title: Physiological and Molecular Responses to Excess Boron in Citrus macrophylla W.
abstract: This work provides insight into several mechanisms involved in boron (B) regulation pathway in response to high B conditions in Citrus. The study was carried out in Citrus macrophylla W. (Cm) seedlings cultured "in vitro" in media with 50 or 400 μM H3BO3 (control, Ct, and B-excess, +B, plants, respectively). Growth parameters, B concentration, leaf chlorophyll (Chl) concentration, the expression of the main putative genes involved in B transport and distribution, and leaf and root proline and malonaldehyde (MDA) concentrations, were assessed. Excess B led to high B concentration in +B plants (3.8- and 1.4-fold in leaves and roots, respectively) when compared with Ct ones. However, a minor effect was recorded in the plant (incipient visual symptoms, less than 27% reduction in root growth and 26% decrease in Chl b concentration). B toxicity down-regulated by half the expression level of putative B transporter genes NIP5 and PIP1. CmBOR1 gene was not repressed in +B plants and B accumulated in the shoots. High B level increased the transcripts of putative gene TIP5, involved in B transport across the tonoplast, by 3.3- and 2.4-fold in leaves and roots, respectively. The activity of V-PPiase proton pump, related with the electrochemical gradient in the vacuole, was also enhanced in +B organs. B toxicity up-regulated putative BOR4 gene (2.1- and 2.7-fold in roots and leaves, respectively), which codifies for an active efflux B transporter. Accordingly, B was located in +B plants preferently in an insoluble form on cell walls. Finally, excess B caused a significant rise in proline concentration (51% and 34% in roots and leaves, respectively), while the MDA level did not exceed 20%. In conclusion, Cm tolerance to a high B level is likely based on the synergism of several specific mechanisms against B toxicity, including: 1/ down-regulation of NIP5 and PIP1 boron transporters; 2/ activation of B efflux from cells due to the up-regulation of putative BOR4 gene; 3/ compartmentation of B in the vacuole through TIP5 transporter activation and the acidification of the organelle; 4/ insolubilisation of B and deposition in cell walls preventing from cytoplasm damage; and, 5/ induction of an efficient antioxidant system through proline accumulation.
authors.affiliation: Department of Citriculture and Vegetal Production, Valencian Institute of Agrarian Research, Moncada, Valencia, Spain.
authors.forename: Mary-Rus
authors.initials: MR
authors.lastname: Martínez-Cuenca
authors.affiliation: Department of Citriculture and Vegetal Production, Valencian Institute of Agrarian Research, Moncada, Valencia, Spain.
authors.forename: Belén
authors.initials: B
authors.lastname: Martínez-Alcántara
authors.affiliation: Department of Citriculture and Vegetal Production, Valencian Institute of Agrarian Research, Moncada, Valencia, Spain.
authors.forename: Ana
authors.initials: A
authors.lastname: Quiñones
authors.affiliation: Department of Vegetal Protection and Biotechnology, Valencian Institute of Agrarian Research, Moncada, Valencia, Spain.
authors.forename: Marta
authors.initials: M
authors.lastname: Ruiz
authors.affiliation: Department of Citriculture and Vegetal Production, Valencian Institute of Agrarian Research, Moncada, Valencia, Spain.
authors.forename: Domingo J
authors.initials: DJ
authors.lastname: Iglesias
authors.affiliation: Department of Citriculture and Vegetal Production, Valencian Institute of Agrarian Research, Moncada, Valencia, Spain.
authors.forename: Eduardo
authors.initials: E
authors.lastname: Primo-Millo
authors.affiliation: Department of Citriculture and Vegetal Production, Valencian Institute of Agrarian Research, Moncada, Valencia, Spain.
authors.forename: M Ángeles
authors.initials: MÁ
authors.lastname: Forner-Giner
mesh_terms: D001895:Boron / Q000378:metabolism*; D002734:Chlorophyll / Q000378:metabolism; D002957:Citrus / Q000235:genetics / Q000254:growth & development / Q000378:metabolism / Q000502:physiology*; D017343:Genes, Plant; D008315:Malondialdehyde / Q000378:metabolism; D018515:Plant Leaves / Q000378:metabolism; D018517:Plant Roots / Q000378:metabolism
chemical_list: D002734:Chlorophyll; D008315:Malondialdehyde; D001895:Boron