Analysis Of Skin Transcriptome Profiling And Effect Of KRTAP Genes On Cashmere Quality In Goats

Cashmere is a high-grade textile material.Ziwuling black goats(low cashmere yield)and Liaoning cashmere goats(high cashmere yield)in this study were selected to discover genes that affect cashmere yield and cashmere quality by using the method of transcriptome sequencing technology(RNA-Seq),real-time fluorescent quantitative PCR(RT-qPCR)and polymerase chain reaction single-strand conformation polymorphism(PCR-SSCP)and so on.In this study,we explored the characteristics of the expression profile of cashmere goat skin tissues and analyzed the effects of three keratin association protein genes(KRTAPs)on cashmere fiber traits.There were four findings in this study.(1)A total of 668 differentially expressed genes(DEGs)were obtained between Ziwuling black goats and Liaoning cashmere goats through analyzing the results of skin transcriptome sequencing,among which 340 were up-regulated and 328 were down-regulated in the skin samples of Liaoning cashmere goats.Among the differentially expressed genes,the most surprising aspect of the expression levels of KRTAP genes and KRTAP-like genes,which were related to the cashmere fiber traits,were all down-regulated.The results of GO and KEGG indicated that the up-regulated differentially expressed genes mainly were enriched in the extracellular matrix and were related to immunity,cell migration,cytokine-cytokine receptor interaction and hematopoiesis.The down-regulated differentially expressed genes mainly were enriched in pigment granules and intermediate filament cytoskeleton and were related to enzyme activity and melanin metabolism.(2)The sequence variations of caprine KRTAP15-1 were detected by the method of PCR-SSCP in 375 Ziwuling black goats,and six variants(named CAPHI-KRTAP15-1*A to CAPHI-KRTAP15-1*F)containing eight single nucleotide polymorphisms(SNPs)were identified.Five of the SNPs were non-synonymous and would lead to putative amino acid changes.Association analyses revealed that variant A of caprine KRTAP15-1 was associated with decreasing mean fiber diameter,and this effect appeared to be dominant;while variant C was found to be associated with increasing mean fiber diameter,the effect being recessive.The findings suggest that caprine KRTAP15-1 is highly polymorphic and that variation in this gene affects cashmere mean fiber diameter.These results suggest that KRTAP15-1 may provide new insight into improving cashmere fiber diameter.(3)In this study,we confirmed the identification of a new KRTAP27-1 in goat genome firstly,and then described its effect on cashmere traits in 375 Ziwuling black goats.The PCR-SSCP analysis was used to screen for sequence variation in this gene,and three sequence variants(named CAPHI-KRTAP27-1*A to CAPHI-KRTAP27-1*C)were found.These sequences had the highest similarity to a human KRTAP27-1 sequence.There were two SNPs detected in the coding sequence,including one nonsynonymous SNP(c.413C/T;p.Ala138Val)and one synonymous SNP(c.495C/T).The analysis results showed that cashmere fiber diameter was higher when variant B was presented.Goats with the genotypes AB and BB produced cashmere fibers of higher mean fiber diameter than goats with genotype AA,but no difference in mean fiber diameter was detected between the AB and BB goats.These results suggested that B was associated with increasing mean fiber diameter.KRTAP27-1 can be used as a candidate marker gene to improve cashmere fiber diameter.(4)In this study,we identified the keratin-associated protein 1-2 gene(KRTAP1-2)in the goat genome and described its effect on cashmere fiber traits in 375 Ziwuling black goats.Six sequence variants(named CAPHI-KRTAP1-2*A to CAPHI-KRTAP1-2*F)were revealed using PCR-SSCP analysis.These sequences had the highest homology with ovine KRTAP1-2 sequences.There were a 60-bp deletion,a 15-bp insertion and five SNPs including two non-synonymous SNPs in the coding sequence.And the expression level of caprine KRTAP1-2 gene was higher in Ziwuling black goats than Liaoning cashmere goats,and the expression level was significantly different(P < 0.05).Variation in caprine KRTAP1-2 was found to be associated with raw cashmere fiber weight,but not with fiber diameter and length.The present of variant B can significantly reduce the weight of cashmere fiber.Therefore,the weight of cashmere fiber can be increased by reducing the content of variant B in goat population.In summary,the gene expression profile of skin tissue from Ziwuling black goat was obtained in this study.The differentially expressed gene caprine KRTAP15-1 were closely related to cashmere fiber quality,and the sequence variations of caprine KRTAP15-1affected the mean cashmere fiber diameter.The sequence variations of newly identified caprine KRTAP27-1 were also associated with cashmere mean fiber diameter,too.The caprine KRTAP1-2,a newly identified KRATP gene,was involved in raw cashmere fiber weight.The caprine KRTAP15-1and newly identified caprine KRTAP27-1 and KRTAP1-2may be used as marker genes for improving mean fiber diameter and cashmere fiber weight.