Amniotes possess variability in sex determining mechanisms, however, this diversity is still only partially known throughout the clade and sex determining systems still remain unknown even in such a popular and distinctive lineage as chameleons (Squamata: Acrodonta: Chamaeleonidae). Here, we present evidence for female heterogamety in this group. The Malagasy giant chameleon (Furcifer oustaleti) (chromosome number 2n = 22) possesses heteromorphic Z and W sex chromosomes with heterochromatic W. The panther chameleon (Furcifer pardalis) (2n = 22 in males, 21 in females), the second most popular chameleon species in the world pet trade, exhibits a rather rare Z1Z1Z2Z2/Z1Z2W system of multiple sex chromosomes, which most likely evolved from W-autosome fusion. Notably, its neo-W chromosome is partially heterochromatic and its female-specific genetic content has expanded into the previously autosomal region. Showing clear evidence for genotypic sex determination in the panther chameleon, we resolve the long-standing question of whether or not environmental sex determination exists in this species. Together with recent findings in other reptile lineages, our work demonstrates that female heterogamety is widespread among amniotes, adding another important piece to the mosaic of knowledge on sex determination in amniotes needed to understand the evolution of this important trait.

] Department of Ecology Faculty of Science Charles University Prague Prague 128 44 Czech Republic [2] Institute of Animal Physiology and Genetics The Academy of Sciences of the Czech Republic Liběchov 277 21 Czech Republic

Department of Ecology Faculty of Science Charles University Prague Prague 128 44 Czech Republic

Zobrazit více v PubMed

Uetz P. & Hošek J. (eds). The reptile database. (2014) Available at: http://www.reptile-database.org (Accessed: 23th January 2015).

Tolley K. A., Townsend T. M. & Vences M. Large-scale phylogeny of chameleons suggests African origins and Eocene diversification. Proc. R. Soc. B. 280, 20130184 (2013). PubMed PMC

Pokorná M. & Kratochvíl L. Phylogeny of sex-determining mechanisms in squamate reptiles: are sex chromosomes an evolutionary trap? Zool. J. Linn. Soc. 156, 168–183 (2009).

Johnson Pokorná M. & Kratochvíl L. What was the ancestral sex-determining mechanism in amniote vertebrates? Biol. Rev. (2014). In press. 10.1111/brv.12156. PubMed DOI

Ezaz T. PubMed

Harlow P. S. in Temperature-Dependent Sex Determination in Vertebrates (eds. Valenzuela N. & Lance V. ), 42–52 (Smithsonian Institution Scholarly Press: Washington, DC, 2004).

Gamble T., Geneva A. J., Glor R. E. & Zarkower D. PubMed PMC

Rovatsos M., Altmanová M., Pokorná M. & Kratochvíl L. Conserved sex chromosomes across adaptively radiated PubMed

Rovatsos M., Pokorná M., Altmanová M. & Kratochvíl L. Cretaceous park of sex determination: sex chromosomes are conserved across iguanas. Biol. Lett. 10, 20131093 (2014). PubMed PMC

Viets B. E., Ewert M. A., Talent L. G. & Nelson C. E. Sex-determining mechanisms in squamate reptiles. J. Exp. Zool. 270, 45–56 (1994).

Andrews R. M. Incubation temperature and sex ratio of the veiled chameleon (

Olmo E. & Signorino G. Chromorep: A reptile chromosomes database. Chamaeleonidae. (2005) Available at: http://ginux.univpm.it/scienze/chromorep/3.2.1.1.2%20Chamaeleonidae.html (Accessed: 23th January 2015).

Bourgat R. M. Biogeographical interest of

Cheke A. & Hume J. Lost Land of the Dodo. (T & A D Poyser: London, 2008).

Cole N. A Field Guide to the Reptiles and Amphibians of Mauritius. (Mauritian Wildlife Foundation: Vacoas, Mauritius, 2009).

Grbic D. PubMed DOI PMC

Jenkins R. K. B.

Jenkins R. K. B.

