The involvement of nuclear factor Y (NF-Y) in transcriptional reprogramming during arbuscular mycorrhizal symbiosis has been demonstrated in several plant species. However, a comprehensive picture is lacking. We showed that the spatial expression of NF-YC3 was observed in cortical cells containing arbuscules via the cis-regulatory element GCC boxes. Moreover, the NF-YC3 promoter was transactivated by the combination of CYCLOPS and autoactive calcium and calmodulin-dependent kinase (CCaMK) via GCC boxes. Knockdown of NF-YC3 significantly reduced the abundance of all intraradical fungal structures and affected arbuscule size. BCP1, SbtM1, and WRI5a, whose expression associated with NF-YC3 levels, might be downstream of NF-YC3. NF-YC3 interacted with NF-YB3a, NF-YB5c, or NF-YB3b, in yeast (Saccharomyces cerevisiae) and in planta, and interacted with NF-YA3a in yeast. Spatial expression of 3 NF-YBs was observed in all cell layers of roots under both mock and mycorrhizal conditions. Simultaneous knockdown of 3 NF-YBs, but not individually, reduced the fungal colonization level, suggesting that there might be functional redundancy of NF-YBs to regulate AM symbiosis. Collectively, our data suggest that NF-YC3 and NF-YBs positively regulate AM symbiosis in tomato, and arbuscule-related NF-YC3 may be an important downstream gene of the common symbiosis signaling pathway.
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