Genetic evidence for co-occurrence of chromosomal and thermal sex-determining systems in a lizard

An individual's sex depends upon its genes (genotypic sex determination or GSD) in birds and mammals, but reptiles are more complex: some species have GSD whereas in others, nest temperatures determine offspring sex (temperature-dependent sex determination). Previous studies suggested that montane scincid lizards (Bassiana duperreyi, Scincidae) possess both of these systems simultaneously: offspring sex is determined by heteromorphic sex chromosomes (XX-XY system) in most natural nests, but sex ratio shifts suggest that temperatures override chromosomal sex in cool nests to generate phenotypically male offspring even from XX eggs. We now provide direct evidence that incubation temperatures can sex-reverse genotypically female offspring, using a DNA sex marker. Application of exogenous hormone to eggs also can sex-reverse offspring (oestradiol application produces XY as well as XX females). In conjunction with recent work on a distantly related lizard taxon, our study challenges the notion of a fundamental dichotomy between genetic and thermally determined sex determination, and hence the validity of current classification schemes for sex-determining systems in reptiles.     

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.     

Phylogeny of sex‐determining mechanisms in squamate reptiles: are sex chromosomes an evolutionary trap?

Squamate reptiles possess two general modes of sex determination: (1) genotypic sex determination (GSD), where the sex of an individual is determined by sex chromosomes, i.e. by sex‐specific differences in genotype; and (2) temperature‐dependent sex determination (TSD), where sex chromosomes are absent and sex is determined by nongenetic factors. After gathering information about sex‐determining mechanisms for more than 400 species, we employed comparative phylogenetic analyses to reconstruct the evolution of sex determination in Squamata. Our results suggest relative uniformity in sex‐determining mechanisms in the majority of the squamate lineages. Well‐documented variability is found only in dragon lizards (Agamidae) and geckos (Gekkota). Polarity of the sex‐determining mechanisms in outgroups identified TSD as the ancestral mode for Squamata. After extensive review of the literature, we concluded that to date there is no known well‐documented transition from GSD to TSD in reptiles, although transitions in the opposite direction are plentiful and well corroborated by cytogenetic evidence. We postulate that the evolution of sex‐determining mechanisms in Squamata was probably restricted to the transitions from ancestral TSD to GSD. In other words, transitions were from the absence of sex chromosomes to the emergence of sex chromosomes, which have never disappeared and constitute an evolutionary trap. This evolutionary trap hypothesis could change the understanding of phylogenetic conservatism of sex‐determining systems in many large clades such as butterflies, snakes, birds, and mammals. © 2009 The Linnean Society of London, Zoological Journal of the Linnean Society, 2009, 156 , 168–183.     

Offspring sex is not related to maternal allocation of yolk steroids in the lizard Bassiana duperreyi (Scincidae)

The eggs of birds and reptiles contain detectable levels of several steroid hormones, and experimental application of such steroids can reverse genetically determined sex of the offspring. However, any causal influence of maternally derived yolk steroids on sex determination in birds and reptiles remains controversial. We measured yolk hormones (dihydrotestosterone, testosterone, and 17 beta-estradiol) in newly laid eggs of the montane scincid lizard Bassiana duperreyi. This species is well suited to such an analysis because (1) offspring sex is influenced by incubation temperatures and egg size as well as by sex chromosomes, suggesting that yolk hormones might somehow be involved in the complex pathways of sex determination, and (2) experimental application of either estradiol or fadrozole to such eggs strongly influences offspring sex. We obtained yolk by biopsy, before incubating the eggs at a temperature that produces a 50:50 sex ratio. Yolk steroid levels varied over a threefold range between eggs from different clutches, but there were no significant differences in yolk steroids, or in relative composition of steroids, between eggs destined to become male versus female. Further, yolk steroid concentrations were not significantly related to egg size. Thus, yolk steroid hormones do not appear to play a critical role in sex determination for B. duperreyi.     

