title: Role of vav1- and src-related tyrosine kinases in macrophage activation by CpG DNA.
abstract: Macrophage activation by CpG DNA requires toll-like receptor 9 and the adaptor protein MyD88. Gram-negative bacterial lipopolysaccharide also activates macrophages via a toll-like receptor pathway (TLR-4), but we and others have reported that lipopolysaccharide also stimulates tyrosine phosphorylation in macrophages. Herein we report that exposure of RAW 264.7 murine macrophages to CpG DNA (but not non-CpG DNA) provoked the rapid tyrosine phosphorylation of vav1. PP1, a selective inhibitor of src-related tyrosine kinases, blocked both the CpG DNA-mediated tyrosine phosphorylation of vav1 and the CpG DNA-mediated up-regulation of macrophage tumor necrosis factor secretion and inducible nitric-oxide synthase protein accumulation. Furthermore, we found that the inducible expression of any of three dominant interfering mutants of vav1 (a truncated protein, vavC; a form containing a point mutation in the regulatory tyrosine residue, vavYF174; and a form with an in-frame deletion of six amino acids required for the guanidine nucleotide exchange factor (GEF) activity of vav1 for rac family GTPases, vavGEFmt) consistently inhibited CpG DNA-mediated up-regulation of tumor necrosis factor secretion and inducible nitric-oxide synthase protein accumulation in RAW-TT10 macrophages. Finally, we determined that CpG DNA-mediated up-regulation of NF-kappaB activity (but not mitogen-activated protein kinase activation) was inhibited by preincubation with PP1 or by expression of the truncated vavC mutant. Taken together, our results indicate that the tyrosine phosphorylation of vav1 by a src-related tyrosine kinase or kinases plays an important role in the macrophage response to CpG DNA.
authors.affiliation: Children's Foundation Research Center at Le Bonheur Children's Medical Center, University of Tennessee Health Science Center, Memphis, Tennessee 38103, USA.
authors.forename: Stephanie H
authors.initials: SH
authors.lastname: Stovall
authors.forename: Ae-Kyung
authors.initials: AK
authors.lastname: Yi
authors.forename: Elizabeth A
authors.initials: EA
authors.lastname: Meals
authors.forename: Ajay J
authors.initials: AJ
authors.lastname: Talati
authors.forename: Sandip A
authors.initials: SA
authors.lastname: Godambe
authors.forename: B Keith
authors.initials: BK
authors.lastname: English
mesh_terms: D000818:Animals; D018797:Cell Cycle Proteins; D002460:Cell Line; D018899:CpG Islands / Q000502:physiology*; D008264:Macrophages / Q000166:cytology / Q000201:enzymology*; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism; D009154:Mutation; D016328:NF-kappa B / Q000378:metabolism; D019001:Nitric Oxide Synthase / Q000378:metabolism; D052247:Nitric Oxide Synthase Type II; D010766:Phosphorylation; D011518:Proto-Oncogene Proteins / Q000235:genetics / Q000378:metabolism*; D051567:Proto-Oncogene Proteins c-vav; D011720:Pyrazoles / Q000378:metabolism; D011743:Pyrimidines / Q000378:metabolism; D014409:Tumor Necrosis Factor-alpha / Q000378:metabolism; D014443:Tyrosine / Q000378:metabolism; D019061:src-Family Kinases / Q000378:metabolism*
chemical_list: C108123:4-amino-5-(4-methylphenyl)-7-(tert-butyl)pyrazolo(3,4-d)pyrimidine; D018797:Cell Cycle Proteins; D016328:NF-kappa B; D011518:Proto-Oncogene Proteins; D051567:Proto-Oncogene Proteins c-vav; D011720:Pyrazoles; D011743:Pyrimidines; D014409:Tumor Necrosis Factor-alpha; D014443:Tyrosine; D019001:Nitric Oxide Synthase; D052247:Nitric Oxide Synthase Type II; D019061:src-Family Kinases; D020928:Mitogen-Activated Protein Kinases