Chloroplast DnaJ-like proteins 3 and 4 (CDJ3/4) from Chlamydomonas reinhardtii contain redox-active Fe-S clusters and interact with stromal HSP70B
Résumé
In this study we report on the identification and characterization of three novel chloroplast-targeted J-domain proteins, CDJ3-5, which in addition to their J-domains contain bacterial-type ferredoxin domains. In the databases we could identify homologs of CDJ3-5 in green algae, moss and higher plants, but not in cyanobacteria. Phylogenetic analyses allowed distinguishing two clades containing CDJ3/4 and CDJ5 that must have diverged early in an ancestor of the green lineage and have further diversified later on. Molecular and biochemical analysis of CDJ3 and CDJ4 from Chlamydomonas reinhardtii revealed that both are weakly expressed proteins that appear to be localized to the stroma and to thylakoid membranes, respectively. The low transcript levels of the CDJ3 and CDJ4 genes declined even further in the initial phase of heat shock, but CDJ3 transcript levels strongly increased after dark-to-light shift. Accordingly, the Arabidopsis ortholog of CDJ5 was also found to be light inducible and to be under strong circadian control. CDJ3 and CDJ4 proteins could both be expressed in Escherichia coli with redox-active Fe-S clusters. In vitro crosslinking studies demonstrated that CDJ3 and CDJ4 interact with chloroplast HSP70B in the ATP state, presumably as dimers, and immunoprecipitation studies showed that CDJ3/4 were in common complexes with HSP70B also in Chlamydomonas cell extracts. Finally, CDJ3 was found in complexes with apparent molecular masses of ~550 to 2800 kDa that appeared to contain RNA. We speculate that CDJ3-5 might represent redox switches that act by recruiting HSP70B for the reorganization of regulatory protein complexes.
Origine : Fichiers produits par l'(les) auteur(s)
Loading...