Role Analysis of the scarb1 Gene in the Pigmentation of Neocaridina denticulata sinensis

Abstract

Body color is a key economic trait for Neocaridina denticulata sinensis, an important ornamental shrimp. Scarb1 may be an important mediator of astaxanthin uptake, changing the shrimp's body color. To discover the relationship between scarb1 and the pigmentation of cherry shrimp, the expression profiles, RNAi, and SNP genotyping of scarb1 were studied. There were significant differences in four color populations and five development stages (p < 0.05). The highest expression level of scarb1 appeared in the red population and the pre-nauplius stage. Exposure to scarb1 dsRNA increased the number and development of chromatophores at the metanauplius stage, but almost no phenotypic changes were observed at the pre-zoea stage. There was a synonymous SNP (G1593A) with a significantly different genotype frequency between the red and yellow populations (p < 0.05). The above results suggested that scarb1 is involved in pigmentation by affecting the development of chromatophores.

Keywords: RNAi; SNP genotyping; carotenoid; scavenger receptor class B type I.

Figure 1

Relative expression of scarb1 in Neocaridina denticulata sinensis. (A) The relative expression levels of scarb1 in different populations; (B) The relative expression levels of scarb1 in different developmental stages. I: pre-nauplius stage; II: metanauplius stage; III: pre-zoea stage; IV: membrane-zoea stage; V: post-larva stage. Different letters indicate significant differences (p < 0.05). Error bars represent the mean ± standard error.

Figure 2

The effect of chromatophores after scarb1 interference in Neocaridina denticulata sinensis embryos. (A) The metanauplius stage after scarb1 interference. EGFP dsRNA was the non-specific control and scarb1 dsRNA was the scarb1 interference group; (B) The pre-zoea stage after scarb1 interference. EGFP dsRNA was the non-specific control and scarb1 dsRNA was the scarb1 interference group; (C) The relative expression level of scarb1 in metanauplius and pre-zoea stages after RNAi; (D) The red pixel brightness (RPB) ratio and the pigment distribution scale (PDS) of the metanauplius stage; (E) The red pixel brightness (RPB) ratio and the pigment distribution scale (PDS) of the pre-zoea stage; (F) The length and width of chromatophore clusters in compound eyes. The black arrow indicates a chromatophore cluster in compound eyes, and the red arrow indicates an erythrophore. Different lowercase letters indicate significant differences (p < 0.05). Error bars represent the mean ± standard error.

Figure 3

Fst of high-quality SNPs in three color populations (red, yellow, and wild). All unigenes of the transcriptome were divided into 22 equal parts based on their ID number as 22 pseudochromosomes. The blue dashed line represents the lowest Fst in the top 5% of SNPs.

Figure 4

Partial sequence of scarb1. Capital letters represent amino acids, lowercase letters represent nucleotides, and underlining represents triplet codons. The red box shows the amino acids coded by SNP 1593 G>A in cherry shrimp and other species, and the upper blue letter of the box represents the amino acid of cherry shrimp after mutation. A black background indicates that the amino acid at this location is identical in all comparison species, and a gray background indicates that the amino acid at this location is similar in all comparison species.

Figure 5

Effect of the SNP on RNA secondary structure. The black lines are RNA secondary structures, the color diagram is the enlargement of the corresponding box part, and the arrow points to the exact position of the SNP. (A) The original RNA structure prediction of scarb1; (B) The influence of the 1593G>A change on the secondary structure of RNA.

Figure 6

Codon usage of serine in Neocaridina denticulata sinensis. (A) RSCU stands for Synonymous codon usage; (B) SCF stands for synonymous codon frequency.