Identification of conserved and new miRNAs that affect nodulation and strain selectivity in the Phaseolus vulgaris-Rhizobium etli symbiosis through differential analysis of host small RNAs.

Phaseolus vulgaris plants from the Mesoamerican centre of genetic diversification establish a preferential and more efficient root nodule symbiosis with sympatric Rhizobium etli strains. This is mediated by changes in host gene expression, which might occur either at the transcriptional or at the post-transcriptional level. However, the implication of small RNA (sRNA)-mediated control of gene expression in strain selectivity has remained elusive. sRNA sequencing was used to identify host microRNAs (miRNAs) differentially regulated in roots at an early stage of the symbiotic interaction, which were further characterized by applying a reverse genetic approach. In silico analysis identified known and new miRNAs that accumulated to a greater extent in the preferential and more efficient interaction. One of them, designated as Pvu-miR5924, participates in the mechanisms that determine the selection of R. etli strains that will colonize the nodules. In addition, the functional analysis of Pvu-miR390b verified that this miRNA is a negative modulator of nodule formation and bacterial infection. This study not only extended the list of miRNAs identified in P. vulgaris but also enabled the identification of miRNAs that play relevant functions in nodule formation, rhizobial infection and the selection of the rhizobial strains that will occupy the nodule.

The PvNF-YA1 and PvNF-YB7 Subunits of the Heterotrimeric NF-Y Transcription Factor Influence Strain Preference in the Phaseolus vulgaris-Rhizobium etli Symbiosis.

Transcription factors of the Nuclear Factor Y (NF-Y) family play essential functions in plant development and plasticity, including the formation of lateral root organs such as lateral root and symbiotic nodules. NF-Ys mediate transcriptional responses by acting as heterotrimers composed of three subunits, NF-YA, NF-YB, and NF-YC, which in plants are encoded by relatively large gene families. We have previously shown that, in the Phaseolus vulgaris × Rhizobium etli interaction, the PvNF-YC1 subunit is involved not only in the formation of symbiotic nodules, but also in the preference exhibited by the plant for rhizobial strains that are more efficient and competitive in nodule formation. PvNF-YC1 forms a heterotrimer with the PvNF-YA1 and PvNF-YB7 subunits. Here, we used promoter:reporter fusions to show that both PvNF-YA1 and PvNF-YB7 are expressed in symbiotic nodules. In addition, we report that knock-down of PvNF-YA1 and its close paralog PvNF-YA9 abolished nodule formation by either high or low efficient strains and arrested rhizobial infection. On the other hand, knock-down of PvNF-YB7 only affected the symbiotic outcome of the high efficient interaction, suggesting that other symbiotic NF-YB subunits might be involved in the more general mechanisms of nodule formation. More important, we present functional evidence supporting that both PvNF-YA1 and PvNF-YB7 are part of the mechanisms that allow P. vulgaris plants to discriminate and select those bacterial strains that perform better in nodule formation, most likely by acting in the same heterotrimeric complex that PvNF-YC1.

Annotation, phylogeny and expression analysis of the nuclear factor Y gene families in common bean (Phaseolus vulgaris).

In the past decade, plant nuclear factor Y (NF-Y) genes have gained major interest due to their roles in many biological processes in plant development or adaptation to environmental conditions, particularly in the root nodule symbiosis established between legume plants and nitrogen fixing bacteria. NF-Ys are heterotrimeric transcriptional complexes composed of three subunits, NF-YA, NF-YB, and NF-YC, which bind with high affinity and specificity to the CCAAT box, a cis element present in many eukaryotic promoters. In plants, NF-Y subunits consist of gene families with about 10 members each. In this study, we have identified and characterized the NF-Y gene families of common bean (Phaseolus vulgaris), a grain legume of worldwide economical importance and the main source of dietary protein of developing countries. Expression analysis showed that some members of each family are up-regulated at early or late stages of the nitrogen fixing symbiotic interaction with its partner Rhizobium etli. We also showed that some genes are differentially accumulated in response to inoculation with high or less efficient R. etli strains, constituting excellent candidates to participate in the strain-specific response during symbiosis. Genes of the NF-YA family exhibit a highly structured intron-exon organization. Moreover, this family is characterized by the presence of upstream ORFs when introns in the 5' UTR are retained and miRNA target sites in their 3' UTR, suggesting that these genes might be subjected to a complex post-transcriptional regulation. Multiple protein alignments indicated the presence of highly conserved domains in each of the NF-Y families, presumably involved in subunit interactions and DNA binding. The analysis presented here constitutes a starting point to understand the regulation and biological function of individual members of the NF-Y families in different developmental processes in this grain legume.