Figure 1.
Genes involved in sexual development.
Table 1.
Male sex-determining/differentiation gene inventory.
Table 2.
Female sex-determining/differentiation gene inventory.
Figure 2.
Expression of male development genes.
Expression levels of male sex-determining/differentiation genes in L. menadoensis liver and testis transcriptomes. Values are expressed as FPKM (Fragments Per Kilobase of exon per Million sequenced fragments). A) genes highly expressed in testis; B) genes poorly expressed in testis. The expression levels of some housekeeping genes (not represented) were also analysed: PGK 96.95 (liver), 342.41 (testis); RPS27a 152.59 (liver), 128.43 (testis); RPL19 744.01 (liver), 64.89 (testis); RPL11 457.35 (liver), 282.59 (testis); RPL32 629.83 (liver), 373.75 (testis); HSPCB 507.99 (liver), 1213.75 (testis). Threshold value = 1. * Expression level assessed on L. chalumnae orthologue.
Figure 3.
Conserved micro-synteny and structure of the DMRT1 genomic locus and transcripts.
A) Genomic representation of DMRT1 on scaffold JH127237 of L. chalumnae. Grey box corresponds to gene. Small boxes and V signs represent the intron/exon map. B) Transcript representation of DMRT1 in L. menadoensis and L. chalumnae. Boxes: exons; V signs: introns; white box: DM domain; light grey box: 3′UTR; dashed box: putative transposable element contained in the 3′UTR. Dotted boxes represent missing exons in the ENSEMBL transcript prediction. C) Micro-syntenic conservation of genomic blocks containing the DMRT1 gene. White pentagons represent DMRT1 genes. The pentagon tip points to the relative gene orientation. Numbers near the pentagons stand for gene size expressed as kb, numbers on lines represent intergene distance expressed as kb. ENSEMBL data: H. sa (Homo sapiens), M. mu (Mus musculus), O. an (Ornithorhynchus anatinus), G. ga (Gallus gallus), A. ca (Anolis carolinensis), L. ch (Latimeria chalumnae), D. re (Danio rerio), T. ru (Takifugu rubripes). L. chalumnae DMRT1 position was clarified using the L. menadoensis transcript, by integrating the L. chalumnae ENSLACT00000015034 coordinates. *In O. anatinus DMRT1 gene size was defined by comparison with other species. **Values obtained in G. gallus from the annotation of NC_006127.3 accession.
Figure 4.
Phylogenetic tree of SOX8, SOX9, and SOX10.
Phylogenetic analyses of vertebrate SOXE amino acid sequences. Midpoint rooting. Total characters: 592, constant: 164, parsimony non-informative: 77, parsimony informative: 351. Numbers close to nodes represent posterior probability in Bayesian Inference/bootstrap percentage in Maximum Parsimony. Danio rerio (SOX8: AAX73357.1; SOX9a: NP_571718.1; SOX9b: NP_571719.1; SOX10: AAK84872.1); Dicentrarchus labrax (SOX8: CBN81184.1; SOX9: CBN81190.1); Gallus gallus (SOX8: AAF73917.1; SOX9: BAA25296.1; SOX10: AAD38050.2); Homo sapiens (SOX8: AAH31797.1; SOX9: CAA86598.1; SOX10: CAG30470.1); Latimeria chalumnae (SOX8: ENSLACP00000018883; SOX9: ENSLACP00000021343; SOX10: ENSLACP00000004990); Latimeria menadoensis (SOX9, SOX10: this study); Mus musculus (SOX8: AAF35837.1; SOX9: AAH23953.1; SOX10: NP_035567.1); Oryzias latipes (SOX8: NP_001158342.1; SOX9a: AAX62152.1; SOX9b: AAX62151.1); Salmo salar (SOX8: ABC24688.1; SOX9: ACN10975.1); Scyliorhinus canicula (SOX9: ABY71239.1); Trachemys scripta (SOX8: AAP59791.1; SOX9: ACG70782.1; SOX10: ENSLACP00000004990); Xenopus laevis (SOX8: AAI69525.1; SOX9: NP_001084276; SOX10: NP_001082358.1). *Only a partial SOX8 sequence, perfectly matching the ENSEMBL prediction of the L. chalumnae SOX8 gene, was retrieved in the transcriptome assembly of L. menadoensis.
Figure 5.
Analysis of micro-syntenic conservation in FGF9, FGF16 and FGF20 blocks.
Micro-syntenic conservation of genomic regions containing the FGF9, FGF20 and FGF16 genes. White pentagons represent FGF genes. The pentagon tip points to the relative gene orientation. The grey mark on the top third of the figure indicates a EFHA1 putative sequence of Latimeria chalumnae. Numbers near pentagons stand for gene size expressed as kb, numbers on lines represent intergene distance expressed as kb. ENSEMBL data: H. sa (Homo sapiens), G. ga (Gallus gallus), A. ca (Anolis carolinensis), X. tr (Xenopus tropicalis), L. ch (Latimeria chalumnae), D. re (Danio rerio), T. ru (Takifugu rubripes). Syntenic blocks for FGF20 in L. chalumnae and X. tropicalis, and FGF16 in A. carolinensis are split between two different scaffolds. The ZDHHC15 genes belonging to the syntenic block of FGF16 in H. sapiens and X. tropicalis lie on the same chromosome or scaffold, but are far removed from the genomic locus of FGF16 and ATRX. *Genes missing in the ENSEMBL prediction.
