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Adenosine receptors play pivotal roles in physiological processes. Adenosine A 3 receptor (A 3 R), the most recently identified adenosine receptor, is expressed in various tissues, exhibiting important roles in neuron, heart, and immune cells, and is often overexpressed in tumors, highlighting the therapeutic potential of A 3 R-selective agents. Recently, we identified RNA-derived N 6 -methyladenosine (m 6 A) as an endogenous agonist for A 3 R, suggesting the relationship between RNA-derived modified adenosine and A 3 R. Despite extensive studies on the other adenosine receptors, the selectivity mechanism of A 3 R, especially for A 3 R-selective agonists such as m 6 A and namodenoson, remained elusive. Here, we identify tRNA-derived N 6 -isopentenyl adenosine (i 6 A) as an A 3 R-selective ligand via screening of modified nucleosides against the adenosine receptors. Like m 6 A, i 6 A is found in the human body and may be an endogenous A 3 R ligand. Our cryo-EM analyses elucidate the A 3 R-G i complexes bound to adenosine, 5'-N-ethylcarboxamidoadenosine (NECA), m 6 A, i 6 A, and namodenoson at overall resolutions of 3.27 Å (adenosine), 2.86 Å (NECA), 3.19 Å (m 6 A), 3.28 Å (i 6 A), and 3.20 Å (namodenoson), suggesting the selectivity and activation mechanism of A 3 R. We further conduct structure-guided engineering of m 6 A-insensitive A 3 R, which may aid future research targeting m 6 A and A 3 R, providing a molecular basis for future drug discovery., (© 2024. The Author(s).)

Supersulphides are inorganic and organic sulphides with sulphur catenation with diverse physiological functions. Their synthesis is mainly mediated by mitochondrial cysteinyl-tRNA synthetase (CARS2) that functions as a principal cysteine persulphide synthase (CPERS). Here, we identify protective functions of supersulphides in viral airway infections (influenza and COVID-19), in aged lungs and in chronic lung diseases, including chronic obstructive pulmonary disease (COPD), idiopathic pulmonary fibrosis (IPF). We develop a method for breath supersulphur-omics and demonstrate that levels of exhaled supersulphides increase in people with COVID-19 infection and in a hamster model of SARS-CoV-2 infection. Lung damage and subsequent lethality that result from oxidative stress and inflammation in mouse models of COPD, IPF, and ageing were mitigated by endogenous supersulphides production by CARS2/CPERS or exogenous administration of the supersulphide donor glutathione trisulphide. We revealed a protective role of supersulphides in airways with various viral or chronic insults and demonstrated the potential of targeting supersulphides in lung disease., (© 2023. The Author(s).)

Cysteine hydropersulfide (CysSSH) occurs in abundant quantities in various organisms, yet little is known about its biosynthesis and physiological functions. Extensive persulfide formation is apparent in cysteine-containing proteins in Escherichia coli and mammalian cells and is believed to result from post-translational processes involving hydrogen sulfide-related chemistry. Here we demonstrate effective CysSSH synthesis from the substrate L-cysteine, a reaction catalyzed by prokaryotic and mammalian cysteinyl-tRNA synthetases (CARSs). Targeted disruption of the genes encoding mitochondrial CARSs in mice and human cells shows that CARSs have a crucial role in endogenous CysSSH production and suggests that these enzymes serve as the principal cysteine persulfide synthases in vivo. CARSs also catalyze co-translational cysteine polysulfidation and are involved in the regulation of mitochondrial biogenesis and bioenergetics. Investigating CARS-dependent persulfide production may thus clarify aberrant redox signaling in physiological and pathophysiological conditions, and suggest therapeutic targets based on oxidative stress and mitochondrial dysfunction.