Pseudomonas aeruginosa utilizes host polyunsaturated phosphatidylethanolamines to trigger theft-ferroptosis in bronchial epithelium
Haider H. Dar, … , Hülya Bayır, Valerian E. Kagan
Haider H. Dar, … , Hülya Bayır, Valerian E. Kagan
Published October 1, 2018; First published September 10, 2018
Citation Information: J Clin Invest. 2018;128(10):4639-4653. https://doi.org/10.1172/JCI99490.
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Categories: Research Article Cell biology Infectious disease

Pseudomonas aeruginosa utilizes host polyunsaturated phosphatidylethanolamines to trigger theft-ferroptosis in bronchial epithelium

Abstract

Ferroptosis is a death program executed via selective oxidation of arachidonic acid–phosphatidylethanolamines (AA-PE) by 15-lipoxygenases. In mammalian cells and tissues, ferroptosis has been pathogenically associated with brain, kidney, and liver injury/diseases. We discovered that a prokaryotic bacterium, Pseudomonas aeruginosa, that does not contain AA-PE can express lipoxygenase (pLoxA), oxidize host AA-PE to 15-hydroperoxy-AA-PE (15-HOO-AA-PE), and trigger ferroptosis in human bronchial epithelial cells. Induction of ferroptosis by clinical P. aeruginosa isolates from patients with persistent lower respiratory tract infections was dependent on the level and enzymatic activity of pLoxA. Redox phospholipidomics revealed elevated levels of oxidized AA-PE in airway tissues from patients with cystic fibrosis (CF) but not with emphysema or CF without P. aeruginosa. We believe that the evolutionarily conserved mechanism of pLoxA-driven ferroptosis may represent a potential therapeutic target against P. aeruginosa–associated diseases such as CF and persistent lower respiratory tract infections.

Authors

Haider H. Dar, Yulia Y. Tyurina, Karolina Mikulska-Ruminska, Indira Shrivastava, Hsiu-Chi Ting, Vladimir A. Tyurin, James Krieger, Claudette M. St. Croix, Simon Watkins, Erkan Bayir, Gaowei Mao, Catherine R. Armbruster, Alexandr Kapralov, Hong Wang, Matthew R. Parsek, Tamil S. Anthonymuthu, Abiola F. Ogunsola, Becca A. Flitter, Cody J. Freedman, Jordan R. Gaston, Theodore R. Holman, Joseph M. Pilewski, Joel S. Greenberger, Rama K. Mallampalli, Yohei Doi, Janet S. Lee, Ivet Bahar, Jennifer M. Bomberger, Hülya Bayır, Valerian E. Kagan

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Figure 1

pLoxA is required for P. aeruginosa supernatant–induced ferroptosis.

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pLoxA is required for P. aeruginosa supernatant–induced ferroptosis. (A)...
(A) HBE cells were treated with supernatants (10 μg each) from WT, ΔwspF, or pLoxA-deficient mutant loxA::Tn (PW3111) with or without ferrostatin-1 (FER, 0.2 μM). Cell death (20 hours) assessed by PI; mean ± SD, *P < 0.05 vs. control (untreated HBE cells), #P < 0.05 vs. corresponding no FER treatment; n = 3. (B) ΔwspF supernatant alone or with z-VAD-fmk (20 μM), necrostatin-1s (Nec-1s, 20 μM), bafilomycin-A1 (Baf-A1, 1 nM). FER, positive control; mean ± SD, *P < 0.05 vs. control (untreated), #P < 0.05 vs. ΔwspF only; n = 3. (C–K) Representative fixed biofilms on glass coverslips stained with anti-pLoxA antibodies (green) (D, G, and J) or analyzed by SEM (E, H, and K) (of 3 performed). (L) Dioxygenase activity: WT, ΔwspF, or loxA::Tn supernatants were incubated with DOPC/AA (1:10) or DOPC/SAPE (1:1) liposomes (10 minutes, 37°C). AA oxidation product 15-HpETE (left panel) or 15-HpETE-PE from AA-PE (right panel) assessed by LC-MS; normalized to bacteria (bac.)/ml of supernatant; *P < 0.05 vs. loxA::Tn supernatant; n = 3. (M) Effect of 15LOX-specific inhibitors (PD146176 and ML351; 1.0 μM) on ΔwspF ferroptosis. RSL3 (200 nM, left panel) was a positive control (both inhibitors were effective against host 15LOX); mean ± SD, #P < 0.05 vs. control (untreated), *P < 0.05 vs. RSL3 or ΔwspF supernatant; n = 3. (N) Bacterial cell lysates (pLoxA-deficient or complemented, 100 μg each) were incubated with SAPE (100 μM, 30 minutes) and then added to RSL3-pretreated (20 nM) HBE cells with or without FER (0.2 μM). Mean ± SD,*P < 0.05 vs. RSL3, #P < 0.05 vs. no FER PW3111 Tn7-loxA; n = 3. (O) HBE cells were incubated with supernatant from MJK8 or its loxA-KO strain (MJK8ΔloxA) in the presence or absence of FER (0.2 μM). Mean ± SD, *P < 0.05 vs. control (untreated HBE cells), #P < 0.05 vs. no FER MJK8 supernatant; n = 3. One-way ANOVA for A, B, and L–O.