The Ku70 autoantigen interacts with p40phox in B lymphocytes.

Ku70, a regulatory component of the DNA-dependent protein kinase, was identified by a yeast two-hybrid screen of a B lymphocyte cDNA library as a partner of p40phox, a regulatory component of the O2--producing NADPH oxidase. Truncated constructs of p40phox and Ku70 were used to map the interacting sites. The 186 ...
C-terminal amino acids (aa) of Ku70 were found to interact with two distinct regions of p40phox, the central core region (aa 50-260) and the C-terminal extremity (aa 260-339). In complementary experiments, it was observed that Ku70 binds to immobilized recombinant p40phox fusion protein and that p40phox and Ku70 from a B lymphocyte cell extract comigrate in successive chromatographies on Q Separose, Superose 12 and hydroxylapatite columns. Moreover, we report that Ku70 and p40phox colocalize in B lymphocytes and in transfected Cos-7 cells. We also show that the two NADPH oxidase activating factors, p47phox and p67phox are substrates for DNA-PK in vitro and that they are present together with p40phox in the nucleus of B cells. These results may help solve the paradox that the phox protein triad, p40phox, p47phox and p67phox, is expressed equally in B lymphocytes and neutrophils, whereas the redox component of the NADPH oxidase, a flavocytochrome b, which is well expressed in neutrophils, is barely detectable in B lymphocytes.
Mesh Terms:
Animals, Antigens, Nuclear, Autoantigens, B-Lymphocytes, Binding Sites, COS Cells, Cell Nucleus, Cytoplasm, DNA Helicases, DNA, Complementary, DNA-Activated Protein Kinase, DNA-Binding Proteins, Fluorescent Antibody Technique, Humans, NADPH Oxidase, Nuclear Proteins, Phosphoproteins, Phosphorylation, Protein-Serine-Threonine Kinases, Recombinant Fusion Proteins, Saccharomyces cerevisiae, Saccharomyces cerevisiae Proteins, Subcellular Fractions, Transfection
J. Cell. Sci.
Date: Feb. 01, 1999
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