times, as they have for amphibians (particularly between male and female heterogamety). It is also likely that thermosensitivity in sex determination is a key factor in those transitions in reptiles, and possibly in amphibians too. New models of sex determination involving temperature thresholds are providing the framework for the investigation of transitions and making possible key predictions about the homologies and sex-determination patterns expected among taxa in these groups. Molecular cytogenetics and other genomic approaches are essential to providing the fundamental material necessary to make advances in this field.

Evolutionary transitions between mechanisms of sex determination in vertebrates

Sex in many organisms is a dichotomous phenotype—individuals are either male or female. The molecular pathways underlying sex determination are governed by the genetic contribution of parents to the zygote, the environment in which the zygote develops or interaction of the two, depending on the species. Systems in which multiple interacting influences or a continuously varying influence (such as temperature) determines a dichotomous outcome have at least one threshold. We show that when sex is viewed as a threshold trait, evolution in that threshold can permit novel transitions between genotypic and temperature-dependent sex determination (TSD) and remarkably, between male (XX/XY) and female (ZZ/ZW) heterogamety. Transitions are possible without substantive genotypic innovation of novel sex-determining mutations or transpositions, so that the master sex gene and sex chromosome pair can be retained in ZW–XY transitions. We also show that evolution in the threshold can explain all observed patterns in vertebrate TSD, when coupled with evolution in embryonic survivorship limits.

An XX/XY sex microchromosome system in a freshwater turtle, Chelodina longicollis (Testudines: Chelidae) with genetic sex determination.

Heteromorphic sex chromosomes are rare in turtles, having been described in only four species. Like many turtle species, the Australian freshwater turtle Chelodina longicollis has genetic sex determination, but no distinguishable (heteromorphic) sex chromosomes were identified in a previous karyotyping study. We used comparative genomic hybridization (CGH) to show that C. longicollis has an XX/XY system of chromosomal sex determination, involving a pair of microchromosomes. C-banding and reverse fluorescent staining also distinguished microchromosomes with different banding patterns in males and females in ∼70% cells examined. GTG-banding did not reveal any heteromorphic chromosomes, and no replication asynchrony on the X or Y microchromosomes was observed using replication banding. We conclude that there is a very small sequence difference between X and Y chromosomes in this species, a difference that is consistently detectable only by high-resolution molecular cytogenetic techniques, such as CGH. This is the first time a pair of microchromosomes has been identified as the sex chromosomes in a turtle species.

Non-homologous sex chromosomes of birds and snakes share repetitive sequences

Snake sex chromosomes provided Susumo Ohno with the material on which he based his theory of how sex chromosomes differentiate from autosomal pairs. Like birds, snakes have a ZZ male/ZW female sex chromosome system, in which the snake Z is a macrochromosome much the same size as the bird Z. However, the gene content shows clearly that the snake and bird Z chromosomes are completely non-homologous. The molecular aspect of W chromosome degeneration in snakes remains largely unexplored. We used comparative genomic hybridization to identify the female-specific region of the W chromosome in representative species of Australian snakes. Using this approach, we show that an increasingly complex suite of repeats accompanies the evolution of W chromosome heteromorphy. In particular, we found that while the python Liasis fuscus exhibits no sex-specific repeats and indeed, no cytologically recognizable sex-specific region, the colubrid Stegonotus cucullatus shows a large domain on the short arm of the W chromosome that consists of female-specific repeats, and the large W of Notechis scutatus is composed almost entirely of repetitive sequences, including Bkm and 18S rDNA-related elements. FISH mapping of both simple and complex probes shows patterns of repeat amplification concordant with the size of the female-specific region in each species examined. Mapping of intronic sequences of genes that are sex-linked in both birds (DMRT1) and snakes (CTNNB1) reveals massive amplification in discrete domains on the W chromosome of the elapid N. scutatus. Using chicken W chromosome paint, we demonstrate that repetitive sequences are shared between the sex chromosomes of birds and derived snakes. This could be explained by ancestral but as yet undetected shared synteny of bird and snake sex chromosomes or may indicate functional homology of the repeats and suggests that degeneration is a convergent property of sex chromosome evolution. We also establish that synteny of snake Z-linked genes has been conserved for at least 166 million years and that the snake Z consists of two conserved blocks derived from the same ancestral vertebrate chromosome.

Are some chromosomes particularly good at sex? Insights from amniotes

Several recent studies have produced comparative maps of genes on amniote sex chromosomes, revealing homology of gene content and arrangement across lineages as divergent as mammals and lizards. For example, the chicken Z chromosome, which shares homology with the sex chromosomes of all birds, monotremes, and a gecko, is a striking example of stability of genome organization and retention, or independent acquisition, of function in sex determination. In other lineages, such as snakes and therian mammals, well conserved but independently evolved sex chromosome systems have arisen. Among lizards, novel sex chromosomes appear frequently, even in congeneric species. Here, we review recent gene mapping data, examine the evolutionary relationships of amniote sex chromosomes and argue that gene content can predispose some chromosomes to a specialized role in sex determination.