HEAT-SHOCK PROTEINS HSP104 AND HSP70 REACTIVE MESSENGER-RNA SPLICING AFTER HEAT INACTIVATION

Background: The heat-shock protein Hsp104 plays a crucial role in the survival of cells exposed to high temperatures and other severe stress es, but its specific functions and the biological pathways on which it operates have been unclear. Indeed, very little is known about the sp ecific cellular processes in which any of the heat-shock proteins acts to affect thermotolerance. One essential process that is particularly sensitive to heat in many organisms is the splicing of intervening se quences from mRNA precursors. Results: We have examined the role of Hs p104 in the repair of splicing after disruption by heat shock. When sp licing in the budding yeast Saccharomyces cerevisiae was disrupted by a brief heat shock, it recovered much more rapidly in wild-type strain s than in strains containing hsp104 mutations. Constitutive expression of Hsp104 promoted the recovery of heat-damaged splicing in the absen ce of other protein synthesis, but did not protect splicing from the i nitial disruption, suggesting that Hsp104 functions to repair splicing after heat damage rather than to prevent the initial damage. A modest reduction in the recovery of splicing after heat shock in an hsp70 mu tant suggested that Hsp70 may also function in the repair of splicing. The roles of Hsp104 and Hsp70 were confirmed by the ability of the pu rified proteins to restore splicing in extracts that had been heat-ina ctivated in vitro. Together, these two proteins were able to restore s plicing to a greater degree than could be accomplished by an optimal c oncentration of either protein alone. Conclusions: Our findings provid e the first demonstration of the roles of heat-shock proteins in a bio logical process that is known to be particularly sensitive to heat in vivo. The results support previous genetic arguments that the Hsp104 a nd Hsp70 proteins have different, but related, functions in protecting cells from the toxic effects of high temperatures. Because Hsp104 and Hsp70 are able to function in vitro, after the heat-damaged substrate or substrates have been generated, neither protein is required to bin d to its target(s) during heating in order to effect repair.