title: Isolation of doublesex- and mab-3-related transcription factor 6 and its involvement in spermatogenesis in tilapia.
abstract: The dmrt6 gene has been isolated from tetrapods and recently from a coelacanth, Latimeria chalumnae. Its evolutionary history and exact function remain unclear. In the present study, dmrt6 was isolated from Perciformes (five cichlids and stickleback), Siluriformes (southern catfish), and Lepisosteiformes (spotted gar). Syntenic and phylogenetic analyses indicated that dmrt6 experienced gene transposition after the divergence of teleosts from other bony fish as gene loci surrounding dmrt6 were conserved among teleosts (but was completely different from gene loci surrounding dmrt6 in tetrapods and spotted gar), while these gene loci were conserved among nonteleost species. Real-time PCR and in situ hybridization revealed that dmrt6 was highly expressed in the XY gonads from 90 days after hatching (dah) onward and was observed exclusively in spermatocytes of the testes in tilapia. Dmrt6 knockout by CRISPR/Cas9 resulted in fewer spermatocytes, down-regulated Cyp11b2 in testes, and consequently produced a lower level of serum 11-ketotestosterone (11-KT) in Dmrt6-deficient XY fish compared with the XY control at 120 dah. From 150 to 180 dah, spermatogenesis gradually recovered, and cyp11b2 expression and serum 11-KT level were restored to the same levels as those of the XY control fish. In addition, a Dmrt6 mutation was observed in genomic DNA of sperm of G0 mutant fish and F1 fish. Taken together, our data suggest that dmrt6 also exists in bony fish. Its absence in most fish genomes was probably due to incomplete sequencing and/or secondary loss. The dmrt6 gene is highly expressed in spermatocytes and is involved in spermatogenesis in tilapia.
authors.affiliation: Key Laboratory of Freshwater Fish Reproduction and Development (Ministry of Education), Key Laboratory of Aquatic Science of Chongqing, School of Life Sciences, Southwest University, Chongqing, China.
authors.forename: Xianbo
authors.initials: X
authors.lastname: Zhang
authors.affiliation: Key Laboratory of Freshwater Fish Reproduction and Development (Ministry of Education), Key Laboratory of Aquatic Science of Chongqing, School of Life Sciences, Southwest University, Chongqing, China.
authors.forename: Hai
authors.initials: H
authors.lastname: Wang
authors.affiliation: Key Laboratory of Freshwater Fish Reproduction and Development (Ministry of Education), Key Laboratory of Aquatic Science of Chongqing, School of Life Sciences, Southwest University, Chongqing, China.
authors.forename: Minghui
authors.initials: M
authors.lastname: Li
authors.affiliation: Key Laboratory of Freshwater Fish Reproduction and Development (Ministry of Education), Key Laboratory of Aquatic Science of Chongqing, School of Life Sciences, Southwest University, Chongqing, China.
authors.forename: Yunying
authors.initials: Y
authors.lastname: Cheng
authors.affiliation: Key Laboratory of Freshwater Fish Reproduction and Development (Ministry of Education), Key Laboratory of Aquatic Science of Chongqing, School of Life Sciences, Southwest University, Chongqing, China.
authors.forename: Dongneng
authors.initials: D
authors.lastname: Jiang
authors.affiliation: Key Laboratory of Freshwater Fish Reproduction and Development (Ministry of Education), Key Laboratory of Aquatic Science of Chongqing, School of Life Sciences, Southwest University, Chongqing, China.
authors.forename: Lina
authors.initials: L
authors.lastname: Sun
authors.affiliation: Key Laboratory of Freshwater Fish Reproduction and Development (Ministry of Education), Key Laboratory of Aquatic Science of Chongqing, School of Life Sciences, Southwest University, Chongqing, China.
authors.forename: Wenjing
authors.initials: W
authors.lastname: Tao
authors.affiliation: Key Laboratory of Freshwater Fish Reproduction and Development (Ministry of Education), Key Laboratory of Aquatic Science of Chongqing, School of Life Sciences, Southwest University, Chongqing, China.
authors.forename: Linyan
authors.initials: L
authors.lastname: Zhou
authors.affiliation: Key Laboratory of Freshwater Fish Reproduction and Development (Ministry of Education), Key Laboratory of Aquatic Science of Chongqing, School of Life Sciences, Southwest University, Chongqing, China wangzj@swu.edu.cn.
authors.forename: Zhijian
authors.initials: Z
authors.lastname: Wang
authors.affiliation: Key Laboratory of Freshwater Fish Reproduction and Development (Ministry of Education), Key Laboratory of Aquatic Science of Chongqing, School of Life Sciences, Southwest University, Chongqing, China wdeshou@swu.edu.cn.
authors.forename: Deshou
authors.initials: D
authors.lastname: Wang
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D030801:Animals, Genetically Modified; D001483:Base Sequence; D003001:Cloning, Molecular; D008297:Male; D008969:Molecular Sequence Data; D010802:Phylogeny; D017386:Sequence Homology, Amino Acid; D013091:Spermatogenesis / Q000235:genetics*; D013737:Testis / Q000378:metabolism; D017210:Tilapia / Q000235:genetics* / Q000502:physiology; D014018:Tissue Distribution; D014157:Transcription Factors / Q000235:genetics / Q000302:isolation & purification* / Q000502:physiology*
chemical_list: D014157:Transcription Factors
keywords: CRISPR/Cas9; dmrt6; gene knockout; gene transposition; spermatogenesis; tilapia