Discovery of the youngest sex chromosomes reveals first case of convergent co-option of ancestral autosomes in turtles

Abstract

Most turtle species possess temperature-dependent sex determination (TSD), but genotypic sex determination (GSD) has evolved multiple times independently from the TSD ancestral condition. GSD in animals typically involves sex chromosomes, yet the sex chromosome system of only 9 out of 18 known GSD turtles has been characterized. Here, we combine comparative genome hybridization (CGH) and BAC clone fluorescent in situ hybridization (BAC FISH) to identify a macro-chromosome XX/XY system in the GSD wood turtle Glyptemys insculpta (GIN), the youngest known sex chromosomes in chelonians (8-20 My old). Comparative analyses show that GIN-X/Y is homologous to chromosome 4 of Chrysemys picta (CPI) painted turtles, chromosome 5 of Gallus gallus chicken, and thus to the X/Y sex chromosomes of Siebenrockiella crassicollis black marsh turtles. We tentatively assign the gene content of the mapped BACs from CPI chromosome 4 (CPI-4) to GIN-X/Y. Chromosomal rearrangements were detected in G. insculpta sex chromosome pair that co-localize with the male-specific region of GIN-Y and encompass a gene involved in sexual development (Wt1-a putative master gene in TSD turtles). Such inversions may have mediated the divergence of G. insculpta sex chromosome pair and facilitated GSD evolution in this turtle. Our results illuminate the structure, origin, and evolution of sex chromosomes in G. insculpta and reveal the first case of convergent co-option of an autosomal pair as sex chromosomes within chelonians.

Sex Reversal in Reptiles: Reproductive Oddity or Powerful Driver of Evolutionary Change?

“…1 ). Recapitulating this point, we observe that the 3 currently identified reptile candidate sex-determining genes are different in squamates ( NR5A1 P. vitticeps; Deakin et al [2016]) and testudines ( WT1 Glyptemys insculpta; Montiel et al [2016] and CIRBP Chelydra serpentina; Schroeder et al [2016] There has been significant concern in the literature about the future of TSD reptiles under climate scenarios with rapid warming events [Cotton and Wedekind, 2009;Sinervo et al, 2010;Boyle et al, 2014]. This is a legitimate concern and indeed the recent discovery of an increasing rate of sex reversal in wild bearded dragon populations suggests that climatic extremes have the potential to perturb even seemingly stable GSD systems [Holleley et al, 2015].…”

Cytogenetic Insights into the Evolution of Chromosomes and Sex Determination Reveal Striking Homology of Turtle Sex Chromosomes to Amphibian Autosomes

“…Cell Culture, Chromosome Preparation, and Chromosome Banding Chromosome preparations were obtained from fibroblast cell cultures established from embryonic or muscle tissue following standard procedures as previously described [Valenzuela, 2009;Badenhorst et al, 2013Badenhorst et al, , 2015Valenzuela et al, 2014;Montiel et al, 2016]. New samples were obtained from private collections or pet trade specimens.…”

“…Of the 18 turtles known to possess GSD, 10 have a cytogenetically characterized system of heterogamety ( figs. 1 , 6 ), 3 in the suborder Pleurodira which are all XX/XY [McBee et al, 1985;Martinez et al, 2008;Ezaz et al, 2009], and 7 in Cryptodira, 3 of which are ZZ/ZW [Sharma et al, 1975;Kawai et al, 2007;Badenhorst et al, 2013] and 4 are XX/XY [Bull et al, 1974;Carr and Bickham, 1981;Montiel et al, 2016]. Of these 10 cases, 3 involved microchromosomes and 7 macrochromosomes.…”

“…Of these 10 cases, 3 involved microchromosomes and 7 macrochromosomes. Relatively little is known about the molecular content of turtle sex chromosomes, although information is increasing [Kawagoshi et al, 2009[Kawagoshi et al, , 2012[Kawagoshi et al, , 2014Montiel et al 2016]. Molecular cytogenetic data from comparative genome hybridization (CGH) has been used to identify the extent of the male-or female-limited region of the turtle sex chromosomes [Ezaz et al, 2006;Kawai et al, 2007;Martinez et al, 2008;Badenhorst et al, 2013;Montiel et al, 2016] where recombination should be reduced or absent.…”

“…Of the 335 recognized turtle species [van Dijk et al, 2014], 87 have a known sex-determining mechanism (SDM) established based on either incubation experiments or cytogenetic evidence [Tree of Sex Consortium, 2014; Montiel et al, 2016]. Phylogenetic analyses recover temperature-dependent sex determination (TSD) as the ancestral SDM in turtles, and most evolutionary transitions involved the evolution of genotypic sex determination (GSD) [Janzen and Krenz, 2004;Valenzuela and Adams, 2011], with 2 potential reversals back to TSD inferred by maximum likelihood analysis [Valenzuela and Adams, 2011] ( fig.…”