Environmental sex reversal,Trojan sex genes,and sex ratio adjustment: conditions and population consequences

The great diversity of sex determination mechanisms in animals and plants ranges from genetic sex determination (GSD, e.g. mammals, birds, and most dioecious plants) to environmental sex determination (ESD, e.g. many reptiles) and includes a mixture of both, for example when an individual’s genetically determined sex is environmentally reversed during ontogeny (ESR, environmental sex reversal, e.g. many fish and amphibia). ESD and ESR can lead to widely varying and unstable population sex ratios. Populations exposed to conditions such as endocrine‐active substances or temperature shifts may decline over time due to skewed sex ratios, a scenario that may become increasingly relevant with greater anthropogenic interference on watercourses. Continuous exposure of populations to factors causing ESR could lead to the extinction of genetic sex factors and may render a population dependent on the environmental factors that induce the sex change. However, ESR also presents opportunities for population management, especially if the Y or W chromosome is not, or not severely, degenerated. This seems to be the case in many amphibians and fish. Population growth or decline in such species can potentially be controlled through the introduction of so‐called Trojan sex genes carriers, individuals that possess sex chromosomes or genes opposite from what their phenotype predicts. Here, we review the conditions for ESR, its prevalence in natural populations, the resulting physiological and reproductive consequences, and how these may become instrumental for population management.     

A tropical oviparous lizard, Calotes versicolor, exhibiting a potentially novel FMFM pattern of temperature-dependent sex determination

Among squamate reptiles, lizards exhibit an impressive array of sex-determining modes viz. genotypic sex determination, temperature-dependent sex determination, co-occurrence of both these and those that reproduce parthenogenetically. The oviparous lizard, Calotes versicolor, lacks heteromorphic sex chromosomes and there are no reports on homomorphic chromosomes. Earlier studies on this species presented little evidence to the sex-determining mechanism. Here we provide evidences for the potential role played by incubation temperature that has a significant effect (P < 0.01) on gonadal sex and sex ratio. The eggs were incubated at 14 different incubation temperatures. Interestingly, 100% males were produced at low (25.5 ± 0.5 ° C) as well as high (34 ± 0.5 ° C) incubation temperatures and 100% females were produced at low (23.5 ± 0.5 ° C) and high (31.5 ± 0.5 ° C) temperatures, clearly indicating the occurrence of TSD in this species. Sex ratios of individual clutches did not vary at any of the critical male-producing or female-producing temperatures within as well as across the seasons. However, clutch sex ratios were female- or male-biased at intermediate temperatures. Thermosensitive period occurred during the embryonic stages 30-33. Three pivotal temperatures operate producing 1:1 sex ratio. Histology of gonad and accessory reproductive structures provide additional evidence for TSD. The sex-determining pattern, observed for the first time in this species, that neither compares to Pattern I [Ia (MF) and Ib (FM)] nor to Pattern II (FMF), is being referred to as FMFM pattern of TSD. This novel FMFM pattern of sex ratio exhibited by C. versicolor may have an adaptive significance in maintaining sex ratio.     

Fitness effects of the timing of hatching may drive the evolution of temperature-dependent sex determination in short-lived lizards

In several species of short-lived Australian agamid lizards, an individual’s sex is determined by the nest temperatures encountered during incubation. The adaptive significance of such systems remains unclear. Here, we explore the hypothesis that (1) the optimal timing of hatching differs between the sexes, and thus (2) temperature-dependent sex determination (TSD) enhances maternal and offspring fitness by generating seasonal shifts in offspring sex ratios. Our model predicts that TSD can indeed enhance maternal fitness returns in short-lived lizards if (1) male–male competition is intense, thus reducing mating success of newly-matured males (but not females), and (2) the nesting season is prolonged, such that seasonal effects become significant. Available data on the distribution of TSD in Australian agamid lizards broadly support these predictions. Because both the level of male–male competition and the length of nesting season can vary at small spatial and temporal scales, selective forces on sex-determining mechanisms also should vary. Hence, our model predicts extensive small-scale (intraspecific) variation in sex-determining systems within agamid lizards, as well as among species.     

