IBR   13079
INSTITUTO DE BIOLOGIA MOLECULAR Y CELULAR DE ROSARIO
Unidad Ejecutora - UE
congresos y reuniones científicas
Título:
Molecular characterization of adaptor proteins ClpT1/2 and ClpS from Arabidopsis thaliana
Autor/es:
COLOMBO, CLARA V.; ROSANO, GERMÁN L.; CECCARELLI, EDUARDO A.
Lugar:
Heidelberg
Reunión:
Simposio; EMBO symposium: Quality Control, from molecules to organelles; 2012
Institución organizadora:
EMBO/EMBL
Resumen:
Proteolysis in plant chloroplast is highly regulated and is an essential tool for organelle protein quality control. The chloroplastic ClpPR system of Arabidopsis thaliana is composed by proteins ClpP1/3-6 and ClpR1-4, which self assemble in a proteolytic chamber. Substrate selection is carried out by the Hsp100 chaperones ClpC1/2 and ClpD. They specifically bind proteins destined to degradation, unfold and translocate them into the chamber. Three recently discovered proteins, ClpT1/2 and ClpS are thought to be adaptors and/or regulators of the Clp system. ClpT1/2 may assist the assembly of the ClpPR complex and may regulate the binding of the Hsp100 chaperones to the core. ClpS is proposed to modulate ClpC1/2 substrate selection and affinity, providing new targets to this system. Its homologous protein in Escherichia coli, ClpS, is a recognin of the N-end rule degradation pathway. This protein recognizes N-end rule substrates and presents them to ClpAP for degradation. Also, ClpS of A. thaliana could have a similar function in chloroplast. Nevertheless, there is little in vitro evidence to support these functions. We determined the oligomerization status of recombinant ClpT1 and ClpS by molecular exclusion chromatography, where ClpT1 is monomeric and ClpS can form homo-oligomers of 2, 3, 4 and 6 subunits. We studied the interaction of these proteins with ClpC2 and ClpD by fast ultrafiltration analysis. The adaptor proteins were found to associate with the chaperones in the presence of ATP. Furthermore, we have investigated the influence of ClpT1 in the ATPase activity of ClpC2 and ClpD. ClpT1 has no influence in the activity of ClpC2, however it increments ClpD ATPase activity in 60 %, demonstrating that ClpT1 is a specific activator of ClpD. Our results represent the first evidence of regulatory function of ClpT1 on Hsp100 chaperones and provide new insights for understanding regulation of proteostasis in plant chloroplast.