Selective Metabolism of Hypothiocyanous Acid by Mammalian Thioredoxin Reductase Promotes Lung Innate Immunity and Antioxidant Defense

• Published in: The Journal of biological chemistry Vol. 288; no. 25; pp. 18421 - 18428
• Main Authors: Chandler, Joshua D., Nichols, David P., Nick, Jerry A., Hondal, Robert J., Day, Brian J.
• Format: Journal Article
• Language: English
• Published: United States Elsevier Inc 21-06-2013; American Society for Biochemistry and Molecular Biology
• Subjects: Amino Acid Substitution; Animals; Antioxidants - metabolism; Auranofin - pharmacology; Bacterial Proteins - antagonists & inhibitors; Bacterial Proteins - genetics; Bacterial Proteins - metabolism; Bronchioles - cytology; Cell Survival - drug effects; Cells, Cultured; Cysteine - genetics; Cysteine - metabolism; Cystic fibrosis; Enzymology; Epithelial Cells - drug effects; Epithelial Cells - metabolism; Humans; Hypochlorous Acid - pharmacology; hypothiocyanite; Immunity, Innate; Infectious diseases; Inflammation; Innate immunity; Kinetics; Lung - immunology; Lung - metabolism; Mice; Microbial Viability - drug effects; Rats; Selenocysteine - genetics; Selenocysteine - metabolism; Substrate Specificity; thiocyanate; Thiocyanates - metabolism; Thiocyanates - pharmacology; Thioredoxin reductase; Thioredoxin-Disulfide Reductase - antagonists & inhibitors; Thioredoxin-Disulfide Reductase - genetics; Thioredoxin-Disulfide Reductase - metabolism; Infectious diseases; hypothiocyanite; Cystic fibrosis; Innate immunity; Inflammation; Thioredoxin reductase; thiocyanate
• ISSN: 0021-9258; 1083-351X
• DOI: 10.1074/jbc.M113.468090

The endogenously produced oxidant hypothiocyanous acid (HOSCN) inhibits and kills pathogens but paradoxically is well tolerated by mammalian host tissue. Mammalian high molecular weight thioredoxin reductase (H-TrxR) is evolutionarily divergent from bacterial low molecular weight thioredoxin reductase (L-TrxR). Notably, mammalian H-TrxR contains a selenocysteine (Sec) and has wider substrate reactivity than L-TrxR. Recombinant rat cytosolic H-TrxR1, mouse mitochondrial H-TrxR2, and a purified mixture of both from rat selectively turned over HOSCN (kcat = 357 ± 16 min−1; Km = 31.9 ± 10.3 μm) but were inactive against the related oxidant hypochlorous acid. Replacing Sec with Cys or deleting the final eight C-terminal peptides decreased affinity and turnover of HOSCN by H-TrxR. Similarly, glutathione reductase (an H-TrxR homologue lacking Sec) was less effective at HOSCN turnover. In contrast to H-TrxR and glutathione reductase, recombinant Escherichia coli L-TrxR was potently inhibited by HOSCN (IC50 = 2.75 μm). Similarly, human bronchial epithelial cell (16HBE) lysates metabolized HOSCN, but E. coli and Pseudomonas aeruginosa lysates had little or no activity. HOSCN selectively produced toxicity in bacteria, whereas hypochlorous acid was nonselectively toxic to both bacteria and 16HBE. Treatment with the H-TrxR inhibitor auranofin inhibited HOSCN metabolism in 16HBE lysates and significantly increased HOSCN-mediated cytotoxicity. These findings demonstrate both the metabolism of HOSCN by mammalian H-TrxR resulting in resistance to HOSCN in mammalian cells and the potent inhibition of bacterial L-TrxR resulting in cytotoxicity in bacteria. These data support a novel selective mechanism of host defense in mammals wherein HOSCN formation simultaneously inhibits pathogens while sparing host tissue.
Secreted hypothiocyanous acid (HOSCN) kills pathogens but paradoxically is tolerated by many mammalian cells.
Mammalian thioredoxin reductase (H-TrxR) reduces HOSCN, whereas bacterial L-TrxR is inhibited by it, corresponding to differential cytotoxicity.
Mammalian H-TrxR confers resistance against HOSCN, enabling its use as a selective biocide.
Findings directly link mammalian H-TrxR to innate immunity and inflammatory lung disease.