Stable transfection of the bovine NRAMP1 gene into murine RAW264.7 cells:: Effect on Brucella abortus survival

被引:76
作者
Barthel, R
Feng, JW
Piedrahita, JA
McMurray, DN
Templeton, JW
Adams, LG [1 ]
机构
[1] Texas A&M Univ, Coll Vet Med, Dept Vet Pathobiol, College Stn, TX 77843 USA
[2] Texas A&M Univ, Coll Vet Med, Dept Vet Anat & Publ Hlth, College Stn, TX 77843 USA
[3] Texas A&M Univ, Coll Med, Dept Med Microbiol & Immunol, College Stn, TX 77843 USA
关键词
D O I
10.1128/IAI.69.5.3110-3119.2001
中图分类号
R392 [医学免疫学]; Q939.91 [免疫学];
学科分类号
100102 ;
摘要
Genetically based natural resistance to brucellosis in cattle provides for novel strategies to control zoonotic diseases. Bovine NRAMP1, the homologue of a murine gene (Bcg), has been identified as a major candidate for controlling the in vivo resistant phenotype. We developed an in vitro model for expression of resistance- and susceptibility-associated alleles of bovine NRAMP1 as stable transgenes under the regulatory control of the bovine NRAMP1 promoter in the murine RAW264.7 macrophage cell line (Bcg;) to analyze the regulation of the NRAMP1 gene and its role in macrophage function. We demonstrated that the 5'-flanking region of bovine NRAMP1, despite the lack of TATA and CAAT boxes, has a functional promoter capable of driving the expression of a transgene in murine macrophages. A polymorphism within a microsatellite in the 3' untranslated region critically affects the expression of bovine NRAMP1 and the control of in vitro replication of Brucella abortus but not Salmonella enterica serovar Dublin. We did not observe any differences in the production of NO by resting or gamma interferon (IFN-gamma)- and IFN-gamma -lipopolysaecharide (LPS)-treated transfected cell lines, yet the resistant transfected cell lines produced significantly less NO than other cell lines, following stimulation with LPS at 24 and 48 h.
引用
收藏
页码:3110 / 3119
页数:10
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