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p97 dysfunction underlies a loss of quality control of damaged membrane proteins and promotes oxidative stress and sickling in sickle cell disease
Author(s) -
Song Anren,
Wen Alexander Q.,
Wen Y. Edward,
Dzieciatkowska Monika,
Kellems Rodney E.,
Juneja Harinder S.,
D'Alessandro Angelo,
Xia Yang
Publication year - 2022
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fj.202101500rr
Subject(s) - oxidative stress , microbiology and biotechnology , membrane protein , ubiquitin , proteostasis , reactive oxygen species , cytosol , biology , ubiquitin ligase , proteasome , mitochondrion , biochemistry , membrane , enzyme , gene
Abstract Sickling is the central pathogenic process of sickle cell disease (SCD), one of the most prevalent inherited hemolytic disorders. Having no easy access to antioxidants in the cytosol, elevated levels of reactive oxygen species (ROS) residing at the plasma membrane in sickle red blood cells (sRBCs) easily oxidize membrane proteins and thus contribute to sickling. Although the ubiquitin‐proteasome system (UPS) is essential to rapidly clear ROS‐damaged membrane proteins and maintain cellular homeostasis, the function and regulatory mechanism of the UPS for their clearance in sRBCs remains unidentified. Elevated levels of polyubiquitinated membrane‐associated proteins in human sRBCs are reported here. High throughput and untargeted proteomic analyses of membrane proteins immunoprecipitated by ubiquitin antibodies detected elevated levels of ubiquitination of a series of proteins including cytoskeletal proteins, transporters, ROS‐related proteins, and UPS machinery components in sRBCs. Polyubiquitination of membrane‐associated catalase was increased in sRBCs, associated with decreased catalase activity and elevated ROS. Surprisingly, shuttling of p97 (ATP‐dependent valosin‐containing chaperone protein), a key component of the UPS to shuttle polyubiquitinated proteins from the membrane to cytosol for proteasomal degradation, was significantly impaired, resulting in significant accumulation of p97 along with polyubiquitinated proteins in the membrane of human sRBCs. Functionally, inhibition of p97 directly promoted accumulation of polyubiquitinated membrane‐associated proteins, excessive ROS levels, and sickling in response to hypoxia. Overall, we revealed that p97 dysfunction underlies impaired UPS and contributes to oxidative stress in sRBCs.