The Whole Genome Of Triplophysa Tibetana And Its Adaptation To The Intense Ultraviolet Radiation Environment On The Tibetan Plateau

Strong ultraviolet radiation is one of the main characteristics of Tibetan Plateau,and the high-altitude adaptation has always been a multidisciplinary research hotspot.Recent studies of high-altitude adaptation mainly focused on terrestrial animals,while the research on the mechanism of plateau fishes and other aquatic animals adapt to strong ultraviolet radiation has not been reported.In conducting environmental surveys on the waters of the Tibet Plateau,we found that shallow water areas such as rivers and lakes on the Qinghai-Tibet Plateau have relatively high ultraviolet radiation intensity,and these areas are important habitats for Triplophysa fishes and the larvae and juveniles of Schizothoracinae fishes.How do high-altitude fishes adapt to strong ultraviolet radiation? In order to clarify this important and interesting scientific question,the indigenous fish Triplophysa tibetana was selected as the research object,and the exotic species Misgurnus anguillicaudatus was selected as a reference.Based on the whole genome of T.tibetana,we performed a comparative genome analysis and clarified the adaptation mechanism of high-altitude fishes to the intense ultraviolet radiation on the Tibetan Plateau.The main results as following:1.The sequence and chromosome-level assembly of T.tibetana using PacBio and Hi-C techniquesIn the present study,we used PacBio sequencing and the Hi-C technique to assemble the T.tibetana genome.A 652-Mb genome with 1,325 contigs,which had an N50 length of 3.1 Mb,was obtained.The 1,137 contigs were further assembled into 25 chromosomes.In total,24,372 protein-coding genes were predicted in the genome,and about 95% of the genes were functionally annotated via a search in public databases.This study obtained the first chromosome-level genomic resources of Triplophysa fishes,laying the foundation for further exploring the environmental adaptation of plateau fishes.2.Explore the mechanisms of adaptive evolution to the intense UV radiation of the Tibetan Plateau in T.tibetana based on comparative genomics analysesIn this study,we performed the whole genome comparative analyses of T.tibetana with other fish species which live in different aquatic environments,such as plateau,plateau-cave,plain,plain-cave,ocean-surface,and deep sea.The results showed that a total of 616 pairs of single-copy orthologous genes were identified in the T.tibetana and other 12 fish species.Further analyses showed that ASP had effects on melanogenesis by suppressing gene expression of MC1 R.CDS of ASP in T.tibetana showed gene deletion in 200-249(50 bp),when compared to that in Sinocyclocheilus anshuiensis and S.rhinocerous,which led to the frameshift mutation and then loss-of-function variants.GO and KEGG enrichment analyses of the expanded gene families of T.tibetana showed that these expanded genes are mainly enriched in GO items related to chromosome stability,DNA stability,DNA recombination repair,and enriched in KEGG items related to disease and immunity.Positive selection analysis showed that there are 46 and 105 genes in T.tibetana which are subject to positive selection compared to D.rerio and M.anguillicaudatus,respectively.Among them,1 and 4 genes were identified to participate in DNA repair,DNA recombination repair,and UV radiation resistance,respectively.Convergent evolution showed that there were 6 groups of orthologous genes related to DNA recombination repair induced by intense UV radiation.The results of comparative genomics analyses clarified that different fish has different daptation mechanism to the strong UVR environment,and there are three adaptive evolution strategies for T.tibetana to UV:(1)Reducing the inhibition of melanin synthesis to produce more melanin and protect the body.(2)Enhancing the stability of chromosomes and DNA to reduce the damage induced by UV;(3)Enhancing the ability of DNA repair.3.Molecular adaptation mechanisms of the alien M.anguillicaudatus to elevated UV radiation on the Tibetan PlateauThe transcriptomes of the dorsal skin and visceral peritoneum of wild M.anguillicaudatus between Chabalang Wetland in Tibet(3592.3 m a.s.l)and Beisha Lake in Hubei(18.3 m a.s.l)were sequenced by RNA-seq,and then the comparative transcriptomics analyses were performed.In the alien M.anguillicaudatus,1,710 differentially expressed genes(DEGs)in dorsal skin and 9,598 DEGs in visceral peritoneum were found,which suggested that the visceral peritoneum was more sensitive to highland environment.KEGG enrichment analyses of DEGs between alien M.anguillicaudatus(HMS)and lowland native M.anguillicaudatus(LMS)were performed.Among the DEGs in dorsal skin,several genes associated with pigment cells were enriched,and the expression of genes related to melanin synthesis were up-regulated in HMS.In addition,the DEGs in visceral peritoneum were significantly enriched into pathways associated with DNA repair system and pigment cell.Our results indicated that M.anguillicaudatus could activate genes related to different DNA repair pathways to respond and repair DNA damage induced by ultraviolet radiation.Moreover,the main defense mechanism for the dorsal skin and visceral peritoneum of alien HMS in response to elevated UV radiation is to carry out photoprotection through the regulation of pigment cell-related genes to reduce the damage caused by UV radiation,which is consistent with the adaptation mechanism of T.tibetana by increasing the synthesis of melanin to defense strong ultraviolet radiation.4.The effects and responses of UVB radiation on T.tibetanaAn artificial environmental dose of UVB radiation on T.tibetana was conducted in the present study.The histopathological changes in the dorsal skin structure were examined,and the time-course expression of DNA repair system-related and pigmentation-related genes were analyzed during UVB-exposed and after UVB exposure,whereas specimens before UVB exposure served as a control group.Histological observations of dorsal skin at 6 h of UVB exposure induced necrosis,degeneration,and reduced number of mucous cells in the epidermis,and increased number of melanophore in the dermis.After 24 h of recovery kept in darkness,the histological structure and the distribution of melanophore in dorsal skin got back to same levels as control group.The gene expression results demonstrated an activation of the p53-dependent cell cycle arrest genes in dorsal skin of T.tibetana exposed to UVB radiation.The expression of genes related to cell cycle arrest increased during the UVB exposure and significantly decreased at early recovery period,and then increased at late recovery period.The expression of DNA repair genes increased to the highest level at late recovery period.The expression levels of anti-apoptotic gene BCL-2 was down-regulated at all the time.Interestingly,the expression levels of genes related to melanin synthesis and transport were up-regulated during UVB exposure and the late recovery period,which was consistent with that in the histological results.Our results indicated that UVB radiation could induce the cellular damage and activated the expression of genes related to synthesis and transport of melanin,as well as DNA repair system,increasing the number of melanocytes in the dermis and ienhancing the repair of damaged DNA,so as to realize the adaptation to the strong UVB radiation environment.This result verifies the molecular mechanism that the T.tibetana adapts to the strong ultraviolet radiation environment of the Qinghai-Tibet Plateau by increasing the synthesis of melanin and enhancing the ability to repair DNA damage.