An ancient route towards salicylic acid and its implications for the perpetual Trichormus-Azolla symbiosis.

Despite its small size, the water fern Azolla is a giant among plant symbioses. Within each of its leaflets, a specialized leaf cavity is home to a population of nitrogen-fixing cyanobacteria (cyanobionts). Although a number of plant-cyanobiont symbioses exist, Azolla is unique in that its symbiosis is perpetual: the cyanobionts are inherited during sexual and vegetative propagation. What underpins the communication between the two partners? In angiosperms, the phytohormone salicylic acid (SA) is a well-known regulator of plant-microbe interactions. Using high-performance liquid chromatography-tandem mass spectrometry, we pinpoint the presence of SA in the fern. Comparative genomics and phylogenetics on SA biosynthesis genes across Chloroplastida reveal that the entire Phenylalanine ammonia-lyase-dependent pathway likely existed in the last common ancestor of land plants. Indeed, Azolla filiculoides secondarily lost its isochorismate synthase but has the genetic competence to derive SA from benzoic acid; the presence of SA in artificially cyanobiont-free Azolla supports the existence of this route. Global gene expression data and SA levels from cyanobiont-containing and -free A. filiculoides link SA synthesis with the symbioses: SA appears to induce cyanobacterial proliferation, whereas removal of the symbiont results in reduced SA levels in a nitrogen-dependent manner.