Studies On Phosphorylated Protein In Skin Tissue Of Early Stage Of Er-mao And Er-mao Stage Of Tan Sheep

Tan sheep is an excellent local sheep breed in China.It's er-mao fur is beautiful,light and warm.It is the top product of fur.But after the period of er-mao,with the increase of the age of Tan sheep,the wool bunch gradually becames thicker and longer,the special characteristics of er-mao fur weakens or disappears.Besides,it will become loose and the flower spike is not beautiful.To understand the cause of formation of fur,TMT label and IMAC phosphorylated peptide enrichment techniques and quantitative proteomics research strategy by liquid chromatography-mass spectrometry(LC-MS/MS)were used to study the phosphorylated proteomics of Tan sheep skin at early stage of er-mao and er-mao stage.The protein phosphorylation modification of skin tissue of Tan sheep at 120 days gestational age and newborn was studied.A total of 10167 phosphorylation modification sites on 3298 proteins were identified.The fetal 120day gestational age group and newborn group were screened with FC>1.3 or<1/1.3,P<0.05 for differentially phosphorylated proteins.Quantitative analysis showed that 900 sites on 570 proteins were up-regulated and 680 sites on 486 proteins were down-regulated.Bioinformatics analysis of differentially modified proteins revealed that acidophilic kinases may play an important role in fetal skin development,and KEGG pathway enrichment analysis showed that changes in BMPR2 protein phosphorylation at Ser683 sites and Smad5 at Ser133 sites could affect hair follicle development cycle by affecting BMP signal transduction.Meanwhile,the high level of phosphorylation modification of vimentin at Ser432 and Ser341 may affect hair follicles in the early stages of fetal development;changes in phosphorylation modification at KAP2.3 and Ser13 sites may affect the early keratinization of hair fibers by affecting the chemical binding of KIFs to the matrix.Phosphorylated proteomics was used to study the skin samples of Tan sheep at newborn and er-mao stages.A total of 2,806 phosphorylated proteins were identified,including 8,184 phosphorylation sites.The newborn group and er-mao group were screened with FC>1.3 or<1/1.3,P<0.05 for differentially phosphorylated proteins.The results of quantitative analysis showed that the phosphorylation levels of 171 sites had been up-regulated in the skin samples at newborn stage when compared with the skin samples at the er-mao stage.Meanwhile,125 sites had been down-regulated at the same stage.As shown by the results of the functional enrichment analysis of the differentially phosphorylated proteins,they had been mainly enriched in the cysteine and methionine metabolism.In addition,the phosphorylation levels of KAP4.7 and KAP13.1 had also varied during the different skin stages.These results indicated that the cysteine metabolism pathways,and the phosphorylation modifications of the keratin associated proteins in the skin,played important roles in the formation of the er-mao stage fur of the Tan sheep.The study of phosphorylated proteome in skin tissue of Tan sheep at different developmental stages provides a theoretical basis for the formation of the unique fur of Tan sheep.A new understanding of the formation mechanism of fur of er-mao of Tan sheep is presented from the perspective of protein phosphorylation modification.