Peptide-Mediated Cyclic Nucleotide Signaling in Plants: Identification and Characterization of Interactor Proteins with Nucleotide Cyclase Activity

During the last decades, an increasing number of plant signaling peptides have been discovered and it appears that many of them are specific ligands for interacting receptor molecules. These receptors can enable the formation of second messengers which in turn transmit the ligand-induced stimuli into complex and tunable downstream responses. In order to perform such complex tasks, receptor proteins often contain several distinct domains such as a kinase and/or adenylate cyclase (AC) or guanylate cyclase (GC) domains. ACs catalyze the conversion of ATP to 3',5'-cyclic adenosine monophosphate (cAMP) while GCs catalyze the reaction of GTP to 3',5'-cyclic guanosine monophosphate (cGMP). Both cAMP and cGMP are now recognized as essential components of many plant responses, including responses to peptidic hormones. Here we describe the approach that led to the discovery of the Plant Natriuretic Peptide Receptor (PNP receptor), including a protocol for the identification of currently undiscovered peptidic interactions, and the subsequent application of computational methods for the identification of AC and/or GC domains in such interacting receptor candidates.