Evidence for functionality of transmembrane domains of predicted non-canonical plant phosphotransmitters

Bioinformatic methods have been used to predict a new subclass of proteins among plant phosphotransmitters involved in the signaling system of multistep phosphorelay. In contrast to the canonical soluble nucleocytosolic forms, the found phosphotransmitter sequences, belonging to a wide range of plant taxa, potentially contain transmembrane domains. This suggests localization of such proteins on cell membranes and, therefore, a different function in signaling than that of canonical phosphotransmitters. We tested the functionality of the transmembrane domain of the phosphotransmitter using the protein of the tea plant Camellia sinensis. Membrane localization of the transiently expressed recombinant phosphotransmitter with this domain was confirmed by microscopy and immunoblotting. Thus, for the first time, experimental evidence was obtained for the existence of membrane-bound plant phosphotransmitters with as yet unknown functions. These data suggest the presence of a non-canonical membrane branch of signal transduction in the multistage phosphotransfer system in plants.