For a long time, snakes were presented as a textbook example of a group with gradual differentiation of homologous ZZ/ZW sex chromosomes. However, recent advances revealed that the ZZ/ZW sex chromosomes characterize only caenophidian snakes and certain species of boas and pythons have nonhomologous XX/XY sex chromosomes. We used genome coverage analysis in four non-caenophidian species to identify their sex chromosomes, and we examined the homology of sex chromosomes across phylogenetically informative snake lineages. We identified sex chromosomes for the first time in 13 species of non-caenophidian snakes, providing much deeper insights into the evolutionary history of snake sex chromosomes. The evolution of sex chromosomes in snakes is more complex than previously thought. Snakes may have had ancestral XX/XY sex chromosomes, which are still present in a blind snake and some boas, and there were several transitions to derived XX/XY sex chromosomes with different gene content and two or even three transitions to ZZ/ZW sex chromosomes. However, we discuss more alternative scenarios. In any case, we document that (1) some genomic regions were likely repeatedly co-opted as sex chromosomes in phylogenetically distant lineages, even with opposite types of heterogamety; (2) snake lineages differ greatly in the rate of differentiation of sex chromosomes; (3) snakes likely originally possessed sex chromosomes prone to turnovers. The sex chromosomes became evolutionarily highly stable once their differentiation progressed in the megadiverse caenophidian snakes. Snakes thus provide an ideal system for studying the evolutionary factors that drive unequal rates of differentiation, turnovers and stability of sex chromosomes.

• Keywords
• DNA-seq, genomics, qPCR, reptiles, sex chromosomes, sex determination,
• MeSH
• Biological Evolution MeSH
• Phylogeny MeSH
• Snakes * genetics   MeSH
• Evolution, Molecular MeSH
• Sex Chromosomes * genetics   MeSH
• Animals MeSH
• Check Tag
• Male MeSH
• Female MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH

Chameleons are well-known lizards with unique morphology and physiology, but their sex determination has remained poorly studied. Madagascan chameleons of the genus Furcifer have cytogenetically distinct Z and W sex chromosomes and occasionally Z1Z1Z2Z2/Z1Z2W multiple neo-sex chromosomes. To identify the gene content of their sex chromosomes, we microdissected and sequenced the sex chromosomes of F. oustaleti (ZZ/ZW) and F. pardalis (Z1Z1Z2Z2/Z1Z2W). In addition, we sequenced the genomes of a male and a female of F. lateralis (ZZ/ZW) and F. pardalis and performed a comparative coverage analysis between the sexes. Despite the notable heteromorphy and distinctiveness in heterochromatin content, the Z and W sex chromosomes share approximately 90% of their gene content. This finding demonstrates poor correlation of the degree of differentiation of sex chromosomes at the cytogenetic and gene level. The test of homology based on the comparison of gene copy number variation revealed that female heterogamety with differentiated sex chromosomes remained stable in the genus Furcifer for at least 20 million years. These chameleons co-opted for the role of sex chromosomes the same genomic region as viviparous mammals, lacertids and geckos of the genus Paroedura, which makes these groups excellent model for studies of convergent and divergent evolution of sex chromosomes.

• Keywords
• Chameleons, Homology, Karyotypes, Microdissection, Sex chromosomes, qPCR,
• MeSH
• Lizards * genetics   MeSH
• Evolution, Molecular MeSH
• Sex Chromosomes genetics   MeSH
• Sex Determination Processes genetics   MeSH
• Mammals genetics   MeSH
• Base Sequence MeSH
• DNA Copy Number Variations * MeSH
• Animals MeSH
• Check Tag
• Male MeSH
• Female MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH

Sex chromosomes are a great example of a convergent evolution at the genomic level, having evolved dozens of times just within amniotes. An intriguing question is whether this repeated evolution was random, or whether some ancestral syntenic blocks have significantly higher chance to be co-opted for the role of sex chromosomes owing to their gene content related to gonad development. Here, we summarize current knowledge on the evolutionary history of sex determination and sex chromosomes in amniotes and evaluate the hypothesis of non-random emergence of sex chromosomes. The current data on the origin of sex chromosomes in amniotes suggest that their evolution is indeed non-random. However, this non-random pattern is not very strong, and many syntenic blocks representing putatively independently evolved sex chromosomes are unique. Still, repeatedly co-opted chromosomes are an excellent model system, as independent co-option of the same genomic region for the role of sex chromosome offers a great opportunity for testing evolutionary scenarios on the sex chromosome evolution under the explicit control for the genomic background and gene identity. Future studies should use these systems more to explore the convergent/divergent evolution of sex chromosomes. This article is part of the theme issue 'Challenging the paradigm in sex chromosome evolution: empirical and theoretical insights with a focus on vertebrates (Part II)'.

