Many essential cellular functions require the action of molecular chaperones. Chaperones function during non-stress conditions to facilitate folding of newly synthesized proteins, to remodel protein complexes, and to target for degradation regulatory proteins and misfolded proteins. During cell stress, chaperones play an essential role in preventing the appearance of folding intermediates that lead to irreversibly damaged and aggregated proteins. They promote recovery from stress by disaggregating and reactivating proteins, a process not long ago thought to be impossible. They are also involved in delivering damaged proteins to compartmentalized proteases. Protein aggregation and misfolding are primary contributors to a large number of human diseases, including Alzheimers, Parkinsons, type II diabetes, cystic fibrosis, and prion diseases. Understanding how chaperones function and how they interact with proteases will provide the foundation for discovering cures and preventions for the devastating diseases caused by protein misfolding, premature degradation, and formation of toxic protein aggregates. Our previous work showed that Escherichia coli ClpA, an AAA+ protein and the regulatory component of ClpAP protease, has molecular chaperone activity. This work and the work of others demonstrated that Clp ATPases comprise a family of ATP-dependent chaperones. Two members of the Clp family, ClpB of prokaryotes and Hsp104 of yeast, collaborate with DnaK/Hsp70 and DnaJ/Hsp40 to rescue insoluble aggregated proteins. However, the mechanisms of protein disaggregation by ClpB and Hsp104 remain unclear. Our recent studies support a mechanism in which polypeptides are extracted from aggregates by forcible unfolding and translocation. We showed that both proteins have the intrinsic ability to disaggregate some aggregates independent of the Hsp70/DnaKsystem. The products of disaggregation are unfolded polypeptides. The innate disaggregating and unfolding activities of both ClpB and Hsp104 are elicited in vitro by using mixtures of ATP and ATPgammaS (a slowly hydrolyzed and nonphysiological ATP analog) or by the use of mutants with defective ATP hydrolytic activity. It is likely that these in vitro conditions allow the substrate to be held (a process requiring ATP binding but not hydrolysis) and, at the same time, unfolded (a process requiring ATP hydrolysis). Because ClpB/Hsp104 acts in conjunction with the DnaK/Hsp70 system in vivo, these results suggest that one role of the DnaK/Hsp70 chaperone system is to synchronize substrate holding and unfolding by ClpB/Hsp104. To gain insight into the individual roles of ClpB and the DnaK system in protein remodeling we tested whether there was a further stimulation by the DnaK chaperone system under conditions that promoted protein remodeling by ClpB alone. Our results demonstrated that ClpB and the DnaK system act synergistically to remodel proteins and dissolve aggregates, implying that ATPgammaS does not simply mimic the function of the DnaK/Hsp70 system. We have also been investigating how another Clp protein, ClpX, acts with a proteolytic component, ClpP, in regulating cell division. FtsZ is the major cytoskeletal protein in bacteria and a tubulin homolog. It polymerizes into filaments at the cell midline where constriction occurs to divide the cell into two daughter cells. We have found that both FtsZ monomers and filaments are degraded by ClpXP in vivo and in vitro. Our results suggest that ClpXP modulates FtsZ assembly and disassembly via regulation of the equilibrium between free and polymeric FtsZ through degradation of FtsZ. Ongoing studies of the E. coli DnaK chaperone and its two co-chaperones, DnaJ and GrpE have been aimed at understanding the function of CbpA, a DnaJ homolog, and its regulation by CbpM. We have shown that in vitro CbpA has cochaperone activity and forms a physical complex with CbpM. Both in vitro and in vivo the activity of CbpA is inhibited by CbpM as a consequence of the interaction. These results reveal that the activity of the DnaK system can be regulated in vivo by the expression of an inhibitor specific for one of the components.
| Camberg, Jodi L; Hoskins, Joel R; Wickner, Sue (2009) ClpXP protease degrades the cytoskeletal protein, FtsZ, and modulates FtsZ polymer dynamics. Proc Natl Acad Sci U S A 106:10614-9 |
| Doyle, Shannon M; Wickner, Sue (2009) Hsp104 and ClpB: protein disaggregating machines. Trends Biochem Sci 34:40-8 |
| Chenoweth, Matthew R; Trun, Nancy; Wickner, Sue (2007) In vivo modulation of a DnaJ homolog, CbpA, by CbpM. J Bacteriol 189:3635-8 |
| Doyle, Shannon M; Hoskins, Joel R; Wickner, Sue (2007) Collaboration between the ClpB AAA+ remodeling protein and the DnaK chaperone system. Proc Natl Acad Sci U S A 104:11138-44 |
| Doyle, Shannon M; Shorter, James; Zolkiewski, Michal et al. (2007) Asymmetric deceleration of ClpB or Hsp104 ATPase activity unleashes protein-remodeling activity. Nat Struct Mol Biol 14:114-22 |
| Bougdour, Alexandre; Wickner, Sue; Gottesman, Susan (2006) Modulating RssB activity: IraP, a novel regulator of sigma(S) stability in Escherichia coli. Genes Dev 20:884-97 |
| Hoskins, Joel R; Wickner, Sue (2006) Two peptide sequences can function cooperatively to facilitate binding and unfolding by ClpA and degradation by ClpAP. Proc Natl Acad Sci U S A 103:909-14 |
| Chae, Chi; Sharma, Suveena; Hoskins, Joel R et al. (2004) CbpA, a DnaJ homolog, is a DnaK co-chaperone, and its activity is modulated by CbpM. J Biol Chem 279:33147-53 |
| Sharma, Suveena; Sathyanarayana, Bangalore K; Bird, Jeremy G et al. (2004) Plasmid P1 RepA is homologous to the F plasmid RepE class of initiators. J Biol Chem 279:6027-34 |
