The plant cysteine oxidases from Arabidopsis thaliana are kinetically tailored to act as oxygen sensors

Group VII ethylene response factors (ERF-VIIs) regulate transcriptional adaptation to flooding-induced hypoxia in plants. ERF-VII stability is controlled in an O(2)-dependent manner by the Cys/Arg branch of the N-end rule pathway whereby oxidation of a conserved N-terminal cysteine residue initiates target degradation. This oxidation is catalyzed by plant cysteine oxidases (PCOs), which use O(2) as cosubstrate to generate Cys-sulfinic acid. The PCOs directly link O(2) availability to ERF-VII stability and anaerobic adaptation, leading to the suggestion that they act as plant O(2) sensors. However, their ability to respond to fluctuations in O(2) concentration has not been established. Here, we investigated the steady-state kinetics of Arabidopsis thaliana PCOs 1–5 to ascertain whether their activities are sensitive to O(2) levels. We found that the most catalytically competent isoform is AtPCO4, both in terms of responding to O(2) and oxidizing AtRAP2.2/2,12 (two of the most prominent ERF-VIIs responsible for promoting the hypoxic response), which suggests that AtPCO4 plays a central role in ERF-VII regulation. Furthermore, we found that AtPCO activity is susceptible to decreases in pH and that the hypoxia-inducible AtPCOs 1/2 and the noninducible AtPCOs 4/5 have discrete AtERF-VII substrate preferences. Pertinently, the AtPCOs had K(m(O2))(app) values in a physiologically relevant range, which should enable them to sensitively react to changes in O(2) availability. This work validates an O(2)-sensing role for the PCOs and suggests that differences in expression pattern, ERF-VII selectivity, and catalytic capability may enable the different isoforms to have distinct biological functions. Individual PCOs could therefore be targeted to manipulate ERF-VII levels and improve stress tolerance in plants.