Studies on the mechanism of HSA32-mediated long-term acquired thermotolerance and HSP101 degradation in Arabidopsis

Heat acclimation enhances plant thermotolerance against severe heat stress, a phenomenon known as acquired thermotolerance. Previous research demonstrated that Arabidopsis and rice seedlings of HSA32 knockout (hsa32) mutant have defect in long-term acquired thermotolerance (LAT), but retain normal short-term acquired thermotolerance (SAT). It was shown that HSA32 regulates LAT by specifically preventing HSP101 from degradation. However, the mechanisms of HSP101 degradation and HSA32-mediated HSP101 stability remain unclear. Proteolysis can be categorized into 26S proteasome and autophagy pathways. The strong 26S proteasome inhibitor bortezomib and autophagy related mutant atg5 were used to specifically block proteolysis pathways to investigate which route is responsible for the degradation of HSP101. The results demonstrated that blocking either pathway in the absence of HSA32 resulted in partial suppression of HSP101 degradation. However, when hsa32 atg5 double mutant was treated with bortezomib, the HSP101 level was restored to that of the wild type. Taken together, the results show that HSP101 undergoes post-stress degradation through both 26S proteasome and macroautophagy pathways, which is suppressed by HSA32.