How to identify sex chromosomes and their turnover

Although sex is a fundamental component of eukaryotic reproduction, the genetic systems that control sex determination are highly variable. In many organisms the presence of sex chromosomes is associated with female or male development. Although certain groups possess stable and conserved sex chromosomes, others exhibit rapid sex chromosome evolution, including transitions between male and female heterogamety, and turnover in the chromosome pair recruited to determine sex. These turnover events have important consequences for multiple facets of evolution, as sex chromosomes are predicted to play a central role in adaptation, sexual dimorphism, and speciation. However, our understanding of the processes driving the formation and turnover of sex chromosome systems is limited, in part because we lack a complete understanding of interspecific variation in the mechanisms by which sex is determined. New bioinformatic methods are making it possible to identify and characterize sex chromosomes in a diverse array of non‐model species, rapidly filling in the numerous gaps in our knowledge of sex chromosome systems across the tree of life. In turn, this growing data set is facilitating and fueling efforts to address many of the unanswered questions in sex chromosome evolution. Here, we synthesize the available bioinformatic approaches to produce a guide for characterizing sex chromosome system and identity simultaneously across clades of organisms. Furthermore, we survey our current understanding of the processes driving sex chromosome turnover, and highlight important avenues for future research.     

Genetics of sex determination in flowering plants

Most flowering plant species are hermaphroditic, but a small number of species in most plant families are unisexual (i.e., an individ-ual will produce only male or female gametes). Because species with unisexual flowers have evolved repeatedly from hermaphroditic progenitors, the mechanisms controlling sex determination in flowering plants are extremely diverse. Sex is most strongly determined by genotype in all species but the mechanisms range from a single controlling locus to sex chromosomes bearing several linked locirequired for sex determination. Plant hormones also influence sex expression with variable effects from species to species. Here, we review the genetic control of sex determination from a number of plant species to illustrate the variety of extant mechanisms. We emphasize species that are now used as models to investigate the molecular biology of sex determination. We also present our own investigations of the structure of plant sex chromosomes of white campion (Silene latifolia - Melan-drium album). The cytogenetic basis of sex determination in white campion is similar to mammals in that it has a male-specific Y-chromosome that carries dominant male determining genes. If one copy of this chromosome is in the genome, the plant is male. Otherwise it is female. Like mammalian Y-chromosomes, the white campion Y-chromosome is rich in repetitive DNA. We isolated repetitive sequences from microdissected Y-chromosomes of white campion to study the distribution of homologous repeated sequences on the Y-chromosome and the other chromosomes. We found the Y to be especially rich in repetitive sequences that were generally dispersed over all the white campion chromosomes. Despite its repetitive character, the Y-chromosome is mainly euchromatic. This may be due to the relatively recent evolution of the white campion sex chromosomes compared to the sex chromosomes of animals. © 1994 Wiley-Liss, Inc.     

Sex without sex chromosomes: genetic architecture of multiple loci independently segregating to determine sex ratios in the copepod Tigriopus californicus

Sex‐determining systems are remarkably diverse and may evolve rapidly. Polygenic sex‐determination systems are predicted to be transient and evolutionarily unstable, yet examples have been reported across a range of taxa. Here, we provide the first direct evidence of polygenic sex determination in Tigriopus californicus, a harpacticoid copepod with no heteromorphic sex chromosomes. Using genetically distinct inbred lines selected for male‐ and female‐biased clutches, we generated a genetic map with 39 SNPs across 12 chromosomes. Quantitative trait locus mapping of sex ratio phenotype (the proportion of male offspring produced by an F2 female) in four F2 families revealed six independently segregating quantitative trait loci on five separate chromosomes, explaining 19% of the variation in sex ratios. The sex ratio phenotype varied among loci across chromosomes in both direction and magnitude, with the strongest phenotypic effects on chromosome 10 moderated to some degree by loci on four other chromosomes. For a given locus, sex ratio phenotype varied in magnitude for individuals derived from different dam lines. These data, together with the environmental factors known to contribute to sex determination, characterize the underlying complexity and potential lability of sex determination, and confirm the polygenic architecture of sex determination in T. californicus.     

