Updated project metadata. The PII family comprises a group of widely distributed signal transduction proteins ubiquitous present in prokaryotes and in the chloroplast of plants. The PII proteins sense the levels of key metabolites ATP, ADP and 2-oxoglutarate which affect the PII structure and the ability of PII to interact with a range of target proteins. Here we performed multiple PII ligand fishing assays to identify proteins that are likely to be part of the PII protein interaction network in plant-growth promoting nitrogen-fixing bacterium Azospirillum brasilense. Among the PII targets identified were enzymes related to nitrogen and fatty acid metabolism, signaling, co-enzyme synthesis, RNA catabolism and transcription. Direct binary PII-target complex was confirmed for xx protein complex using pull-down assays with recombinant proteins. Untargeted metabolome showed that PII is required to proper maintenance of Coenzyme A, panthotenate, PRPP and SAICAR levels. Two enzymes involved in c-di-GMP metabolism were among the identified PII targets. A PII minus strain showed reduced c-di-GMP levels and swimming behavior. These data support that PII acts as a major metabolic hub controlling key metabolic enzymes and c-di-GMP homeostasis accordingly to the prevailing nutritional status.