Medicago SPX1 and SPX3 recruit nitrogen-fixing microbes to promote growth through controlling flavonoid biosynthesis

Forming symbiotic associations with beneficial microbes are important strategies for sessile plants to acquire nitrogen and phosphorus nutrients from the soil. Root exudates play key roles on set-up of the rhizosphere microbiome. According to the needs for nitrogen or phosphorus, plants can adjust the root exudates composition to attract proper microbes. Flavonoids are a group of secondary metabolites that are well studied in shaping the root microbiome, especially the root nodule symbiosis in legumes. Here, we show the medicago truncatula phosphate sensors SPX1 and SPX3 regulate flavonoids biosynthesis to recruit nitrogen-fixing microbes for nitrogen acquisition. Nitrogen-fixing microbes were less recruited in spx1spx3 double mutant root rhizosphere. This was caused by lower flavonoids biosynthesis related genes expression, which resulted in lower flavonoids levels in the root exudates compared to wild type plant R108. Further analysis indicates the regulation of flavonoids biosynthesis is through the SPX1 and SPX3 interaction transcription factor PHR2. We propose the SPX-PHR phosphate homeostasis regulation network also control microbe-dependent nitrogen acquisition according to phosphate levels. Thus, SPX1 and SPX3 play important roles to keep a microbe-dependent nitrogen and phosphorus absorption balance for optimal growth. Overall design: R108 wild type and spx1spx3 mutant roots grew in low Pi conditions