| Body color is one of the basic characteristics of animals,which is mainly determined by the type,quantity and distribution of pigment cells deposited in the dermis of animals.It is of great significance in the survival and reproduction of animals.Compared with mammals,fish have more abundant body color patterns and pigment cell types.At present,there are many studies on the formation and regulation mechanism of fish body color,but mainly focus on model organisms such as zebrafish(Dannio rerio),medaka(Oryzias latipes)and some ornamental fish.Sinibrama taeniatus is an endemic fish in the upper reaches of the Yangtze River with both ornamental and edible value.At present,the reports on S.taeniatus mainly focus on the basic research of its resource status,phylogeny and reproductive biology,while the related research on the body color of S.taeniatus has not yet been carried out.Therefore,the preliminary study on the development and regulation of body color in S.taeniatus is helpful to analyze the molecular regulation mechanism of body color occurrence and breeding of excellent body color phenotypic strains.In this study,microscopic observation,high performance liquid chromatography and transcriptome sequencing analysis were used to study the distribution of pigment cells,pigment content changes and differentially expressed genes in the skin of S.taeniatus at different developmental stages.The main results and conclusions are as follows:1.Changes of pigment cells and pigment content in different developmental stages of S.taeniatus.In this study,the types of pigment cells and pigment content in the whole body of S.taeniatus at different developmental stages were compared.Melanocytes,xanthophore and iridophores were identified in S.taeniatus.Melanocytes first appeared in the eye socket on3 day post fertilization(dpf).The iridophores first appeared on the eyes at 4dpf.The xanthophore first appeared in the trunk and heart at 5dpf.At 1 month post fertilization(mpf),the abdomen is transparent and the viscera can be clearly observed.At 3mpf,the abdomen was covered with a large number of melanocytes and iridescent cells,showing translucent;at9mpf,the ink line on both sides of S.taeniatus was formed;at 12 mpf,the muscles between the anal fin and the lateral line are full of pigment cells,and the overall body color pattern is completely formed.Melanin content continued to increase during development,guanine content increased most at 3mpf,and pteridine and β-carotene increased most at 12 mpf.2.Transcriptome sequencing and analysis of S.taeniatus at different developmental stages.The skin tissues of 3dpf,15 dpf,1mpf,3mpf,6mpf,9mpf and 12 mpf were subjected to transcriptome comparative analysis,and 14,332 differentially expressed genes were identified.Functional enrichment analysis showed that differentially expressed genes were significantly enriched in retinol metabolism,thyroid hormone synthesis,purine metabolism,pyrimidine metabolism and other pathways,suggesting that these pathways were closely related to body color development.Weighted gene co-expression network analysis found four modules related to pigment synthesis,namely darked,thistle,darkorange2 and darkmagenta.Combined with KEGG enrichment analysis and co-expression network analysis,eight hub genes(mitfa,wnt9 a,wnt10b,wnt11,adcy5,edn1,adcy8 and rdh8)were obtained,suggesting that these genes may play an important role in pigmentation.3.Effects of thyroxine on body color formation of juvenile S.taeniatus.In this study,different concentrations(0,1,10,20,40 and 80 n M)of thyroxine were used to soak S.taeniatus for 30 days.The results showed that the body color of S.taeniatus was lighter and the melanin cells were less after thyroxine treatment.With the increase of thyroxine concentration,the number of melanocytes decreased significantly(p < 0.01).The content of guanine increased significantly(p < 0.01),the content of β-carotene decreased significantly(p < 0.01),the content of pteridine decreased significantly only in the silent concentration group(p < 0.05),and the content of melanin increased significantly(p < 0.01).At the same time,with the increase of thyroxine concentration,the expression of genes related to iridescent cells and yellow pigment cells was up-regulated,such as pnp4 a,mpv17,xdh,bco1,urah and so on.Among the genes related to melanin,mitf and plcb4 were significantly down-regulated;gsk3b and wnt10 b were significantly up-regulated;and many genes showed dynamic changes in different concentration groups.In summary,this study identified melanocytes,xanthophore and iridophores in S.taeniatus,and morphologically observed their body color formation.At the same time,the differentially expressed genes of mitfa,wnt9 a,wnt10b,wnt11,adcy5,edn1,adcy8 and rdh8 were obtained by transcriptome sequencing technology,and the key regulatory pathways such as retinol metabolism,thyroid hormone synthesis,purine metabolism and pyrimidine metabolism pathway were screened out.Finally,through thyroxine treatment,it was shown that thyroxine promoted the formation of yellow pigment cells and iridophores,and the regulation of melanocyte differentiation and sedimentation was inconsistent.This study explored the changes of pigment cells and pigment content in the process of fish body color formation,and preliminarily explored the molecular mechanism of its changes.Our results lay a foundation for studying the molecular mechanism of fish pigmentation and provide a theoretical basis for color-based molecular breeding programs in aquaculture. |