Reproductive ecology of the jacky dragon (Amphibolurus muricatus): an agamid lizard with temperature‐dependent sex determination

Abstract The jacky dragon, Amphibolurus muricatus (White, ex Shaw 1790) is a medium sized agamid lizard from the southeast of Australia. Laboratory incubation trials show that this species possesses temperature‐dependent sex determination. Both high and low incubation temperatures produced all female offspring, while varying proportions of males hatched at intermediate temperatures. Females may lay several clutches containing from three to nine eggs during the spring and summer. We report the first field nest temperature recordings for a squamate reptile with temperature‐dependent sex determination. Hatchling sex is determined by nest temperatures that are due to the combination of daily and seasonal weather conditions, together with maternal nest site selection. Over the prolonged egg‐laying season, mean nest temperatures steadily increase. This suggests that hatchling sex is best predicted by the date of egg laying, and that sex ratios from field nests will vary over the course of the breeding season. Lizards hatching from eggs laid in the spring (October) experience a longer growing season and should reach a larger body size by the beginning of their first reproductive season, compared to lizards from eggs laid in late summer (February). Adult male A. muricatus attain a greater maximum body size and have relatively larger heads than females, possibly as a consequence of sexual selection due to male‐male competition for territories and mates. If reproductive success in males increases with larger body size, then early hatching males may obtain a greater fitness benefit as adults, compared to males that hatch in late summer. We hypothesize that early season nests should produce male‐biased sex ratios, and that this provides an adaptive explanation for temperature‐dependent sex determination in A. muricatus.     

早期胚胎的发育选择:性别决定

性别决定是一个复杂的发育调控过程, 早期胚胎发育过程中, 雌雄二者必居其一的发育选择是胚胎性腺形成必须的发育决定。文章综述了动物性别决定的遗传系统、性腺发生、性别决定关键基因及其作用机制, 从分子进化的角度分析了性染色体与性别决定形成机制, 提示性别决定基因在进化中总是趋向异配性染色体。     

CONSERVED SEX CHROMOSOMES ACROSS ADAPTIVELY RADIATED ANOLIS LIZARDS

Vertebrates possess diverse sex‐determining systems, which differ in evolutionary stability among particular groups. It has been suggested that poikilotherms possess more frequent turnovers of sex chromosomes than homoiotherms, whose effective thermoregulation can prevent the emergence of the sex reversals induced by environmental temperature. Squamate reptiles used to be regarded as a group with an extensive variability in sex determination; however, we document how the rather old radiation of lizards from the genus Anolis, known for exceptional ecomorphological variability, was connected with stability in sex chromosomes. We found that 18 tested species, representing most of the phylogenetic diversity of the genus, share the gene content of their X chromosomes. Furthermore, we discovered homologous sex chromosomes in species of two genera (Sceloporus and Petrosaurus) from the family Phrynosomatidae, serving here as an outgroup to Anolis. We can conclude that the origin of sex chromosomes within iguanas largely predates the Anolis radiation and that the sex chromosomes of iguanas remained conserved for a significant part of their evolutionary history. Next to therian mammals and birds, Anolis lizards therefore represent another adaptively radiated amniote clade with conserved sex chromosomes. We argue that the evolutionary stability of sex‐determining systems may reflect an advanced stage of differentiation of sex chromosomes rather than thermoregulation strategy.     

