title: Knockout B lymphoma cell lines as biochemical tools to explore multiple signalling pathways.
abstract: 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- micro, 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 (Ca2+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 Ca2+ mobilization. Extracellular signal-regulated kinase 1/2 phosphorylation in response to anti- micro 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.
authors.affiliation: The Walter and Eliza Hall Institute of Medical Research, 1G Royal Parade, Victoria 3050, Australia.
authors.forename: Margaret F
authors.initials: MF
authors.lastname: Veale
authors.forename: Wendy M
authors.initials: WM
authors.lastname: Dietrich
authors.forename: Lynn M
authors.initials: LM
authors.lastname: Corcoran
mesh_terms: D000077329:Agammaglobulinaemia Tyrosine Kinase; D000818:Animals; D001402:B-Lymphocytes / Q000276:immunology / Q000502:physiology*; D020013:Calcium Signaling; D018797:Cell Cycle Proteins; D045744:Cell Line, Tumor; D004268:DNA-Binding Proteins / Q000235:genetics; D005816:Genetic Complementation Test; D016130:Immunophenotyping; D015847:Interleukin-4 / Q000276:immunology; D008213:Lymphocyte Activation; D016393:Lymphoma, B-Cell / Q000235:genetics / Q000276:immunology; D051379:Mice; D018345:Mice, Knockout; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism; D050813:Octamer Transcription Factor-2; D010766:Phosphorylation; D011505:Protein-Tyrosine Kinases / Q000235:genetics / Q000378:metabolism; D011518:Proto-Oncogene Proteins / Q000235:genetics; D051567:Proto-Oncogene Proteins c-vav; D050800:STAT6 Transcription Factor; D015398:Signal Transduction; D015534:Trans-Activators / Q000235:genetics / Q000378:metabolism; D014157:Transcription Factors / Q000235:genetics
chemical_list: D018797:Cell Cycle Proteins; D004268:DNA-Binding Proteins; D050813:Octamer Transcription Factor-2; C490424:Pou2af1 protein, mouse; C489206:Pou2f2 protein, mouse; D011518:Proto-Oncogene Proteins; D051567:Proto-Oncogene Proteins c-vav; D050800:STAT6 Transcription Factor; C494100:Stat6 protein, mouse; D015534:Trans-Activators; D014157:Transcription Factors; C496619:Vav1 protein, mouse; D015847:Interleukin-4; D011505:Protein-Tyrosine Kinases; D000077329:Agammaglobulinaemia Tyrosine Kinase; C000625951:Btk protein, mouse; D020928:Mitogen-Activated Protein Kinases