Identification And Function Validation Of Bovine Mastitis-related LncRNAs

Mastitis is one of the most popular diseases in dairy industry,causing great economic loss worldwide.The disease is due to invasion of the mammary glands by pathogenic bacteria and the outcome of the multiplication followed in the udder.And E.coli and S.aureus in the environment are mainly ascribed to the inflammation.Lipopolysaccharides(LPS),the major structural elements of Gram-negative bacteria as E.coli,trigger strong immune responses via TLR4,while Lipoteichoic acid(LTA),an important cell wall component of from Gram-positive bacteria as S.aureus,introduce inflammation by TLR2.Increasing evidences indicate that lncRNA also exerts great influence in inflammation cascade.However,the noncoding parts of gene network in immunity of bovine mammary epithelial cell remain largely unknown.To establish the cell model of mastitis appropriately in this study,Heat-killed bacterial was applied to treat MAC-T cells,in which tumor necrosis factor alpha(TNF-?),a well-known master cytokine governing the early stage of inflammation,was considered as the parameter of immune reaction.Then lncRNA expression profile of healthy MAC-T cells and mastitis model MAC-T cells of were obtained by high-throughput sequencing,then from which we analysed the differentially expressed lncRNA by bioinformatics method coupled with further study on the function of lncRNA by using CRISPR-Cas9 System.A total of 1323 novel lncRNA were identified,of which 53 were differentially expressed.And the functional analysis of lncRNA indicated that differentially expressed lncRNA were closely related to cell immunity.Then a differently expressed lncRNA(named lncRNA-TUB)was selected and knockout by crisper cas9 system,which triggered sever EMT phenomenon and other physiological change of MAC-T cell including proliferation,migration,?-casein secretion and immune response.In this study,we identified a class of previously undetected inflammation-related lncRNAs,in which lncRNA-TUB was found played a vital role in many biological processes including immune reaction,aimed to elucidate the pathogenesis of mastitis in epigenetic perspective and provide new targets to further research on the link between these lncRNAs and mastitis.