Sex determination: the amphibian models

We review and discuss current knowledge about sex determination in amphibians. The astonishing wide variety of mechanisms of genotypic sex determination is presented and discussed in an evolutionary context. We recall the natural occurrence of transitory juvenile hermaphroditism in some species. Our present knowledge of the mechanisms of sex determination in amphibians is compared to that in mammals. The influence of epigenetic factors, and especially temperature is highlighted. In amphibians, the influence of temperature on sexual differentiation, that can prevail over genotypic sex determination, remains poorly considered in publications. We suggest that studies on genetic and epigenetic factors of gonadal sex differentiation in amphibians could provide substantial information on the evolutionary process of sex determination mechanisms in current living vertebrates.     

Diverse and variable sex determination mechanisms in vertebrates

Sex is prevalent in nature and sex determination is one of the most fundamental biological processes, while the way of initiating female and male development exhibits remarkable diversity and variability across vertebrates. The knowledge on why and how sex determination mechanisms evolve unusual plasticity remains limited. Here, we summarize sex determination systems, master sex-determining genes and gene-regulatory networks among vertebrates. Recent research advancements on sex determination system transition are also introduced and discussed in some non-model animals with multiple sex determination mechanisms. This review will provide insights into the origin, transition and evolutionary adaption of different sex determination strategies in vertebrates, as well as clues for future perspectives in this field.     

非哺乳类脊椎动物性别决定的研究进展

比较了非哺乳类脊椎动物和哺乳动物性别决定的差异,从影响非哺乳类脊椎动物性别决定的环境因子及其机制两方面回顾了性别决定的研究进展,分析环境因素和类固醇激素在性别决定中的作用,指出了非哺乳类脊椎动物性别决定研究中需要进一步探讨的问题。     

Sex, genes, and heat: triggers of diversity.

In vertebrates, sex is determined by a surprising variety of mechanisms. In many reptiles, the primary testis or ovary-determining trigger is regulated by egg incubation temperature. This temperature dependent sex determining (TSD) mechanism occurs in all crocodilians and marine turtles examined to date and is common in terrestrial turtles and viviparous lizards (Ewert et al. 1994. J Exp Zool 270:3-15; Lang and Andrews. 1994. J Exp Biol 270:28-44; Mrosovsky. 1994. J Exp Zool 270:16-27; Pieau. 1996. Bioessays 18:19-26; Viets et al. 1994. J Exp Zool 270:45-56; Wibbels et al. 1998. J Exp Zool 281:409-416). In contrast, sex in mammals and birds is determined chromosomally (CSD). Despite these differences, morphological development of the gonads in all these vertebrate groups appears to have been conserved through evolution. Therefore, the genetic mechanisms triggering sex determination appear not to have been conserved through evolution, although the basic genetic pathway controlling the morphological differentiation of the gonads appears to have been conserved.     

Ecdysteroids： the overlooked sex steroids of insects？ Males： the black box

The paradigm, still around in textbooks, that ＇in insects sex is strictly genetic, thus that they do not have sex hormones＇, is mainly based on a wrong interpretation of the ＇gynandromorph argument＇. It is no longer tenable. Given the fact that vertebrates and invertebrates probably had a common, sexually reproducing ancestor, there is no reason to assume that only vertebrates need sex hormones. The major function of sex hormones is to inform the somatoplasm about developmental changes that take place in the gonads. In contrast to juvenile hormone and neuropeptides, ecdysteroids meet all criteria to act as sex hormones, which was probably their ancient role. Their much better documented role in moulting and metamorphosis was a secondary acquisition that enabled arthropods to cope with growth problems, imposed by a rigid cuticle. Female insects use 20-hydroxyecdysone （20E）, secreted by the follicle cells of the ovary, in a similar way as females of egg-laying vertebrates use estrogens. For a variety of reasons, the possibility that ecdysteroids, in particular ecdysone （E）, might also act as sex hormones in male insects, thus as the counterpart of testosterone of vertebrates, has been very much overlooked. Thanks to the recent discovery of the molecular basis of the haploid-diploid system of sex determination in the honeybee, the characterization of Halloween genes, proteomics, RNAi and so on, it now becomes possible to verify whether in insects, as with vertebrates, males are the endocrinologically default gender form.     

