Persulfidome of Sweet Pepper Fruits during Ripening: The Case Study of Leucine Aminopeptidase That Is Positively Modulated by H2S.

Protein persulfidation is a thiol-based oxidative posttranslational modification (oxiPTM) that involves the modification of susceptible cysteine thiol groups present in peptides and proteins through hydrogen sulfide (H2S), thus affecting their function. Using sweet pepper (Capsicum annuum L.) fruits as a model material at different stages of ripening (immature green and ripe red), endogenous persulfidated proteins (persulfidome) were labeled using the dimedone switch method and identified using liquid chromatography and mass spectrometry analysis (LC-MS/MS). A total of 891 persulfidated proteins were found in pepper fruits, either immature green or ripe red. Among these, 370 proteins were exclusively present in green pepper, 237 proteins were exclusively present in red pepper, and 284 proteins were shared between both stages of ripening. A comparative analysis of the pepper persulfidome with that described in Arabidopsis leaves allowed the identification of 25% of common proteins. Among these proteins, glutathione reductase (GR) and leucine aminopeptidase (LAP) were selected to evaluate the effect of persulfidation using an in vitro approach. GR activity was unaffected, whereas LAP activity increased by 3-fold after persulfidation. Furthermore, this effect was reverted through treatment with dithiothreitol (DTT). To our knowledge, this is the first persulfidome described in fruits, which opens new avenues to study H2S metabolism. Additionally, the results obtained lead us to hypothesize that LAP could be involved in glutathione (GSH) recycling in pepper fruits.

Analysis of sulfide signaling in rice highlights specific drought responses.

Hydrogen sulfide regulates essential plant processes, including adaptation responses to stress situations, and the best characterized mechanism of action of sulfide consists of the posttranslational modification of persulfidation. In this study, we reveal the first persulfidation proteome described in rice including 3443 different persulfidated proteins that participate in a broad range of biological processes and metabolic pathways. In addition, comparative proteomics revealed specific proteins involved in sulfide signaling during drought responses. Several proteins involved in the maintenance of cellular redox homeostasis, the TCA cycle and energy-related pathways, and ion transmembrane transport and cellular water homeostasis, highlighting the aquaporin family, showed the highest differential levels of persulfidation. We revealed that water transport activity is regulated by sulfide which correlates to an increasing level of persulfidation of aquaporins. Our findings emphasize the impact of persulfidation on total ATP levels, fatty acid composition, ROS levels, antioxidant enzymatic activities, and relative water content. Interestingly, the persulfidation role on aquaporin transport activity as an adaptation response in rice differs from the current knowledge in Arabidopsis, which emphasizes the distinct role of sulfide improving rice tolerance to drought.