• Keywords
• amniotes, co-option, sex chromosomes, vertebrates,
• MeSH
• Biological Evolution * MeSH
• Reptiles genetics   growth & development   MeSH
• Sex Chromosomes genetics   MeSH
• Sex Determination Processes * MeSH
• Birds genetics   growth & development   MeSH
• Mammals genetics   growth & development   MeSH
• Animals MeSH
• Check Tag
• Animals MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH

Amniotes possess astonishing variability in sex determination ranging from environmental sex determination (ESD) to genotypic sex determination (GSD) with highly differentiated sex chromosomes. Geckos are one of the few amniote groups with substantial variability in sex determination. What makes them special in this respect? We hypothesized that the extraordinary variability of sex determination in geckos can be explained by two alternatives: 1) unusual lability of sex determination, predicting that the current GSD systems were recently formed and are prone to turnovers; and 2) independent transitions from the ancestral ESD to later stable GSD, which assumes that geckos possessed ancestrally ESD, but once sex chromosomes emerged, they remain stable in the long term. Here, based on genomic data, we document that the differentiated ZZ/ZW sex chromosomes evolved within carphodactylid geckos independently from other gekkotan lineages and remained stable in the genera Nephrurus, Underwoodisaurus, and Saltuarius for at least 15 Myr and potentially up to 45 Myr. These results together with evidence for the stability of sex chromosomes in other gekkotan lineages support more our second hypothesis suggesting that geckos do not dramatically differ from the evolutionary transitions in sex determination observed in the majority of the amniote lineages.

• Keywords
• DNA-seq, genomics, qPCR, reptiles, sex chromosomes, sex determination,
• MeSH
• Biological Evolution MeSH
• Phylogeny MeSH
• Lizards * genetics   MeSH
• Sex Chromosomes genetics   MeSH
• Sex Determination Processes genetics   MeSH
• Animals MeSH
• Check Tag
• Animals MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH

Transitions from environmental sex determination (ESD) to genotypic sex determination (GSD) require an intermediate step of sex reversal, i.e., the production of individuals with a mismatch between the ancestral genotypic and the phenotypic sex. Among amniotes, the sole well-documented transition in this direction was shown in the laboratory in the central bearded dragon, Pogona vitticeps, where very high incubation temperatures led to the production of females with the male-typical (ZZ) genotype. These sex-reversed females then produced offspring whose sex depended on the incubation temperature. Sex-reversed animals identified by molecular and cytogenetic markers were also reported in the field, and their increasing incidence was speculated as a climate warming-driven transition in sex determination. We show that the molecular and cytogenetic markers normally sex-linked in P. vitticeps are also sex-linked in P. henrylawsoni and P. minor, which points to quite ancient sex chromosomes in this lineage. Nevertheless, we demonstrate, based on a crossing experiment with a male bearded dragon who possesses a mismatch between phenotypic sex and genotype, that the used cytogenetic and molecular markers might not be reliable for the identification of sex reversal. Sex reversal should not be considered as the only mechanism causing a mismatch between genetic sex-linked markers and phenotypic sex, which can emerge also by other processes, here most likely by a rare recombination between regions of sex chromosomes which are normally sex-linked. We warn that sex-linked, even apparently for a long evolutionary time, and sex-specific molecular and cytogenetic markers are not a reliable tool for the identification of sex-reversed individuals in a population and that sex reversal has to be verified by other approaches, particularly by observation of the sex ratio of the progeny.