What was the ancestral sex‐determining mechanism in amniote vertebrates?

Amniote vertebrates, the group consisting of mammals and reptiles including birds, possess various mechanisms of sex determination. Under environmental sex determination (ESD), the sex of individuals depends on the environmental conditions occurring during their development and therefore there are no sexual differences present in their genotypes. Alternatively, through the mode of genotypic sex determination (GSD), sex is determined by a sex‐specific genotype, i.e. by the combination of sex chromosomes at various stages of differentiation at conception. As well as influencing sex determination, sex‐specific parts of genomes may, and often do, develop specific reproductive or ecological roles in their bearers. Accordingly, an individual with a mismatch between phenotypic (gonadal) and genotypic sex, for example an individual sex‐reversed by environmental effects, should have a lower fitness due to the lack of specialized, sex‐specific parts of their genome. In this case, evolutionary transitions from GSD to ESD should be less likely than transitions in the opposite direction. This prediction contrasts with the view that GSD was the ancestral sex‐determining mechanism for amniote vertebrates. Ancestral GSD would require several transitions from GSD to ESD associated with an independent dedifferentiation of sex chromosomes, at least in the ancestors of crocodiles, turtles, and lepidosaurs (tuataras and squamate reptiles). In this review, we argue that the alternative theory postulating ESD as ancestral in amniotes is more parsimonious and is largely concordant with the theoretical expectations and current knowledge of the phylogenetic distribution and homology of sex‐determining mechanisms.     

Z and W chromosomes of chickens: studies on their gene functions in sex determination and sex differentiation

Since the discovery of SRY/SRY as a testis-determining gene on the mammalian Y chromosome in 1990, extensive studies have been carried out on the immediate target of SRY/SRY and genes functioning in the course of testis development. Comparative studies in non-mammalian vertebrates including birds have failed to find a gene equivalent to SRY/SRY, whereas they have suggested that most of the downstream factors found in mammals including SOX9 are also involved in the process of gonadal differentiation. Although a gene whose function is to trigger the cascade of gene expression toward gonadal differentiation has not been identified yet on either W or Z chromosomes of birds, a few interesting genes have been found recently on the sex chromosomes of chickens and their possible roles in sex determination or sex differentiation are being investigated. It is the purpose of this review to summarize the present knowledge of these sex chromosome-linked genes in chickens and to give perspectives and point out questions concerning the mechanisms of avian sex determination.     

Decoding sex: Elucidating sex determination and how high-quality genome assemblies are untangling the evolutionary dynamics of sex chromosomes

Sexual reproduction is a diverse and widespread process. In gonochoristic species, the differentiation of sexes occurs through diverse mechanisms, influenced by environmental and genetic factors. In most vertebrates, a master-switch gene is responsible for triggering a sex determination network. However, only a few genes have acquired master-switch functions, and this process is associated with the evolution of sex-chromosomes, which have a significant influence in evolution. Additionally, their highly repetitive regions impose challenges for high-quality sequencing, even using high-throughput, state-of-the-art techniques. Here, we review the mechanisms involved in sex determination and their role in the evolution of species, particularly vertebrates, focusing on sex chromosomes and the challenges involved in sequencing these genomic elements. We also address the improvements provided by the growth of sequencing projects, by generating a massive number of near-gapless, telomere-to-telomere, chromosome-level, phased assemblies, increasing the number and quality of sex-chromosome sequences available for further studies.     

How do germ cells choose their sex? Drosophila as a paradigm

Sex determination in the germ line may either rely on cell-autonomous genetic information, or it may be imposed during development by inductive somatic signals. In Drosophila, both mechanisms contribute to ensure that germ cells are oogenic when differentiating in females and spermatogenic when differentiating in males. Some of the genes that are involved in germ line sex determination have been identified. In other species, including vertebrates, inductive signals are commonly used to determine the sex of germ cells.     