A General Model to Explain Repeated Turnovers of Sex Determination in the Salicaceae

Dioecy, the presence of separate sexes on distinct individuals, has evolved repeatedly in multiple plant lineages. However, the specific mechanisms by which sex systems evolve and their commonalities among plant species remain poorly understood. With both XY and ZW sex systems, the family Salicaceae provides a system to uncover the evolutionary forces driving sex chromosome turnovers. In this study, we performed a genome-wide association study to characterize sex determination in two Populus species, P. euphratica and P. alba. Our results reveal an XY system of sex determination on chromosome 14 of P. euphratica, and a ZW system on chromosome 19 of P. alba. We further assembled the corresponding sex-determination regions, and found that their sex chromosome turnovers may be driven by the repeated translocations of a Helitron-like transposon. During the translocation, this factor may have captured partial or intact sequences that are orthologous to a type-A cytokinin response regulator gene. Based on results from this and other recently published studies, we hypothesize that this gene may act as a master regulator of sex determination for the entire family. We propose a general model to explain how the XY and ZW sex systems in this family can be determined by the same RR gene. Our study provides new insights into the diversification of incipient sex chromosomes in flowering plants by showing how transposition and rearrangement of a single gene can control sex in both XY and ZW systems.     

Effects of global warming on sex ratios in fishes

In fishes, sex is determined by genetics, the environment or an interaction of both. Temperature is among the most important environmental factors that can affect sex determination. As a consequence, changes in temperature at critical developmental stages can induce biases in primary sex ratios in some species. However, early sex ratios can also be biased by sex-specific tolerances to environmental stresses that may, in some cases, be amplified by changes in water temperature. Sex-specific reactions to environmental stress have been observed at early larval stages before gonad formation starts. It is therefore necessary to distinguish between temperature effects on sex determination, generally acting through the stress axis or epigenetic mechanisms, and temperature effects on sex-specific mortality. Both are likely to affect sex ratios and hence population dynamics. Moreover, in cases where temperature effects on sex determination lead to genotype–phenotype mismatches, long-term effects on population dynamics are possible, for example temperature-induced masculinization potentially leading to the loss of Y chromosomes or feminization to male-biased operational sex ratios in future generations. To date, most studies under controlled conditions conclude that if temperature affects sex ratios, elevated temperatures mostly lead to a male bias. The few studies that have been performed on wild populations seem to confirm this general trend. Recent findings suggest that transgenerational plasticity could mitigate the effects of warming on sex ratios in some populations.     

The contrasting role of heterochromatin in the differentiation of sex chromosomes: an overview from Neotropical fishes

During the evolutionary process of the sex chromosomes, a general principle that arises is that cessation or a partial restriction of recombination between the sex chromosome pair is necessary. Data from phylogenetically distinct organisms reveal that this phenomenon is frequently associated with the accumulation of heterochromatin in the sex chromosomes. Fish species emerge as excellent models to study this phenomenon because they have much younger sex chromosomes compared to higher vertebrates and many other organisms making it possible to follow their steps of differentiation. In several Neotropical fish species, the heterochromatinization, accompanied by amplification of tandem repeats, represents an important step in the morphological differentiation of simple sex chromosome systems, especially in the ZZ/ZW sex systems. In contrast, multiple sex chromosome systems have no additional increase of heterochromatin in the chromosomes. Thus, the initial stage of differentiation of the multiple sex chromosome systems seems to be associated with proper chromosomal rearrangements, whereas the simple sex chromosome systems have an accumulation of heterochromatin. In this review, attention has been drawn to this contrasting role of heterochromatin in the differentiation of simple and multiple sex chromosomes of Neotropical fishes, highlighting their surprising evolutionary dynamism.     

