Regulation of the Zinc Deficiency Response in the Legume Model Medicago truncatula

Abstract

The zinc deficiency response in Arabidopsis thaliana is regulated by F-group basic region leucine-zipper (F-bZIP) transcription factors, and there is evidence of evolutionary conservation of this regulatory network in land plants. Fundamental knowledge on the zinc homeostasis regulation in crop species will contribute to improving their zinc nutritional value. Legumes are protein-rich crops, used worldwide as part of traditional diets and as animal forage, being therefore a good target for micronutrient biofortification. Here, we identified F-bZIP transcription factors in representative legume species and functionally characterized the two F-bZIPs from Medicago truncatula. Results indicate that MtFbZIP1 is the functional homolog of A. thaliana bZIP19 and bZIP23, while MtFbZIP2 does not play a role in the zinc deficiency response. Additionally, analysis of M. truncatula genes from the Zrt/Irt-like protein (ZIP) family of zinc transporters or encoding nicotianamine synthase enzymes that produce the zinc ligand nicotianamine, support the conservation of the F-bZIP-regulated zinc deficiency response in M. truncatula. Phylogenetic analysis of F-bZIP homologs enriched in legume species reinforces the branching into two groups, with MtFbZIP1 and MtFbZIP2 mapping in Groups 1 and 2, respectively. This phylogeny combined with the functional characterization of MtFbZIPs supports the suggested conservation of the zinc deficiency response associated with Group 1 F-bZIPs, and the more variable evolutionary paths associated with Group 2. Overall, we provide novel insight on the mechanisms of response to zinc deficiency in M. truncatula, which contributes to developing strategies for improving zinc content in legume crops.

The F-bZIP-regulated Zn deficiency response in land plants

“…In addition to the phylogenetic analyses of F-bZIP homologs, analysis of the promoter region of Arabidopsis ZIP4 homologs across land plants identified an enrichment in ZDREs and a conserved substitution in the consensus sequence (A/GTGT/CCGACAC/T) (Castro et al 2017). The analysis of ZIP and NAS genes from barley, wheat, rice and M. truncatula revealed an association between Zn-deficiency induced expression and the presence of ZDREs in the promoter region (Evens et al 2017;Nazri et al 2017;Lilay et al 2020;Liao et al 2022). This further supports the conservation of the F-bZIP-regulated Zn deficiency response in land plants, which seems to comprise a conserved F-bZIP-regulon module.…”

“…The phylogenetic analysis across land plants indicated divergence of F-bZIP homologs during seed plant evolution into two groups (Castro et al 2017), which was also supported by later phylogenetic analyses with enlarged datasets of Monocot and legume species (Lilay et al 2020;Liao et al 2022). These two groups, within F-bZIPs, were named Group 1 and Group 2.…”

“…Thus, the combination of phylogenetic analysis and functional characterization of F-bZIPs supports the conservation of the Zn deficiency response associated with Group 1 members, while the Group 2 F-bZIPs show more variable evolutionary paths that can lead to non-, sub-or neo-functionalization (Castro et al 2017). Nonetheless, all F-bZIP homologs analysed across land plants show a high level conservation of the ZSM (Assunção et al 2013;Castro et al 2017;Lilay et al 2020;Liao et al 2022), suggesting that possible neo-functionalization for Group 2 F-bZIPs might still retain a Zn-sensing function.…”

The F-bZIP-regulated Zn deficiency response in land plants

“…It is thus conceivable that the F-bZIP-regulated Zn deficiency response played a role in the transition to land by equipping plants with a mechanism to cope with Zn deficiency within a range of variation in Zn availability from the soil. The phylogenetic analysis across land plants indicated divergence of F-bZIP homologs during seed plant evolution into two groups (Castro et al 2017), which was also supported by later phylogenetic analyses with enlarged datasets of Monocot and legume species (Lilay et al 2020;Liao et al 2022). These two groups, within F-bZIPs, were named Group 1 and Group 2.…”

“…The phylogenetic analysis across land plants indicated divergence of F-bZIP homologs during seed plant evolution into two groups (Castro et al 2017 ), which was also supported by later phylogenetic analyses with enlarged datasets of Monocot and legume species (Lilay et al 2020 ; Liao et al 2022 ). These two groups, within F-bZIPs, were named Group 1 and Group 2.…”