Bourgat R. M. Caryotypes des caméléons malgaches et systématique: Synthèse des données acquises. Annales de l´Université de Madagascar, Série Sciences de la Nature et Mathématiques 9, 77–85 (1972).

Matthey R. & van Brink J. M. Note préliminaire sur la cytologie chromosomique comparée des Caméléons. Rev. Suisse Zool. 63, 241–246 (1956).

Matthey R. Cytologie comparée et taxonomie des Chamaeleontidae (Reptilia – Lacertilia). Rev. Suisse Zool. 64, 709–732 (1957).

Tolley K. A. & Herrel A. The Biology of Chameleons. (University of California Press: California, Berkeley and Los Angeles, 2013).

Nagy Z. T., Sonet G., Glaw F. & Vences M. First large-scale DNA barcoding assessment of reptiles in the biodiversity hotspot of Madagascar, based on newly designed COI primers. PLoS One 7, e34506 (2012). PubMed PMC

Johnson Pokorná M., Rovatsos M. & Kratochvíl L. Sex chromosomes and karyotype of the (nearly) mythical creature, the Gila monster, PubMed PMC

Gamble T. PubMed DOI

Pokorná M., Rens W., Rovatsos M. & Kratochvíl L. A ZZ/ZW sex chromosome system in the thick-tailed gecko ( PubMed

Odierna G., Olmo E. & Cobror O. C-band variability in some Lacertidae (Sauria, Reptilia). Experientia 41, 944–946 (1985).

Koubová M. PubMed

Oguiura N., Ferrarezzi H. & Batistic R. F. Cytogenetics and molecular data in snakes: A phylogenetic approach. Cytogenet. Genome Res. 127, 128–142 (2009). PubMed

Mezzasalma M.

Ezaz T., Sarre S. D., O’Meally D., Graves J. & Georges A. Sex chromosome evolution in lizards: independent origins and rapid transitions. Cytogenet. Genome Res. 127, 249–260 (2009). PubMed

Young M. J., O’Meally D., Sarre S. D., Georges A. & Ezaz T. Molecular cytogenetic map of the central bearded dragon, PubMed

Schmid M., Steinlein C., Haaf T. & Mijares-Urrutia A. Nascent ZW sex chromosomes in PubMed

Rovatsos M. T. PubMed

Rovatsos M. T. PubMed

Bachtrog D., Hom E., Wong K. M., Maside X. & de Jong P. Genomic degradation of a young Y chromosome in PubMed PMC

Zhou Q. & Bachtrog D. Sex-specific adaptation drives early sex chromosome evolution in PubMed PMC

Pokorná M., Altmanová M. & Kratochvíl L. Multiple sex chromosomes in the light of female meiotic drive in amniote vertebrates. Chromosome Res. 22, 35–44 (2014). PubMed

Srikulnath K. PubMed

Srikulnath K. PubMed PMC

Rojo V. PubMed

Srikulnath K., Uno Y., Nishida C. & Matsuda Y. Karyotype evolution in monitor lizards: cross-species chromosome mapping of cDNA reveals highly conserved synteny and gene order in the Toxicofera clade. Chromosome Res. 21, 805–819 (2013). PubMed

Meyne J. PubMed

Ruiz-Herrera A., Nergadze S. G., Santagostino M. & Giulotto E. Telomeric repeats far from the ends: mechanisms of origin and role in evolution. Cytogenet. Genome Res. 122, 219–228 (2008). PubMed

Hebert P. D. N. & Gregory T. R. The promise of DNA barcoding for taxonomy. Syst. Biol. 54, 852–859 (2005). PubMed

Hawlitschek O., Nagy Z. T., Berger J. & Glaw F. Reliable DNA barcoding performance proved for species and island populations of comoran squamate reptiles. PLoS One 8, e73368 (2013). PubMed PMC

Jeong T. J. PubMed

Murphy R. W.

Glaw F.

