title: RNA-seq analysis of the gonadal transcriptome during Alligator mississippiensis temperature-dependent sex determination and differentiation.
abstract: BACKGROUND
  The American alligator (Alligator mississippiensis) displays temperature-dependent sex determination (TSD), in which incubation temperature during embryonic development determines the sexual fate of the individual. However, the molecular mechanisms governing this process remain a mystery, including the influence of initial environmental temperature on the comprehensive gonadal gene expression patterns occurring during TSD.
  
  
  RESULTS
  Our characterization of transcriptomes during alligator TSD allowed us to identify novel candidate genes involved in TSD initiation. High-throughput RNA sequencing (RNA-seq) was performed on gonads collected from A. mississippiensis embryos incubated at both a male and a female producing temperature (33.5 °C and 30 °C, respectively) in a time series during sexual development. RNA-seq yielded 375.2 million paired-end reads, which were mapped and assembled, and used to characterize differential gene expression. Changes in the transcriptome occurring as a function of both development and sexual differentiation were extensively profiled. Forty-one differentially expressed genes were detected in response to incubation at male producing temperature, and included genes such as Wnt signaling factor WNT11, histone demethylase KDM6B, and transcription factor C/EBPA. Furthermore, comparative analysis of development- and sex-dependent differential gene expression revealed 230 candidate genes involved in alligator sex determination and differentiation, and early details of the suspected male-fate commitment were profiled. We also discovered sexually dimorphic expression of uncharacterized ncRNAs and other novel elements, such as unique expression patterns of HEMGN and ARX. Twenty-five of the differentially expressed genes identified in our analysis were putative transcriptional regulators, among which were MYBL2, MYCL, and HOXC10, in addition to conventional sex differentiation genes such as SOX9, and FOXL2. Inferred gene regulatory network was constructed, and the gene-gene and temperature-gene interactions were predicted.
  
  
  CONCLUSIONS
  Gonadal global gene expression kinetics during sex determination has been extensively profiled for the first time in a TSD species. These findings provide insights into the genetic framework underlying TSD, and expand our current understanding of the developmental fate pathways during vertebrate sex determination.
authors.affiliation: Department of Basic Biology, Faculty of Life Science, SOKENDAI (Graduate University for Advanced Studies), 5-1 Higashiyama, Myodaiji, Okazaki, Aichi, 444-8787, Japan. ryoheiy@nibb.ac.jp.
authors.forename: Ryohei
authors.initials: R
authors.lastname: Yatsu
authors.affiliation: Department of Basic Biology, Faculty of Life Science, SOKENDAI (Graduate University for Advanced Studies), 5-1 Higashiyama, Myodaiji, Okazaki, Aichi, 444-8787, Japan. miyagawa@nibb.ac.jp.|Okazaki Institute for Integrative Bioscience, National Institute for Basic Biology, National Institutes of Natural Sciences, 5-1 Higashiyama, Myodaiji, Okazaki, Aichi, 444-8787, Japan. miyagawa@nibb.ac.jp.
authors.forename: Shinichi
authors.initials: S
authors.lastname: Miyagawa
authors.affiliation: Department of Obstetrics and Gynecology, Medical University of South Carolina, and Marine Biomedicine and Environmental Science Center, Hollings Marine Laboratory, Charleston, SC, 29412, USA. kohnos@gmail.com.
authors.forename: Satomi
authors.initials: S
authors.lastname: Kohno
authors.affiliation: Department of Obstetrics and Gynecology, Medical University of South Carolina, and Marine Biomedicine and Environmental Science Center, Hollings Marine Laboratory, Charleston, SC, 29412, USA. benbparrott@gmail.com.
authors.forename: Benjamin B
authors.initials: BB
authors.lastname: Parrott
authors.affiliation: National Institute for Basic Biology, National Institutes of Natural Sciences, 38 Nishigonaka, Myodaiji, Okazaki, Aichi, 444-8585, Japan. kyamaguc@nibb.ac.jp.
authors.forename: Katsushi
authors.initials: K
authors.lastname: Yamaguchi
authors.affiliation: Department of Basic Biology, Faculty of Life Science, SOKENDAI (Graduate University for Advanced Studies), 5-1 Higashiyama, Myodaiji, Okazaki, Aichi, 444-8787, Japan. ogino@nibb.ac.jp.|Okazaki Institute for Integrative Bioscience, National Institute for Basic Biology, National Institutes of Natural Sciences, 5-1 Higashiyama, Myodaiji, Okazaki, Aichi, 444-8787, Japan. ogino@nibb.ac.jp.
authors.forename: Yukiko
authors.initials: Y
authors.lastname: Ogino
authors.affiliation: Center for Bioscience Research and Education, Utsunomiya University, 350 Mine-machi, Utsunomiya, Tochigi, 321-8505, Japan. h-miya@cc.utsunomiya-u.ac.jp.
authors.forename: Hitoshi
authors.initials: H
authors.lastname: Miyakawa
authors.affiliation: Innovative Health Applications, Kennedy Space Center, Merritt Island, FL, 32899, USA. Russell.h.lowers@nasa.gov.
authors.forename: Russell H
authors.initials: RH
authors.lastname: Lowers
authors.affiliation: Department of Basic Biology, Faculty of Life Science, SOKENDAI (Graduate University for Advanced Studies), 5-1 Higashiyama, Myodaiji, Okazaki, Aichi, 444-8787, Japan. shige@nibb.ac.jp.|National Institute for Basic Biology, National Institutes of Natural Sciences, 38 Nishigonaka, Myodaiji, Okazaki, Aichi, 444-8585, Japan. shige@nibb.ac.jp.
authors.forename: Shuji
authors.initials: S
authors.lastname: Shigenobu
authors.affiliation: Department of Obstetrics and Gynecology, Medical University of South Carolina, and Marine Biomedicine and Environmental Science Center, Hollings Marine Laboratory, Charleston, SC, 29412, USA. kohno@musc.edu.
authors.forename: Louis J
authors.initials: LJ
authors.lastname: Guillette
authors.affiliation: Department of Basic Biology, Faculty of Life Science, SOKENDAI (Graduate University for Advanced Studies), 5-1 Higashiyama, Myodaiji, Okazaki, Aichi, 444-8787, Japan. taisen@nibb.ac.jp.|Okazaki Institute for Integrative Bioscience, National Institute for Basic Biology, National Institutes of Natural Sciences, 5-1 Higashiyama, Myodaiji, Okazaki, Aichi, 444-8787, Japan. taisen@nibb.ac.jp.
authors.forename: Taisen
authors.initials: T
authors.lastname: Iguchi
mesh_terms: D000489:Alligators and Crocodiles / Q000235:genetics* / Q000502:physiology; D000818:Animals; D005260:Female; D018507:Gene Expression Regulation, Developmental; D059014:High-Throughput Nucleotide Sequencing; D008297:Male; D012313:RNA / Q000235:genetics*; D019849:Sex Determination Processes / Q000235:genetics* / Q000502:physiology; D012733:Sex Differentiation / Q000235:genetics* / Q000502:physiology; D013696:Temperature; D059467:Transcriptome / Q000235:genetics*
chemical_list: D012313:RNA
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