title: Removal of copper from aqueous solutions by rhizofiltration using genetically modified hairy roots expressing a bacterial Cu-binding protein.
abstract: The aim of this work was to develop a biotechnological tool to hyperaccumulate high copper (Cu) concentrations from wastewaters. Transgenic tobacco hairy roots were obtained by expressing, either the wild-type version of the gene copC from Pseudomonas fluorescens in the cytoplasm of plant cells (CuHR), or a modified version targeted to the vacuole (CuHR-V). Control hairy roots transformed with the empty vector (HR) were also generated. The roots were incubated in the presence of solutions containing Cu (from 1 to 50 mM). At 5 mM external copper, transgenic hairy roots accumulated twice the amount of copper accumulated by control hairy roots. However, at 50 mM Cu, accumulation in both transgenic and control roots reached similar values. Maximum Cu accumulation achieved by transgenic hairy roots was 45,000 µg g-1at 50 mM external Cu. Despite the high Cu accumulation, transgenic hairy roots, particularly CuHR-V, showed less toxicity symptoms, in correlation with lower activity of several antioxidant enzymes and lower malondialdehyde (MDA) levels. Moreover, CuHR-V roots displayed low values of the oxidative stress index (OSI) - a global parameter proposed for oxidative stress - indicating that targeting CopC to the vacuole could alleviate the oxidative stress caused by Cu. Our results suggest that expressing copC in transgenic hairy roots is a suitable strategy to obtain Cu-hyperaccumulator hairy roots with less toxicity stress symptoms.
  
  
  BACKGROUND
  APX: ascorbate peroxidase; ATSDR: Agency for Toxic Substances and Disease Registry (U.S.); BCF: bioconcentration factor; CuHR: copper-hairy roots; EDTA: ethylenediamine tetracetic acid; EPA: Environmental Protection Agency (U.S.); GSH: glutathione; HM: heavy metals; HR: control hairy roots; ICP-OES: Inductively Coupled Plasma/Optical Emission Spectrometry; MDA: malondialdehyde; NBT: nitroblue tetrazolium; OD: optical density; OSI: oxidative stress index; PCR: polymerase chain reaction; PVP: polyvynilpirrolidone; PX: peroxidase; ROS: reactive oxygen species; SOD: superoxide dismutase.
authors.affiliation: a Departamento de Microbiología y Parasitología , Facultad de Farmacia, Universidad de Sevilla , Sevilla , Spain.
authors.forename: Patricia
authors.initials: P
authors.lastname: Pérez-Palacios
authors.affiliation: b Departamento de Biología Molecular , FCEFQyN, Universidad Nacional de Río Cuarto , Córdoba , Argentina.
authors.forename: Elizabeth
authors.initials: E
authors.lastname: Agostini
authors.affiliation: b Departamento de Biología Molecular , FCEFQyN, Universidad Nacional de Río Cuarto , Córdoba , Argentina.
authors.forename: Sabrina G
authors.initials: SG
authors.lastname: Ibáñez
authors.affiliation: b Departamento de Biología Molecular , FCEFQyN, Universidad Nacional de Río Cuarto , Córdoba , Argentina.
authors.forename: Melina A
authors.initials: MA
authors.lastname: Talano
authors.affiliation: a Departamento de Microbiología y Parasitología , Facultad de Farmacia, Universidad de Sevilla , Sevilla , Spain.
authors.forename: Ignacio D
authors.identifier: http://orcid.org/0000-0001-7621-8303
authors.initials: ID
authors.lastname: Rodríguez-Llorente
authors.affiliation: a Departamento de Microbiología y Parasitología , Facultad de Farmacia, Universidad de Sevilla , Sevilla , Spain.
authors.forename: Miguel A
authors.identifier: http://orcid.org/0000-0001-8184-0455
authors.initials: MA
authors.lastname: Caviedes
authors.affiliation: a Departamento de Microbiología y Parasitología , Facultad de Farmacia, Universidad de Sevilla , Sevilla , Spain.
authors.forename: Eloísa
authors.identifier: http://orcid.org/0000-0002-1084-5301
authors.initials: E
authors.lastname: Pajuelo
mesh_terms: D001426:Bacterial Proteins / Q000235:genetics* / Q000378:metabolism; D001673:Biodegradation, Environmental; D003300:Copper / Q000378:metabolism*; D005374:Filtration; D018384:Oxidative Stress; D018517:Plant Roots / Q000378:metabolism / Q000382:microbiology; D030821:Plants, Genetically Modified / Q000235:genetics / Q000378:metabolism; D011551:Pseudomonas fluorescens / Q000235:genetics*; D014026:Nicotiana / Q000235:genetics / Q000378:metabolism*; D014865:Waste Disposal, Fluid / Q000379:methods*; D014874:Water Pollutants, Chemical / Q000378:metabolism
chemical_list: D001426:Bacterial Proteins; C071265:CopC protein, Bacteria; D014874:Water Pollutants, Chemical; D003300:Copper
keywords: Agrobacterium rhizogenes; bio-ore; copC; oxidative stress; tobacco