Glaw F. & Vences M. A Field Guide to the Amphibians and Reptiles of Madagascar. (Vences & Glaw Verlags GbR: Köln, 2007).

Gamble T. A review of sex determining mechanisms in geckos (Gekkota: Squamata). Sex Dev. 4, 88–103 (2010). PubMed PMC

Hebert P. D. N., Cywinska A., Ball S. L. & deWaard J. R. Biological identifications through DNA barcodes. Proc. Biol. Sci. 270, 313–321 (2003). PubMed PMC

Folmer O. PubMed

Thompson J. D., Higgins D. G. & Gibson T. J. CLUSTAL W: improving the sensitivity of progressive multiple sequence alignment through sequence weighting, position-specific gap penalties and weight matrix choice. Nucleic Acids Res. 22, 4673–4680 (1994). PubMed PMC

Hall T. BioEdit: a user-friendly biological sequence alignment editor and analysis program for Windows 95/98/NT. Nucleic Acids Symp. Ser. 41, 95–98 (1999).

Librado P. & Rozas J. DnaSP v5: A software for comprehensive analysis of DNA polymorphism data. Bioinformatics 25, 1451–1452 (2009). PubMed

Tamura K., Stecher G., Peterson D., Filipski A. & Kumar S. MEGA6: Molecular evolutionary genetics analysis version 6.0. Mol. Biol. Evol. 30, 2725–2729 (2013). PubMed PMC

Sumner A. T. A simple technique for demonstrating centromeric heterochromatin. Exp. Cell Res. 75, 304–306 (1972). PubMed

Ijdo J. W., Wells R. A., Baldini A. & Reeders S. T. Improved telomere detection using a telomere repeat probe (TTAGGG) PubMed PMC

Endow S. A. Polytenization of the ribosomal genes on the X and Y chromosomes of PubMed PMC

Heteromorphic ZZ/ZW sex chromosomes sharing gene content with mammalian XX/XY are conserved in Madagascan chameleons of the genus Furcifer

Madagascar Leaf-Tail Geckos (Uroplatus spp.) Share Independently Evolved Differentiated ZZ/ZW Sex Chromosomes

ZZ/ZW Sex Chromosomes in the Madagascar Girdled Lizard, Zonosaurus madagascariensis (Squamata: Gerrhosauridae)

Cytogenetic Analysis of the Members of the Snake Genera Cylindrophis, Eryx, Python, and Tropidophis

Sex chromosome evolution among amniotes: is the origin of sex chromosomes non-random?

Are Geckos Special in Sex Determination? Independently Evolved Differentiated ZZ/ZW Sex Chromosomes in Carphodactylid Geckos

Cytogenetic Evidence for Sex Chromosomes and Karyotype Evolution in Anguimorphan Lizards

With or Without W? Molecular and Cytogenetic Markers are Not Sufficient for Identification of Environmentally-Induced Sex Reversal in the Bearded Dragon

Poorly differentiated XX/XY sex chromosomes are widely shared across skink radiation

Interstitial Telomeric Repeats Are Rare in Turtles

Sex is determined by XX/XY sex chromosomes in Australasian side-necked turtles (Testudines: Chelidae)

ZZ/ZW Sex Determination with Multiple Neo-Sex Chromosomes is Common in Madagascan Chameleons of the Genus Furcifer (Reptilia: Chamaeleonidae)

Cytogenetic Analysis Did Not Reveal Differentiated Sex Chromosomes in Ten Species of Boas and Pythons (Reptilia: Serpentes)

Little evidence for switches to environmental sex determination and turnover of sex chromosomes in lacertid lizards

Turtles of the genera Geoemyda and Pangshura (Testudines: Geoemydidae) lack differentiated sex chromosomes: the end of a 40-year error cascade for Pangshura

Evolutionary insight on localization of 18S, 28S rDNA genes on homologous chromosomes in Primates genomes

Evolutionary Dynamics of the W Chromosome in Caenophidian Snakes

Evolution of Karyotypes in Chameleons

Evolutionary stability of sex chromosomes in snakes