title: Inducible expression of enhanced green fluorescent protein by interleukin-1α, interleukin-1β and Toll-like receptor 2 promoters in goat mammary epithelial cells in response to bacterial challenges.
abstract: The development of a bacteria-inducible expression system has several advantages compared with persistent expression of anti-bacterial proteins in milk to prevent and treat mastitis. The present study determined whether mastitis responsive promoters could regulate enhanced green fluorescent protein (EGFP) expression in goat mammary epithelial cells (GMECs) in response to challenges with Escherichia coli, Staphylococcus aureus or Streptococcus agalactiae. The level of expression of interleukin (IL)-1α was significantly increased in GMECs challenged with E. coli, S. aureus or S. agalactiae compared with untreated GMECs. IL-1β was induced by E. coli and S. aureus, while Toll-like receptor 2 (TLR2) was induced by E. coli only. GMECs were transfected with IL-1α, IL-1β and TLR2 promoter-EGFP reporter gene lentiviral expression vectors and the levels of expression of EGFP were measured by flow cytometry and Western blot analysis after bacterial challenge. EGFP expression driven by the IL-1α and IL-1β promoters was higher in GMECs challenged with E. coli, S. aureus or S. agalactiae than in untreated GMECs. There were no differences in EGFP expression driven by the TLR2 promoter between GMECs challenged with S. aureus or S. agalactiae and untreated GMECs, but EGFP expression was significantly increased in GMECs challenged with E. coli. Overall, these results indicate that the promoters of some bacteria-inducible genes can regulate EGFP expression in GMECs in response to bacterial challenges. This bacteria-inducible expression strategy could be used for production of mastitis resistant animals by regulating the expression of anti-bacterial proteins in the mammary gland.
authors.affiliation: College of Veterinary Medicine, Northwest A&F University, Key Laboratory of Animal Biotechnology of the Ministry of Agriculture, Yangling 712100, Shaanxi, China.
authors.forename: Kun
authors.initials: K
authors.lastname: Ru
authors.affiliation: College of Veterinary Medicine, Northwest A&F University, Key Laboratory of Animal Biotechnology of the Ministry of Agriculture, Yangling 712100, Shaanxi, China.
authors.forename: Feng
authors.initials: F
authors.lastname: Su
authors.affiliation: College of Veterinary Medicine, Northwest A&F University, Key Laboratory of Animal Biotechnology of the Ministry of Agriculture, Yangling 712100, Shaanxi, China.
authors.forename: Yuemao
authors.initials: Y
authors.lastname: Zheng
authors.affiliation: College of Veterinary Medicine, Northwest A&F University, Key Laboratory of Animal Biotechnology of the Ministry of Agriculture, Yangling 712100, Shaanxi, China.
authors.forename: Yijun
authors.initials: Y
authors.lastname: Zhang
authors.affiliation: College of Veterinary Medicine, Northwest A&F University, Key Laboratory of Animal Biotechnology of the Ministry of Agriculture, Yangling 712100, Shaanxi, China.
authors.forename: Yan
authors.initials: Y
authors.lastname: Luo
authors.affiliation: College of Veterinary Medicine, Northwest A&F University, Key Laboratory of Animal Biotechnology of the Ministry of Agriculture, Yangling 712100, Shaanxi, China.
authors.forename: Zekun
authors.initials: Z
authors.lastname: Guo
authors.affiliation: College of Veterinary Medicine, Northwest A&F University, Key Laboratory of Animal Biotechnology of the Ministry of Agriculture, Yangling 712100, Shaanxi, China.
authors.forename: Xiaoli
authors.initials: X
authors.lastname: He
authors.affiliation: College of Veterinary Medicine, Northwest A&F University, Key Laboratory of Animal Biotechnology of the Ministry of Agriculture, Yangling 712100, Shaanxi, China.
authors.forename: Xin
authors.initials: X
authors.lastname: Liu
authors.affiliation: College of Veterinary Medicine, Northwest A&F University, Key Laboratory of Animal Biotechnology of the Ministry of Agriculture, Yangling 712100, Shaanxi, China.
authors.forename: Jingcheng
authors.initials: J
authors.lastname: Zhang
authors.affiliation: College of Veterinary Medicine, Northwest A&F University, Key Laboratory of Animal Biotechnology of the Ministry of Agriculture, Yangling 712100, Shaanxi, China. Electronic address: liuluo1028@163.com.
authors.forename: Jun
authors.initials: J
authors.lastname: Liu
authors.affiliation: College of Veterinary Medicine, Northwest A&F University, Key Laboratory of Animal Biotechnology of the Ministry of Agriculture, Yangling 712100, Shaanxi, China.
authors.forename: Yong
authors.initials: Y
authors.lastname: Zhang
mesh_terms: D000818:Animals; D001419:Bacteria / Q000276:immunology*; D002478:Cells, Cultured; D004847:Epithelial Cells / Q000276:immunology / Q000378:metabolism* / Q000382:microbiology; D005260:Female; D005786:Gene Expression Regulation / Q000276:immunology; D006041:Goats / Q000378:metabolism*; D049452:Green Fluorescent Proteins / Q000378:metabolism*; D057809:HEK293 Cells; D006801:Humans; D053582:Interleukin-1alpha / Q000235:genetics / Q000378:metabolism*; D053583:Interleukin-1beta / Q000235:genetics / Q000378:metabolism; D008321:Mammary Glands, Animal / Q000166:cytology*; D051195:Toll-Like Receptor 2 / Q000235:genetics / Q000378:metabolism
chemical_list: D053582:Interleukin-1alpha; D053583:Interleukin-1beta; D051195:Toll-Like Receptor 2; C485184:enhanced green fluorescent protein; D049452:Green Fluorescent Proteins
keywords: Goat mammary epithelial cells; Inducible expression; Interleukin-1; Mastitis; Toll-like receptors