Evolutionary stability of sex chromosomes in snakes

Amniote vertebrates possess various mechanisms of sex determination, but their variability is not equally distributed. The large evolutionary stability of sex chromosomes in viviparous mammals and birds was believed to be connected with their endothermy. However, some ectotherm lineages seem to be comparably conserved in sex determination, but previously there was a lack of molecular evidence to confirm this. Here, we document a stability of sex chromosomes in advanced snakes based on the testing of Z-specificity of genes using quantitative PCR (qPCR) across 37 snake species (our qPCR technique is suitable for molecular sexing in potentially all advanced snakes). We discovered that at least part of sex chromosomes is homologous across all families of caenophidian snakes (Acrochordidae, Xenodermatidae, Pareatidae, Viperidae, Homalopsidae, Colubridae, Elapidae and Lamprophiidae). The emergence of differentiated sex chromosomes can be dated back to about 60 Ma and preceded the extensive diversification of advanced snakes, the group with more than 3000 species. The Z-specific genes of caenophidian snakes are (pseudo)autosomal in the members of the snake families Pythonidae, Xenopeltidae, Boidae, Erycidae and Sanziniidae, as well as in outgroups with differentiated sex chromosomes such as monitor lizards, iguanas and chameleons. Along with iguanas, advanced snakes are therefore another example of ectothermic amniotes with a long-term stability of sex chromosomes comparable with endotherms.     

SEX RATIO VARIATION IN THE LESSONIA NIGRESCENS COMPLEX (LAMINARIALES,PHAEOPHYCEAE): EFFECT OF LATITUDE,TEMPERATURE, AND MARGINALITY1

Little is known about variation of sex ratio, the proportion of males to females, in natural populations of seaweed, though it is a major determinant of the mating system. The observation of sexual chromosomes in kelps suggested that sex is partly genetically determined. However, it is probably not purely genetic since the sex ratio can be modified by environmental factors such as salinity or temperature. In this paper, sex ratio variation was studied in the kelp Lessonia nigrescens Bory complex, recently identified as two cryptic species occurring along the Chilean coast: one located north and the other south of the biogeographic boundary at latitude 29°–30° S. The life cycle of L. nigrescens is characterized by an alternation of microscopic haploid gametophytic individuals and large macroscopic fronds of diploid sporophytes. The sex ratio was recorded in progenies from 241 sporophytic individuals collected from 13 populations distributed along the Chilean coast in order (i) to examine the effect of an environmental gradient coupled with latitude, and (ii) to compare marginal populations to central populations of the two species. In addition, we tested the hypothesis that the sex ratios of the two cryptic species would be affected differently by temperature. First, our results demonstrate that sex ratio seems to be mainly genetically determined and temperature can significantly modify it. Populations of the northern species showed a lower frequency of males at 14°C than at 10°C, whereas populations of the southern species showed the opposite pattern. Second, both species displayed an increased variation in sex ratio at the range limits. This greater variation at the margins could be due either to differential mortality between sexes or to geographic parthenogenesis (asexual reproduction).     

Maintenance of polygenic sex determination in a fluctuating environment: an individual‐based model

R. A. Fisher predicted that individuals should invest equally in offspring of both sexes, and that the proportion of males and females produced (the primary sex ratio) should evolve towards 1:1 when unconstrained. For many species, sex determination is dependent on sex chromosomes, creating a strong tendency for balanced sex ratios, but in other cases, multiple autosomal genes interact to determine sex. In such cases, the maintenance of multiple sex‐determining alleles at multiple loci and the consequent among‐family variability in sex ratios presents a puzzle, as theory predicts that such systems should be unstable. Theory also predicts that environmental influences on sex can complicate outcomes of genetic sex determination, and that population structure may play a role. Tigriopus californicus, a copepod that lives in splash‐pool metapopulations and exhibits polygenic and environment‐dependent sex determination, presents a test case for relevant theory. We use this species as a model for parameterizing an individual‐based simulation to investigate conditions that could maintain polygenic sex determination. We find that metapopulation structure can delay the degradation of polygenic sex determination and that periods of alternating frequency‐dependent selection, imposed by seasonal fluctuations in environmental conditions, can maintain polygenic sex determination indefinitely.