ATP2B3 Inhibition Alleviates Erastin-Induced Ferroptosis in HT-22 Cells through the P62-KEAP1-NRF2-HO-1 Pathway

Shihui Guo; Aiying Zhong; Dongxu Zhang; Jiang Gao; Yingdong Ni; Ruqian Zhao; Wenqiang Ma

doi:10.3390/ijms24119199

ATP2B3 Inhibition Alleviates Erastin-Induced Ferroptosis in HT-22 Cells through the P62-KEAP1-NRF2-HO-1 Pathway

Int J Mol Sci. 2023 May 24;24(11):9199. doi: 10.3390/ijms24119199.

Authors

Shihui Guo^{1

2}, Aiying Zhong^{1

2}, Dongxu Zhang^{1

2}, Jiang Gao^{1

2}, Yingdong Ni^{1

2}, Ruqian Zhao^{1

2}, Wenqiang Ma^{1

2}

Affiliations

¹ Key Laboratory of Animal Physiology and Biochemistry, Ministry of Agriculture and Rural Affairs, College of Veterinary Medicine, Nanjing Agricultural University, Nanjing 210095, China.
² MOE Joint International Research Laboratory of Animal Health & Food Safety, Nanjing Agricultural University, Nanjing 210095, China.

Abstract

Ferroptosis participates in the occurrence and development of neurological disorders. Modulating ferroptosis may have therapeutic potential in nervous system diseases. Therefore, TMTbased proteomic analysis in HT-22 cells was performed to identify erastin-induced differentially expressed proteins. The calcium-transporting ATP2B3 (ATP2B3) was screened as a target protein. ATP2B3 knockdown markedly alleviated the erastin-induced decrease in cell viability and elevated ROS (p < 0.01) and reversed the up-regulation of oxidative stress-related proteins polyubiquitin-binding protein p62 (P62), nuclear factor erythroid 2-related factor2 (NRF2), heme oxygenase-1 (HO-1), and NAD(P)H quinone oxidoreductase-1 (NQO1) protein expression (p < 0.05 or p < 0.01) and the down-regulation of Kelch-like ECH-associated protein 1(KEAP1) protein expression (p < 0.01). Moreover, NRF2 knockdown, P62 inhibition, or KEAP1 overexpression rescued the erastin-induced decrease in cell viability (p < 0.05) and increase in ROS production (p < 0.01) in HT-22 cells, while simultaneous overexpression of NRF2 and P62 and knockdown of KEAP1 partially offset the relief effect of ATP2B3 inhibition. In addition, knockdown of ATP2B3, NRF2, and P62 and overexpression of KEAP1 significantly down-regulated erastin-induced high expression of the HO-1 protein, while HO-1 overexpression reversed the alleviating effects of ATP2B3 inhibition on the erastin-induced decrease in cell viability (p < 0.01) and increase in ROS production (p < 0.01) in HT-22 cells. Taken together, ATP2B3 inhibition mediates the alleviation of erastin-induced ferroptosis in HT-22 cells through the P62-KEAP1-NRF2-HO-1 pathway.

Keywords: ATP2B3; HO-1; HT-22; P62–KEAP1–NRF2; ferroptosis.

MeSH terms

Ferroptosis*
Heme Oxygenase-1 / genetics
Heme Oxygenase-1 / metabolism
Kelch-Like ECH-Associated Protein 1 / genetics
Kelch-Like ECH-Associated Protein 1 / metabolism
NF-E2-Related Factor 2* / genetics
NF-E2-Related Factor 2* / metabolism
Oxidative Stress
Proteomics
Reactive Oxygen Species / metabolism
Signal Transduction

Substances

Kelch-Like ECH-Associated Protein 1
NF-E2-Related Factor 2
erastin
Reactive Oxygen Species
Heme Oxygenase-1

Grants and funding

This work was supported by grants from the National Key Research and Development Program of China (No. 2017YFE0129900), the National Natural Science Foundation of China (No. 31872439), the Priority Academic Program Development of Jiangsu Higher Education Institutions, and the Jiangsu Collaborative Innovation Center of Meat Production and Processing, Quality and Safety Control.