Proteome Analysis of the ROF-FKBP Mutants Reveals Functional Relations among Heat Stress Responses, Plant Development, and Protein Quality Control during Heat Acclimation in Arabidopsis thaliana

In the present study, a differential screening following heat stress acclimation was performed in Arabidopsis thaliana WT and ROF-FKBP mutated plants using mass spectrometry, and the results were used to understand and analyze the effect of the ROF PPIases during thermotolerance acquisition in plants. Our data highlight the central role of these two PPIases in heat stress and point to their direct or indirect effect on other proteins participating in cellular functions such as protein folding and quality control, cell division, photosynthesis, and other metabolic and signaling processes. Specifically, the heat stress response, protein folding, and protein ER processing pathways are enhanced following a 37 °C acclimation period independent of the mutation state. However, at 37 °C, and in the double-mutated rof1^–/2^– plants, a higher accumulation of proteins belonging to the above pathways is observed compared with all other conditions (WT, single mutants, control, and heat-acclimated plants). Furthermore, the proteasomal pathway, involving the common member of both the protasomal and the lysosomal degradation pathway, CDC48, is over-represented in the extracts of both the untreated and heat-stressed rof1^–/2^– mutants compared with the other extracts. In contrast, in the single rof1^– mutation, the heat acclimation pathway is suppressed at 37 °C when compared to the WT. Protein accumulation related to the heat stress and the protein quality control pathways points to a differential but also synergistic role of the two proteins. Protein complexes of other biochemical and developmental mechanisms, such as the light-harvesting complex of the photosynthetic pathway and the phosphoinositide binding proteins involved in membrane-trafficking events during cell plate formation and cytokinesis (patellin 1, 2, and 4), are negatively regulated in the rof1^–/2^– mutant. Our results suggest that ROF1 and ROF2 FKBPs regulate stress response, and developmental and metabolic pathways via a complex feedback mechanism involving partners that ensure protein quality control and plant survival during heat stress.