• Keywords
• Molecular markers, Reversal, Sex chromosomes, Sex linkage, Vertebrates,
• MeSH
• Sex Determination Analysis MeSH
• Lizards * genetics   MeSH
• Sex Chromosomes genetics   MeSH
• Sex Ratio MeSH
• Sex Determination Processes * genetics   MeSH
• Animals MeSH
• Check Tag
• Male MeSH
• Female MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH

Differentiated sex chromosomes are believed to be evolutionarily stable, while poorly differentiated sex chromosomes are considered to be prone to turnovers. With around 1700 currently known species forming ca 15% of reptile species diversity, skinks (family Scincidae) are a very diverse group of squamates known for their large ecological and morphological variability. Skinks generally have poorly differentiated and cytogenetically indistinguishable sex chromosomes, and their sex determination was suggested to be highly variable. Here, we determined X-linked genes in the common sandfish (Scincus scincus) and demonstrate that skinks have shared the same homologous XX/XY sex chromosomes across their wide phylogenetic spectrum for at least 85 million years, approaching the age of the highly differentiated ZZ/ZW sex chromosomes of birds and advanced snakes. Skinks thus demonstrate that even poorly differentiated sex chromosomes can be evolutionarily stable. The conservation of sex chromosomes across skinks allows us to introduce the first molecular sexing method widely applicable in this group.

• Keywords
• genome, molecular sexing, qPCR, sex chromosomes, sex determination, vertebrates,
• MeSH
• Sex Determination Analysis MeSH
• Phylogeny MeSH
• Snakes MeSH
• Lizards * genetics   MeSH
• Sex Chromosomes * genetics   MeSH
• Sex Determination Processes MeSH
• Animals MeSH
• Check Tag
• Animals MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH

Dragon lizards (Squamata: Agamidae) comprise about 520 species in six subfamilies distributed across Asia, Australasia and Africa. Only five species are known to have sex chromosomes. All of them possess ZZ/ZW sex chromosomes, which are microchromosomes in four species from the subfamily Amphibolurinae, but much larger in Phrynocephalus vlangalii from the subfamily Agaminae. In most previous studies of these sex chromosomes, the focus has been on Australian species from the subfamily Amphibolurinae, but only the sex chromosomes of the Australian central bearded dragon (Pogona vitticeps) are well-characterized cytogenetically. To determine the level of synteny of the sex chromosomes of P. vitticeps across agamid subfamilies, we performed cross-species two-colour FISH using two bacterial artificial chromosome (BAC) clones from the pseudo-autosomal regions of P. vitticeps. We mapped these two BACs across representative species from all six subfamilies as well as two species of chameleons, the sister group to agamids. We found that one of these BAC sequences is conserved in macrochromosomes and the other in microchromosomes across the agamid lineages. However, within the Amphibolurinae, there is evidence of multiple chromosomal rearrangements with one of the BACs mapping to the second-largest chromosome pair and to the microchromosomes in multiple species including the sex chromosomes of P. vitticeps. Intriguingly, no hybridization signal was observed in chameleons for either of these BACs, suggesting a likely agamid origin of these sequences. Our study shows lineage-specific evolution of sequences/syntenic blocks and successive rearrangements and reveals a complex history of sequences leading to their association with important biological processes such as the evolution of sex chromosomes and sex determination.

• Keywords
• BACs, FISH, agamid lizards, evolution, sex chromosomes, synteny,
• MeSH
• Sex Determination Analysis methods   MeSH
• Cytogenetics methods   MeSH
• Snakes genetics   growth & development   MeSH
• Lizards genetics   growth & development   MeSH
• Karyotyping MeSH
• Evolution, Molecular * MeSH
• Sex Chromosomes genetics   MeSH
• Sex Determination Processes genetics   MeSH
• Synteny genetics   MeSH
• Chromosomes, Artificial, Bacterial genetics   MeSH
• Animals MeSH
• Check Tag
• Female MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH

Lacertid lizards are a widely radiated group of squamate reptiles with long-term stable ZZ/ZW sex chromosomes. Despite their family-wide homology of Z-specific gene content, previous cytogenetic studies revealed significant variability in the size, morphology, and heterochromatin distribution of their W chromosome. However, there is little evidence about the accumulation and distribution of repetitive content on lacertid chromosomes, especially on their W chromosome. In order to expand our knowledge of the evolution of sex chromosome repetitive content, we examined the topology of telomeric and microsatellite motifs that tend to often accumulate on the sex chromosomes of reptiles in the karyotypes of 15 species of lacertids by fluorescence in situ hybridization (FISH). The topology of the above-mentioned motifs was compared to the pattern of heterochromatin distribution, as revealed by C-banding. Our results show that the topologies of the examined motifs on the W chromosome do not seem to follow a strong phylogenetic signal, indicating independent and species-specific accumulations. In addition, the degeneration of the W chromosome can also affect the Z chromosome and potentially also other parts of the genome. Our study provides solid evidence that the repetitive content of the degenerated sex chromosomes is one of the most evolutionary dynamic parts of the genome.

