Functional Analysis Of ABC Gene In Pigmentation Transport Of Neocaridina Denticulata Sinensis

Neocaridina denticulata sinensis belongs to the phylum arthropods,crustaceans,decapods,dactylidae,Neocaridina,It is loved by more people because of gorgeous color.The preliminary transcriptional comparison of N.denticulata sinensis showed that four ABC genes,namely NdABCG2,NdABCG5,NdABCB6 and NdABCB8,were differentially expressed in red and yellow lineages.The development and tissue expression in different kinds of shrimp were analyzed by q-PCR in red,yellow,sunkist,chocolate and wild type of 5 strains.The development of pigmentation cells in the embryo was observed after the NdABCG2 gene was knocked down by RNAi.The results are as follows:1.These four ABC transporters are all semi-transporters,with 1 NBD and 1 TMD.NdABCG2,NdABCG5,NdABCB6 and NdABCB8 contain 670,640,539 and 717 amino acid residues respectively.The sequences of these 4 proteins are highly conservative in arthropods,and all have the conservative motif of Walker A,Walker B,Q-loop.D-loop,H-loop and ABC transporter signature motif.2.In each group of the same strain.Nd4BCG2 expression level was higher in the epidermis of red,yellow,sunkist and wild strain.which was significantly higher than that of muscle and digestive tract(P<0.05).The compound eye showed the highest expression in the chocolate,which was significantly higher than the epidermis(P<0.05).The differences of NdABCG2 between different strains were mainly reflected in epidermis and compound eye tissues.The epidermal expression levels of red and sunkist strains were significantly higher than those of the other 3 strains(P<0.05),while those of yellow and wild type compound eyes were significantly higher than those of the other 3 strains(P<0.05).The expression level of NdABCG5 in compound eyes of all strains was the lowest,and the significance was lower than that of the other 4 strains(P<0.05).However,the lowest expression level in the epidermis of the red strain was lower than that of the other 4 strains(P<0.05).NdABCB6 was mainly expressed in compound eyes and epidermis,the expression level of chocolate compound eyes was significantly higher than that of other strains(P<0.05),and the expression level of red strain epidermis was significantly lower than that of other strains except sunkist(P<0.05).NdABCB8 was mainly expressed in compound eyes,epidermis and hepatopancreas.The expression levels of wild-type tissues were significantly higher than those of other strains except compound eyes.3.Compared with normally developed embryos,the number of chromatophore in embryos was decreased,the color became lighter,and the development was significantly delayed.After RNAi,the chromatophore were not clustered,but distributed alternately.The morphology of red chromatophore becomes clearer,but less numerous and more granular.The high expression of ABC transporter gene in eye and skin was associated with the abundance of chromatophore in these two tissues.The chromatophore was also synchronized with the increase of the expression of ABC gene.This suggests that the ABC gene is involved in the formation and maintenance of normal function of chromatophore.Combined with the functional features of domains,it can be inferred that NdABCG2 and NdABCG5 are involved in the transport of pigment from the chromatophore.NdABCB6 and NdABCB8 are involved in the assembly of mitochondrial iron-sulfur(Fe/S)cluster proteins or chromatosomal iron-sulfur(Fe/S)cluster protein analogs,and may play a role in Oxidation and reduction of pigment.In a word,these four ABC genes are involved in the transport or metabolism of melanosomes,affecting the formation and maintenance.Therefore,there is a strong correlation between its expression level and chromatophore.After interfering with the NdABCG2 gene,the number of embryonic chromatophore decreased and the diameter became shorter,which furtherly indicated that this gene was involved in the formation of pigmentation and the maintenance of normal morphology.However,there was no significant effect on the color and size of the compound eye,which indicated that the gene was functionally different from the homologous gene white in Drosophila melanogaster.