ИСТИНА

Side-by-side with passive uptake and release of oxygen and metabolically-derived gases erythrocytes exhibit diverse oxygen- sensitive responses involved in regulation of their own and neigh- boring cell functions. To goal of this study was to determine if the attenuated integrity of deoxygenated RBC could be linked to any changes in the composition erythrocyte membrane. Twenty- min exposure to oxygen-free environment decreases RBC integ- rity documented by 3-fold elevation of hemoglobin release. Elec- tron magnetic resonance spectroscopy of spin-labeled stearic acid analogues did not detect any action of hypoxia on the membrane fluidity. The proteomics technology in combination with relative label free quantification analysis allowed us to compare the con- tent of 1176 membrane-bound proteins in the ghosts obtained normoxic and hypoxic RBC. Using this approach we found that sustained deoxygenation of rat erythrocytes alters the composi- tion of membrane-bound proteins including 2–3 fold elevation of the content of hemoglobin. Mapping the identified proteins in the KEGG pathway database we found that the proteins of multi subunit Cullin-Rbx E3 ubiquitin ligase complex presented in normoxic RBC ghosts but not in the hypoxic samples. We hypothesized that Cullin-Rbx E3 complex, associated with RBC membrane in normoxia. In hypoxic conditions, deoxy-Hb binds to band 3 protein, resulting in dissociation of Cullin-Rbx E3 complex from RBC membrane and impaired clearance of damaged cytoskeleton proteins. We believe that these changes underlie attenuated integrity of RBC membrane and contribute to augment release of hemoglobin and ATP seen in hypoxic conditions. This work was supported by grants from the Russian Foundation for Basic Research (#18-04-00063) and the Russian Scientific Foundation (#16-15-10026).