The gene for Hsp30, the only known α-crystallin-related heat shock protein of Neurospora crassa, was disrupted by repeat-induced point mutagenesis, leading to loss of cell survival at high temperature. Hsp30, which is not synthesized at 30°C, associates reversibly with the mitochondria at high temperature (45°C). In this study, we found that import of selected proteins into internal compartments of mitochondria, following their synthesis in the cytosol, was severely impaired at high temperature in a strain mutant in Hsp30. After 70 min of cell incubation at 45°C, most matrix, inner membrane, and intermembrane-space proteins tested were reduced in import by about 50-70% in the mutant, as compared to wild-type cells. In contrast, assembly of selected proteins into the outer mitochondrial membrane was not reduced, except for one component of the preprotein translocase complex of the mitochondrial outer membrane. Three proteins of this complex co-immunoprecipitated with Hsp30 of wild-type cells incubated at 45°C. We propose that Hsp30 interacts with the preprotein translocase of the mitochondrial outer membrane and that it chaperones the activity of one or more components of this translocase complex at high temperature.
Bibliographical noteFunding Information:
We are grateful to Michael P. Yaffe for the gift of the antibody to Saccharomyces ATPase F1-β subunit and Carmen A. Mannella for antibodies to porin and the outer mitochondrial membrane proteins. This research was supported by the National Institute of General Medical Sciences (GM-19398) and the National Research Initiative of the USDA Competitive Research Grants Office (94-37100-0290).
- Heat shock
- Neurospora crassa
- Protein import