• Keywords
• C-banding, FISH, GATA, evolution, heterochromatin, karyotype, microsatellites, sex chromosomes, telomeres,
• MeSH
• Chromosomes genetics   MeSH
• Species Specificity MeSH
• Phylogeny MeSH
• Heterochromatin genetics   ultrastructure   MeSH
• In Situ Hybridization, Fluorescence MeSH
• Lizards genetics   MeSH
• Karyotype MeSH
• Microsatellite Repeats genetics   MeSH
• Evolution, Molecular * MeSH
• Nucleotide Motifs MeSH
• Sex Chromosomes genetics   MeSH
• Chromosome Banding MeSH
• Repetitive Sequences, Nucleic Acid MeSH
• Telomere genetics   MeSH
• Animals MeSH
• Check Tag
• Male MeSH
• Female MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Comparative Study MeSH
• Names of Substances
• Heterochromatin MeSH

Chameleons are well-known, highly distinctive lizards characterized by unique morphological and physiological traits, but their karyotypes and sex determination system have remained poorly studied. We studied karyotypes in six species of Madagascan chameleons of the genus Furcifer by classical (conventional stain, C-banding) and molecular (comparative genomic hybridization, in situ hybridization with rDNA, microsatellite, and telomeric sequences) cytogenetic approaches. In contrast to most sauropsid lineages, the chameleons of the genus Furcifer show chromosomal variability even among closely related species, with diploid chromosome numbers varying from 2n = 22 to 2n = 28. We identified female heterogamety with cytogenetically distinct Z and W sex chromosomes in all studied species. Notably, multiple neo-sex chromosomes in the form Z1Z1Z2Z2/Z1Z2W were uncovered in four species of the genus (F. bifidus, F. verrucosus, F. willsii, and previously studied F. pardalis). Phylogenetic distribution and morphology of sex chromosomes suggest that multiple sex chromosomes, which are generally very rare among vertebrates with female heterogamety, possibly evolved several times within the genus Furcifer. Although acrodontan lizards (chameleons and dragon lizards) demonstrate otherwise notable variability in sex determination, it seems that female heterogamety with differentiated sex chromosomes remained stable in the chameleons of the genus Furcifer for about 30 million years.

• Keywords
• Comparative genome hybridization (CGH), Madagascar, evolution, fluorescence in situ hybridization (FISH), heterochromatin, karyotype, microsatellites, rDNA, sex chromosomes, telomeres.,
• MeSH
• Phylogeny * MeSH
• Lizards classification   genetics   MeSH
• Karyotype * MeSH
• Evolution, Molecular * MeSH
• Sex Chromosomes * MeSH
• Sex Determination Processes * MeSH
• Animals MeSH
• Check Tag
• Male MeSH
• Female MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH

Most pleurodont lizard families (anoles, iguanas and their relatives), with the exception of the basilisks and casquehead lizards (family Corytophanidae), share homologous XX/XY sex chromosomes, syntenic with chicken chromosome 15. Here, we used a suite of methods (i.e. RADseq, RNAseq and qPCR) to identify corytophanid sex chromosomes for the first time. We reveal that all examined corytophanid species have partially degenerated XX/XY sex chromosomes, syntenic with chicken chromosome 17. Transcriptomic analyses showed that the expression of X-linked genes in the corytophanid, Basiliscus vittatus, is not balanced between the sexes, which is rather exceptional under male heterogamety, and unlike the dosage-balanced sex chromosomes in other well-studied XX/XY systems, including the green anole, Anolis carolinensis. Corytophanid sex chromosomes may represent a rare example of a turnover away from stable, differentiated sex chromosomes. However, because of poor phylogenetic resolution among pleurodont families, we cannot reject the alternative hypothesis that corytophanid sex chromosomes evolved independently from an unknown ancestral system.

• Keywords
• RADseq, RNAseq, dosage compensation, reptiles, sex chromosomes, sex determination,
• MeSH
• Phylogeny MeSH
• Genes, X-Linked MeSH
• Lizards * MeSH
• Iguanas * MeSH
• Evolution, Molecular MeSH
• Sex Chromosomes MeSH
• Animals MeSH
• Check Tag
• Male MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Research Support, U.S. Gov't, Non-P.H.S. MeSH