Abstract
Proteasomes are multi-catalytic complexes with important roles in protein control. Their activity in stored red blood cells (RBCs) is affected by both storage time and the donor’s characteristics. However, apart from their abundancy in the membrane proteome, not much is known about their topology, activity, and networking during the storage of RBCs from beta-thalassemia trait donors (βThal+). For this purpose, RBC units from fourteen βThal+ donors were fractionated and studied for proteasome activity distribution and interactome through fluorometric and correlation analyses against units of sex- and aged-matched controls. In all the samples examined, we observed a time-dependent translocation and/or activation of the proteasome in the membrane and a tight connection of activity with the oxidative burden of cells. Proteasomes were more active in the βThal+ membranes and supernatants, while the early storage networking of 20S core particles and activities showed a higher degree of connectivity with chaperones, calpains, and peroxiredoxins, which were nonetheless present in all interactomes. Moreover, the βThal+ interactomes were specially enriched in kinases, metabolic enzymes, and proteins differentially expressed in βThal+ membrane, including arginase-1, piezo-1, and phospholipid scramblase. Overall, it seems that βThal+ erythrocytes maintain a considerable “proteo-vigilance” during storage, which is closely connected to their distinct antioxidant dynamics and membrane protein profile.
Highlights
IntroductionProteasomes are supramolecular, multi-catalytic complexes tasked with the degradation of aberrant and damaged proteins, which is needed to ensure cell homeostasis [1]
Intracellular and extracellular proteasome activities vary as a function of the cytosolic reactive oxygen species (ROS) levels, the storage age of red blood cells (RBCs), and the presence of βThal+ mutations
Storage is associated with decreased proteasome activity in the cytosol but increased activity in the membrane and the extracellular supernatant
Summary
Proteasomes are supramolecular, multi-catalytic complexes tasked with the degradation of aberrant and damaged proteins, which is needed to ensure cell homeostasis [1]. As such, their proteolytic core, the 20S complex, is equipped with three proteolytic activities: caspase (CASP)-like, trypsin (TR)-like, and chymotrypsin (CH)-like, the active centers of which are located at the β1, β2, and β5 subunits of the catalytic chamber, respectively. Misfolded proteins are transferred in the 20S cylinder after being selected by 19S regulatory complexes that recognize ubiquitinated molecules [2]. While the function of 26S holoenzymes is ATP- and ubiquitin-dependent, the 20S proteasome requires neither [3,4]
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