Archaeal Hsp14 drives substrate shuttling between small heat shock proteins and thermosome: insights into a novel substrate transfer pathway

Heat shock proteins maintain protein homeostasis and facilitate the survival of an organism under stress. Archaeal heat shock machinery usually consists of only sHsps, Hsp70, and Hsp60. Moreover, Hsp70 is absent in thermophilic and hyperthermophilic archaea. In the absence of Hsp70, how aggregating protein substrates are transferred to Hsp60 for refolding remains elusive. Here, we investigated the crosstalk in the heat shock response pathway of thermoacidophilic crenarchaeon Sulfolobus acidocaldarius. In the present study, we biophysically and biochemically characterized one of the small heat shock proteins, Hsp14, of S. acidocaldarius. Moreover, we investigated its ability to interact with Hsp20 and Hsp60 to facilitate the substrate proteins' folding under stress conditions. Like Hsp20, we demonstrated that the dimer is the active form of Hsp14, and it forms an oligomeric storage form at a higher temperature. More importantly, the dynamics of the Hsp14 oligomer are maintained by rapid subunit exchange between the dimeric states, and the rate of subunit exchange increases with increasing temperature. We also tested the ability of Hsp14 to form hetero-oligomers via subunit exchange with Hsp20. We observed hetero-oligomer formation only at higher temperatures (50 degrees C-70 degrees C). Furthermore, experiments were performed to investigate the interaction between small heat shock proteins and Hsp60. We demonstrated an enthalpy-driven direct physical interaction between Hsp14 and Hsp60. Our results revealed that Hsp14 could transfer sHsp-captured substrate proteins to Hsp60, which then refolds them back to their active form.

Automated summary

The study investigated the crosstalk in the heat shock response pathway of thermoacidophilic crenarchaeon Sulfolobus acidocaldarius, focusing on the small heat shock protein Hsp14. It was found that Hsp14 interacts with Hsp20 and Hsp60 to facilitate substrate proteins' folding under stress conditions, demonstrating the dynamic oligomeric structure of Hsp14 and its ability to form hetero-oligomers with Hsp20 at higher temperatures. Moreover, enthalpy-driven direct physical interaction between Hsp14 and Hsp60 was revealed, showing that Hsp14 can transfer substrate proteins captured by sHsp to Hsp60 for refolding.