Sexual determination and differentiation in teleost fish

The present work reviews the latest information on the cellular, molecular and physiological aspects of sexual determination and differentiation in teleost fish. The group exhibits a large variety of mechanisms of sexual determination. These may be genetic, or depend on environmental conditions such as temperature, pH, and social factors, all of which can influence the proportion of the sexes. Additionally, sex steroids play an important role in the regulation of sexual differentiation. The patterns of gonadal sexual differentiation are diverse, and species may be hermaphroditic or gonochoristic, some of the latter displaying juvenile hermaphroditism. In recent years, several genes involved in the sexual determination and differentiation pathways in vertebrates, particularly in mammals, have also been characterized in teleosts. Conserved as well as diversified functions have been proposed.     

Characterisation and expression of Sox9 in the Leopard gecko, Eublepharis macularius

Since the discovery of the sex-determining gene, Sry, a number of genes have been identified which are involved in sex determination and gonadogenesis in mammals. Although Sry is known to be the testis-determining factor in mammals, this is not the case in non-mammalian vertebrates. Sox9 is another gene that has been shown to have a male-specific role in sex determination, but, unlike Sry, Sox9 has been shown to be involved in sex determination in mammals, birds, and reptiles. This is the first gene to be described that has a conserved role in sex determination in species with either chromosomal or environmental sex-determining mechanisms. Many reptiles do not have sex chromosomes but exhibit temperature-dependent sex determination (TSD). Sox9 has been shown to be expressed in both turtle and alligator during gonadogenesis. To determine if Sox9 also has a role in a gecko species with TSD, we studied gonadal expression of Sox9 during embryonic development of the Leopard gecko (Eublepharis macularius). Gecko Sox9 was found to be highly conserved at the nucleotide level when compared to other vertebrate species including human, chick, alligator, and turtle. Sox9 was found to be expressed in embryos incubated at the male-producing temperature (32.5 degrees C) as well as in embryos incubated at the female-producing temperatures (26 and 34 degrees C), Northern blot analysis showed that Sox9 was expressed at both temperatures from morphological stages 31 to 37. mRNA in situ hybridisation on isolated urogenital systems showed expression at both female- and male-producing temperatures up to stage 36. After this stage, no expression was seen in the female gonads but expression remained in the male. These data provide further evidence that Sox9 is an essential component of a testis-determining pathway that is conserved in species with differing sex-determining mechanisms.     

Novel evolutionary pathways of sex‐determining mechanisms

Evolutionary transitions between sex‐determining mechanisms (SDMs) are an enigma. Among vertebrates, individual sex (male or female) is primarily determined by either genes (genotypic sex determination, GSD) or embryonic incubation temperature (temperature‐dependent sex determination, TSD), and these mechanisms have undergone repeated evolutionary transitions. Despite this evolutionary lability, transitions from GSD (i.e. from male heterogamety, XX/XY, or female heterogamety, ZZ/ZW) to TSD are an evolutionary conundrum, as they appear to require crossing a fitness valley arising from the production of genotypes with reduced viability owing to being homogametic for degenerated sex chromosomes (YY or WW individuals). Moreover, it is unclear whether alternative (e.g. mixed) forms of sex determination can persist across evolutionary time. It has previously been suggested that transitions would be easy if temperature‐dependent sex reversal (e.g. XX male or XY female) was asymmetrical, occurring only in the homogametic sex. However, only recently has a mechanistic model of sex determination emerged that may allow such asymmetrical sex reversal. We demonstrate that selection for TSD in a realistic sex‐determining system can readily drive evolutionary transitions from GSD to TSD that do not require the production of YY or WW individuals. In XX/XY systems, sex reversal (female to male) occurs in a portion of the XX individuals only, leading to the loss of the Y allele (or chromosome) from the population as XX individuals mate with each other. The outcome is a population of XX individuals whose sex is determined by incubation temperature (TSD). Moreover, our model reveals a novel evolutionarily stable state representing a mixed‐mechanism system that has not been revealed by previous approaches. This study solves two long‐standing puzzles of the evolution of sex‐determining mechanisms by illuminating the evolutionary pathways and endpoints.     

