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Knockout B lymphoma cell lines as biochemical tools to explore multiple signalling pathways
Author(s) -
Veale Margaret F,
Dietrich Wendy M,
Corcoran Lynn M
Publication year - 2003
Publication title -
immunology and cell biology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.999
H-Index - 104
eISSN - 1440-1711
pISSN - 0818-9641
DOI - 10.1046/j.0818-9641.2003.01172.x
Subject(s) - bruton's tyrosine kinase , signal transduction , microbiology and biotechnology , biology , cell culture , b cell receptor , b cell , intracellular , receptor , tyrosine kinase , immunology , biochemistry , antibody , genetics
Studies on B lymphocyte signalling pathways using B lymphocytes from genetically modified mice have the disadvantages of primary cell polyclonality and finite life span. B lymphoma cell lines have been generated from mice with targeted mutations in the oct‐2 , OBF‐1 , vav‐1 and btk genes, as a model system that lacks these limitations and possesses additional potential for experimental manipulation. To assess their utility, activation of the B cell receptor using anti‐µ, the Toll‐like receptor‐4 using lipopolysaccharide and the interleukin‐4 receptor were assessed in these cell lines. Differential tyrosine phosphorylation of intracellular proteins was measured in the wild‐type controls compared to the corresponding mutant cell lines after B cell receptor stimulation. Intracellular calcium (Ca 2+ i ) was mobilized in the control cell lines but not in the OBF‐1 and Vav1‐deficient cells, while Xid B cell lines ( btk mutant) showed a reduced Ca 2+ mobilization. Extracellular signal‐regulated kinase 1/2 phosphorylation in response to anti‐µ or lipopolysaccharide stimulation was significantly reduced in Vav1‐deficient cells. Interleukin‐4 stimulation of wild‐type cells resulted in a 2−3‐fold increase in Stat‐6 phosphorylation. These results indicate that the cell lines mimic the biochemical responses of the corresponding primary B cells. They therefore represent a useful model system to investigate the regulation and roles of these and other gene products in B cell signal transduction and activation.