Tenuazonic acid from Stemphylium loti inhibits the plant plasma membrane H(+)‐ATPase by a mechanism involving the C‐terminal regulatory domain

Pathogenic fungi often target the plant plasma membrane (PM) H(+)‐ATPase during infection. To identify pathogenic compounds targeting plant H(+)‐ATPases, we screened extracts from 10 Stemphylium species for their effect on H(+)‐ATPase activity. We identified Stemphylium loti extracts as potential H(+)‐ATPase inhibitors, and through chemical separation and analysis, tenuazonic acid (TeA) as a potent H(+)‐ATPase inhibitor. By assaying ATP hydrolysis and H(+) pumping, we confirmed TeA as a H(+)‐ATPase inhibitor both in vitro and in vivo. To visualize in planta inhibition of the H(+)‐ATPase, we treated pH‐sensing Arabidopsis thaliana seedlings with TeA and quantified apoplastic alkalization. TeA affected both ATPase hydrolysis and H(+) pumping, supporting a direct effect on the H(+)‐ATPase. We demonstrated apoplastic alkalization of A. thaliana seedlings after short‐term TeA treatment, indicating that TeA effectively inhibits plant PM H(+)‐ATPase in planta. TeA‐induced inhibition was highly dependent on the regulatory C‐terminal domain of the plant H(+)‐ATPase. Stemphylium loti is a phytopathogenic fungus. Inhibiting the plant PM H(+)‐ATPase results in membrane potential depolarization and eventually necrosis. The corresponding fungal H(+)‐ATPase, PMA1, is less affected by TeA when comparing native preparations. Fungi are thus able to target an essential plant enzyme without causing self‐toxicity.