Increase of cortisol levels after temperature stress activates dmrt1a causing female‐to‐male sex reversal and reduced germ cell number in medaka

In vertebrates, there is accumulating evidence that environmental factors as triggers for sex determination and genetic sex determination are not two opposing alternatives but that a continuum of mechanisms bridge those extremes. One prominent example is the model fish species Oryzias latipes which has a stable XX/XY genetic sex determination system, but still responds to environmental cues, where high temperatures lead to female‐to‐male sex reversal. However, the mechanisms behind are still unknown. We show that high temperatures increase primordial germ cells (PGC) numbers before they reach the genital ridge, which, in turn, regulates the germ cell proliferation. Complete ablation of PGCs led to XX males with germ cell less testis, whereas experimentally increased PGC numbers did not reverse XY genotypes to female. For the underlying molecular mechanism, we provide support for the explanation that activation of the dmrt1a gene by cortisol during early development of XX embryos enables this autosomal gene to take over the role of the male determining Y‐chromosomal dmrt1bY.     

Sex determination: insights from the chicken

Not all vertebrates share the familiar system of XX:XY sex determination seen in mammals. In the chicken and other birds, sex is determined by a ZZ:ZW sex chromosome system. Gonadal development in the chicken has provided insights into the molecular genetics of vertebrate sex determination and how it has evolved. Such comparative studies show that vertebrate sex-determining pathways comprise both conserved and divergent elements. The chicken embryo resembles lower vertebrates in that estrogens play a central role in gonadal sex differentiation. However, several genes shown to be critical for mammalian sex determination are also expressed in the chicken, but their expression patterns differ, indicating functional plasticity. While the genetic trigger for sex determination in birds remains unknown, some promising candidate genes have recently emerged. The Z-linked gene, DMRT1, supports the Z-dosage model of avian sex determination. Two novel W-linked genes, ASW and FET1, represent candidate female determinants.     

Mammalian sex--Origin and evolution of the Y chromosome and SRY

Sex determination in vertebrates is accomplished through a highly conserved genetic pathway. But surprisingly, the downstream events may be activated by a variety of triggers, including sex determining genes and environmental cues. Amongst species with genetic sex determination, the sex determining gene is anything but conserved, and the chromosomes that bear this master switch subscribe to special rules of evolution and function. In mammals, with a few notable exceptions, female are homogametic (XX) and males have a single X and a small, heterochromatic and gene poor Y that bears a male dominant sex determining gene SRY. The bird sex chromosome system is the converse in that females are the heterogametic sex (ZW) and males the homogametic sex (ZZ). There is no SRY in birds, and the dosage-sensitive Z-borne DMRT1 gene is a credible candidate sex determining gene. Different sex determining switches seem therefore to have evolved independently in different lineages, although the complex sex chromosomes of the platypus offer us tantalizing clues that the mammal XY system may have evolved directly from an ancient reptile ZW system. In this review we will discuss the organization and evolution of the sex chromosomes across a broad range of mammals, and speculate on how the Y chromosome, and SRY, evolved.     

Sex-ratio adjustment when relatives interact: a test of constraints on adaptation

Studies of sex allocation offer excellent opportunities for examining the constraints and limits on adaptation. A major topic of debate within this field concerns the extent to which the ability of individuals to adaptively manipulate their offspring sex ratio is determined by constraints such as the method of sex determination. We address this problem by comparing the extent of sex-ratio adjustment across taxa with different methods of sex determination, under the common selective scenario of interactions between relatives. These interactions comprise the following: local resource competition (LRC), local mate competition (LMC), and local resource enhancement (LRE). We found that: (1) species with supposedly constraining methods of sex determination showed consistent sex-ratio adjustment in the predicted direction; (2) vertebrates with chromosomal sex determination (CSD) showed less adjustment then haplodiploid invertebrates; (3) invertebrates with possibly constraining sex-determination mechanisms (CSD and pseudo-arrhenotoky) did not show less adjustment then haplodiploid invertebrates; (4) greater sex-ratio adjustment was seen in response to LRC and LMC than LRE; (5) greater sex-ratio adjustment was seen in response to interactions between relatives (LRC, LMC, and LRE) compared to responses to other environmental factors. Our results also illustrate the problem that sex-determination mechanism and selective pressure are confounded across taxa because vertebrates with CSD are influenced primarily by LRE whereas invertebrates are influenced by LRC and LMC. Overall, our analyses suggest that sex-allocation theory needs to consider simultaneously the influence of variable selection pressures and variable constraints when applying general theory to specific cases.