Figure 6.
Phylogenetic tree of FGF9, FGF16, and FGF20.
Phylogenetic analysis of amino acid sequences of the vertebrate FGF9/16/20. Midpoint rooting. Total characters: 237, constant: 91, parsimony non-informative: 35, parsimony informative: 111. Numbers close to nodes represent posterior probability in Bayesian Inference/bootstrap percentage in Maximum Parsimony. Danio rerio (FGF16: ENSDART00000061928; FGF20a: NP_001032180.1; FGF20b: NP_001034261.1); Gallus gallus (FGF9: NP_989730.1; FGF16; NP_001038115.1; FGF20: XP_426335.2); Homo sapiens (FGF9: NP_002001.1; FGF16: NP_003859.1; FGF20: NP_062825.1); Latimeria chalumnae (FGF9: manually inferred from JH128123; FGF16: ENSLACT00000011509; FGF20: ENSLACT00000014939); Mus musculus (FGF9: ADL60500.1; FGF16: BAB16405.1; FGF20: NP_085113.2); Oryzias latipes (FGF16: ENSORLT00000007651; FGF20a: ENSORLT00000012578; FGF20b: ENSORLT00000025767); Takifugu rubripes (FGF16: ENSTRUT00000021181; FGF20(1): ENSTRUT00000008788; FGF20(2): ENSTRUT00000039390); Xenopus tropicalis (FGF9: XP_002938621.1; FGF16: ENSXETT00000009790; FGF20: NP_001137399.1). Latimeria menadoensis is missing in this analysis because FGF9 and FGF20 are poorly or not expressed in the transcriptomes.
Figure 7.
Phylogenetic tree of GSDF, AMH, and inhibin-α.
Phylogenetic analysis of amino acid sequences of vertebrate GSDF, inhibin-α and AMH. Total characters: 849, constant: 84, parsimony non-informative: 225, parsimony informative: 540. Outgroup: human glial-derived nerve growth factor (GDNF). Numbers close to nodes represent posterior probabilities in Bayesian Inference/bootstrap percentage in Maximum Parsimony. Anolis carolinensis (inhibin-α: ENSACAT00000014331); Danio rerio (GSDFa: AEL99890.1; GSDFb: AEL99889.1; AMH: NP_001007780.1; inhibin-α: ENSDART00000057348); Gallus gallus (AMH: NP_990361.1; inhibin-α: NP_001026428.1); Gasterosteus aculeatus (GSDF: ENSGACT00000021595; inhibin-α: ENSGACT00000018909); Homo sapiens (AMH AAC25614.1; GDNF: NP_000505.1); Latimeria chalumnae (inhibin-α: ENSLACT00000017535); Latimeria menadoensis (GSDF, AMH this study); Mus musculus (AMH: AAI50478.1; inhibin-α: AAH56627.1); Oreochromis niloticus (GSDF: BAJ78985.1); Oryzias latipes (GSDF: NP_001171213.1); Oncorhynchus mykiss (GSDF: ABF48201.1); Takifugu rubripes (GSDF: ENSTRUT00000036269; AMH: ENSTRUT00000045919); Xenopus laevis (inhibin-α: NP_001106349.1). The reliability of L. menadoensis CDSs is supported by the same sequence resulting from application of two different assembly procedures.
Figure 8.
Multiple alignment of the TGF-β domain in GSDF, AMH, and inhibin-α.
Conserved amino acids of the cysteine knot are boxed. Anolis carolinensis (inhibin-α: ENSACAT00000014331); Danio rerio (GSDFa: AEL99890.1, GSDFb: AEL99889.1; AMH: NP_001007780.1; inhibin-α: ENSDART00000057348); Gallus gallus (AMH: NP_990361.1; inhibin-α: NP_001026428.1); Gasterosteus aculeatus (GSDF: ENSGACT00000021595; inhibin-α: ENSGACT00000018909); Homo sapiens (AMH: AAC25614.1; GDNF: NP_000505.1); Latimeria chalumnae (inhibin-α: ENSLACT00000017535); Latimeria menadoensis (this study); Mus musculus (AMH: AAI50478.1; inhibin-α: AAH56627.1); Oreochromis niloticus (GSDF: BAJ78985.1); Oryzias latipes (GSDF: NP_001171213.1); Oncorhynchus mykiss (GSDF: ABF48201.1); Takifugu rubripes (GSDF: ENSTRUT00000036269; AMH: ENSTRUT00000045919); Xenopus laevis (inhibin-α: NP_001106349.1). The reliability of L. menadoensis CDSs is supported by the same sequence resulting from application of two different assembly procedures.
Figure 9.
Expression of female development genes.
Expression of female-determining/differentiation genes in L. menadoensis liver and testis transcriptomes. Expression levels are reported as FPKM (Fragments Per Kilobase of exon per Million sequenced fragments). The expression levels of some housekeeping genes were also analysed: PGK 96.95 (liver), 342.41 (testis); RPS27a 152.59 (liver), 128.43 (testis); RPL19 744.01 (liver) 64.89 (testis); RPL11 457.35 (liver), 282.59 (testis); RPL32 629.83 (liver), 373.75 (testis); HSPCB 507.99 (liver), 1213.75 (testis). Threshold value = 1. * Expression level assessed on L. chalumnae orthologue.