G. Tanguy, L. Drevillon, N. Arous, M. Goossens, P. Fanen
INSERM, U654; Université Paris 12; HôpitaI Henri Mondor; Créteil, France
In order to identify news proteins that might interact with CFTR and modify the processing of the protein, we performed a yeast two hybrid screening with the third cytoplasmic loop (CL3) of CFFR as a bait. CSN5/Jabl, the fifth component of the COP9 signalosome, was found to interact with CL3. This interaction was later tested in yeast with class II mutants of CL3 ($945L, H949Y and D979A) and a loss of interaction was observed with one of them.
In vivo experiments showed that CSN5/Jabl and CFIR could be coimmumo-precipitated from HeLa cells stably expressing wt or delF508 CFTR. Our results show that the direct interaction between CFTR and CSN5/Jabl is strongest with delF508 than with wt CFIR.
Cellular localization of CSN5/Jabl is wide: i) as a component of the signalosome, it is predominantly in the nucleus, ii) as a monomer it is both ha the nucleus and the cytoplasm and iii) as a small loose complex ha the cytoplasm. Thus, we have looked at the localization of CSN5/Jabl in the presence of wt or delF508 CFFR in HeLa cells and observed different patterns.
CSN5/Jabl is able to interact with a variety of signaling molecules and is a regulator of their stability in mammalian cells. We are studying the over expression (tr ansfection) and the down-expression (siRNA) of CSN5/Jabl on the processing of wt and delF508 CFIR by pulse chase and in vitro/phosphorylation experiments. In addition, it is known that CSN5/Jabl activity is inhibited by curcumin and that this drug might improve CFTR delF508 prccessing toward the plasma membrane. We are therefore testing the effect of curcumin combined with CSN5/Jabl on CFTR processing.
We have identified CSN5/Jabl as a partner of CFFR but further studies are necessary to investigate the functional role of CSN5/Jabl on CFFR processing and how it could be targeted for a therapeutic benefit of cystic fibrosis.
This work was supported by grants from INSERM, Vaincre la Mucoviscidose and Chancellerie des Universités de paris.