Cystolic chaperones influence the fate of a toxin dislocated from the endoplasmic reticulum.
Spooner, R.A., Hart, P.J., Cook, J.P., et al. Proc. Nat'l Acad. Sci., 105(45): 17408 (November, 2008).
A study of the mechanism by which ricin regains activity after passage through the endoplasmic reticulum has identified the involvement of several cochaperones in determining a cell's sensitivity to the cytotoxic agent. This poison enters the cell by binding with the exterior membrane, triggering endocytosis and its translocation to the endoplasmic reticulum. There, ricin is split into its two components, a membrane binding region and the catalytic subunit (RTA) that causes ribosome depurination and ultimately, cell death. RTA passes into the cytoplasm via channels that are normally used to direct proteins for degradation. This involves a loss of tertiary structural integrity that causes other proteins to aggregate or renders them susceptible to polyubiquitinylation.
Using several inhibitors of cochaperone binding, the authors of the present report demonstrated that chaperone-mediated pathways are involved in restoring ricin to its active, cytotoxic state or preventing it from refolding and thereby protecting the cell. Chaperone involvement in determining ricin's lethality was first established with an inhibitor to Hsc70 ATPase and an antagonist to the Hsp90 ATP-binding site. The results show that inhibition of Hsc70 protects cells from ricin, while blocking Hsp90 render them more susceptible to it. A simple in vitro experiment further demonstrated that Hsc70 interacts directly with RTA and is capable of restoring the catalytic activity of heat-denatured toxin. Genetic manipulation of Hsc70 cochaperones - Hop (Hsc70-Hsp90 organizing protein), CHIP (C terminus of Hsp70-interacting protein), and Hip (Hsp70-interacting protein) - added detail to the participation of this chaperone in vivo. Specifically, the research determined that increasing the interaction of Hsc70 with RTA favors productive folding of the cytotoxin , resulting in greater toxicity, while increasing the interaction with Hsc70 and Hsp90 favors inactivation of ricin.
This is the first report to demonstrate the molecular pathway involved in the transport and reconstitution of ricin within mammalian cells. It is noteworthy that Hsc70 is directly implicated in the refolding of the cytotoxic moleculre and that cochaperones play a key role in determining its ultimate fate in the cytoplasm.
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