The Common Bean miR172c microRNA, a Relevant Regulator of the N‐Fixing Symbiosis, Is Activated by SPL and AGL/MADS‐Domain Transcription Factors

Nitrogen‐fixing symbiosis (NFS) between rhizobia and legume plants is a complex and tightly regulated process. Modules of microRNAs (miRNAs) and their targets from different legumes, such as miR156‐SPL and miR172‐AP2‐1, form part of complex cascades relevant for the regulation of NFS. Based on our previous analysis of the regulatory role of the common bean ( Phaseolus vulgaris ) miR172c‐AP2‐1 module, as well as in reports from other legumes about the transcriptional regulation of MIR172 in the NFS, in this work, we demonstrated that PvMIR172c is transcriptionally activated by PvSPL9 and PvFUL‐like transcription factors from the SQUAMOSA PROMOTER BINDING PROTEIN‐LIKE and AGL (AGAMOUS‐LIKE/MADS domain) families, respectively. Bioinformatic analysis of the PvMIR172c gene promoter revealed statistically enriched binding sites for both SPL and AGL transcription factors. The PvSPL9 and PvFUL‐ like genes are highly expressed in roots/nodules from inoculated plants as compared to roots from fertilized plants. Effector/reporter assays in Nicotiana benthamiana leaves transiently transformed with fluorescent reporter constructs confirmed that PvSPL9d and PvFUL‐like activate PvMIR172c expression. In addition, we showed an increased level of miR172c in common bean transgenic roots and nodules overexpressing PvSPL9d or PvFUL ‐like, while the opposite results held in silenced transgenic tissues. These findings provide an additional understanding of the complex regulatory network controlling NFS in common bean plants.