Overexpression of miR399d impairs arbuscular mycorrhizal symbiosis in tomato.

Arbuscular mycorrhizal symbiosis (AMS) is a pervasive mutualistic interaction that is prevalent among fungi and the majority of terrestrial plant species. AMS host plants possess an alternative phosphate (Pi) acquisition pathway via arbuscular mycorrhizal fungi (AMF) in addition to direct Pi uptake by the root epidermis. In the present study, we found that miR399d homologues were consistently downregulated in multiple angiosperms during AMS. Genetic approaches were used to study its role in AMS in a tomato model. The overexpression of tomato miR399d significantly inhibited the colonization of plants by AMF and the development of arbuscules. A similar phenotype was observed by inactivation of PHO2 (PHOSPHATE2), a target gene of miR399d. Considering that both miR399d overexpression and PHO2 deficiency increase the accumulation of Pi transporters in the direct Pi uptake pathway, a Pi transporter gene PT1, which is involved in direct Pi uptake, was overexpressed in tomato. The resulting transgenic plants presented elevated direct Pi uptake and a decreased degree of AMF colonization. These findings suggest that the downregulation of miR399d is required for AMS establishment and that miR399d may act as a negative regulator of AMS by fine-tuning distinct Pi uptake pathways in tomato plants under phosphorus starvation conditions.