Direct evidence for the intracellular localization of Hsp104 in Saccharomyces cerevisiae by immunoelectron microscopy

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Cell Stress Society International

RESUMO

To reveal the intracellular localization of Hspl04 in the yeast Saccharomyces cerevisiae before and after heat-shock, we performed immunoelectron microscopy after immunogold labeling with anti-Hsp104 antibody. At normal temperature (25°C), a small amount of Hspl04 was located in the cytoplasm and nucleus. On exposure to mild heat-shock at 40°C, protein aggregates appeared in the cytoplasm and nucleus, and Hspl04 increased around the aggregates with increasing time of the mild heat-shock treatment. Moreover, at lethal heat-shock temperature (51°C) for 20 min after mild heat treatment at 40°C, the intracellular localization of Hsp104 and intracellular structures were similar to those of the mild heat-shocked cells. However, in the lethally heat-shocked cells, certain intracellular structures were destroyed, and Hsp104 was not expressed. In the hsp104 null mutant strain Δhsp104 which was treated at 40°C, Hsp104 was not localized around the aggregates. Additionally, in the Δhsp104 strain, even mild heat-shocked cells at 37°C or 4O°C, showed destruction of intracellular structure compared to the wild-type strain. Our data suggest the following: (1) Hsp104 is associated closely with protein aggregates during heat-shock treatment, (2) Hsp104 is important for maintenance of the intracellular structure under lethal heat-shock conditions, (3) acquisition of thermotolerance depends on the amount of Hsp104 produced during mild heat-shock treatment.

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