Related Experiment Videos
The Ku70 autoantigen interacts with p40phox in B lymphocytes
N Grandvaux1, S Grizot, P V Vignais
1Laboratoire de Biochimie et Biophysique des Systèmes Intégrés (UMR 314/CNRS), Département de Biologie Moléculaire et Structurale, Commissariat à l'Energie Atomique-Grenoble, 38054 Grenoble Cedex 9, France. Nathalie@tuckett.ceng.cea.fr
Journal of Cell Science
|January 23, 1999
Summary
Ku70 protein interacts with p40phox, a component of NADPH oxidase, in B lymphocytes. This interaction and the role of DNA-PK in activating other phox proteins may explain the low levels of NADPH oxidase in B cells.
Area of Science:
- Molecular Biology
- Immunology
- Cell Biology
Background:
- Ku70 is a regulatory component of DNA-dependent protein kinase (DNA-PK).
- p40phox is a regulatory component of the O2--producing NADPH oxidase.
- The phox protein triad (p40phox, p47phox, p67phox) is expressed in B lymphocytes and neutrophils, but NADPH oxidase activity is low in B cells.
Purpose of the Study:
- To investigate the interaction between Ku70 and p40phox.
- To elucidate the role of Ku70 and DNA-PK in the function of NADPH oxidase components in B lymphocytes.
Main Methods:
- Yeast two-hybrid screen to identify interacting proteins.
- Truncated protein constructs to map interaction sites.
- Co-immunoprecipitation and chromatography to confirm protein binding.
- Cellular localization studies (colocalization) in B lymphocytes and Cos-7 cells.
- In vitro kinase assays to assess substrate phosphorylation.
Main Results:
- Ku70 directly interacts with p40phox through its C-terminal region.
- Ku70 and p40phox colocalize in the nucleus of B lymphocytes.
- p47phox and p67phox are phosphorylated by DNA-PK in vitro.
- The phox proteins and Ku70 form complexes within B cells.
Conclusions:
- Ku70 is a binding partner of p40phox, suggesting a role in regulating NADPH oxidase assembly or activity.
- DNA-PK's interaction with and phosphorylation of phox proteins may contribute to the regulation of NADPH oxidase in B lymphocytes.
- These findings offer a potential explanation for the low levels of functional NADPH oxidase in B lymphocytes compared to neutrophils.