title: Nitrate efflux at the root plasma membrane: identification of an Arabidopsis excretion transporter.
abstract: Root NO(3)(-) efflux to the outer medium is a component of NO(3)(-) net uptake and can even overcome influx upon various stresses. Its role and molecular basis are unknown. Following a functional biochemical approach, NAXT1 (for NITRATE EXCRETION TRANSPORTER1) was identified by mass spectrometry in the plasma membrane (PM) of Arabidopsis thaliana suspension cells, a localization confirmed using a NAXT1-Green Fluorescent Protein fusion protein. NAXT1 belongs to a subclass of seven NAXT members from the large NITRATE TRANSPORTER1/PEPTIDE TRANSPORTER family and is mainly expressed in the cortex of mature roots. The passive NO(3)(-) transport activity (K(m) = 5 mM) in isolated root PM, electrically coupled to the ATP-dependant H(+)-pumping activity, is inhibited by anti-NAXT antibodies. In standard culture conditions, NO(3)(-) contents were altered in plants expressing NAXT-interfering RNAs but not in naxt1 mutant plants. Upon acid load, unidirectional root NO(3)(-) efflux markedly increased in wild-type plants, leading to a prolonged NO(3)(-) excretion regime concomitant with a decrease in root NO(3)(-) content. In vivo and in vitro mutant phenotypes revealed that this response is mediated by NAXT1, whose expression is upregulated at the posttranscriptional level. Strong medium acidification generated a similar response. In vitro, the passive efflux of NO(3)(-) (but not of Cl(-)) was strongly impaired in naxt1 mutant PM. This identification of NO(3)(-) efflux transporters at the PM of plant cells opens the way to molecular studies of the physiological role of NO(3)(-) efflux in stressed or unstressed plants.
authors.affiliation: Biochimie et Physiologie Moléculaire des Plantes, Agro-M/Centre National de la Recherche Scientifique/Institut National de la Recherche Agronomique/Université Montpellier 2, France.
authors.forename: Cécile
authors.initials: C
authors.lastname: Segonzac
authors.forename: Jean-Christophe
authors.initials: JC
authors.lastname: Boyer
authors.forename: Emilie
authors.initials: E
authors.lastname: Ipotesi
authors.forename: Wojciech
authors.initials: W
authors.lastname: Szponarski
authors.forename: Pascal
authors.initials: P
authors.lastname: Tillard
authors.forename: Brigitte
authors.initials: B
authors.lastname: Touraine
authors.forename: Nicolas
authors.initials: N
authors.lastname: Sommerer
authors.forename: Michel
authors.initials: M
authors.lastname: Rossignol
authors.forename: Rémy
authors.initials: R
authors.lastname: Gibrat
mesh_terms: D000595:Amino Acid Sequence; D027321:Anion Transport Proteins / Q000378:metabolism; D000906:Antibodies; D017360:Arabidopsis / Q000166:cytology* / Q000201:enzymology / Q000235:genetics / Q000378:metabolism*; D029681:Arabidopsis Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D001692:Biological Transport; D002462:Cell Membrane / Q000201:enzymology / Q000378:metabolism*; D003470:Culture Media; D004269:DNA, Bacterial / Q000378:metabolism; D018506:Gene Expression Regulation, Plant; D049452:Green Fluorescent Proteins / Q000378:metabolism; D008969:Molecular Sequence Data; D016254:Mutagenesis, Insertional; D009566:Nitrates / Q000378:metabolism*; D010641:Phenotype; D010940:Plant Proteins / Q000378:metabolism; D018517:Plant Roots / Q000166:cytology* / Q000201:enzymology / Q000378:metabolism; D018520:Plant Shoots / Q000378:metabolism; D006180:Proton-Translocating ATPases / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000378:metabolism
chemical_list: D027321:Anion Transport Proteins; D000906:Antibodies; D029681:Arabidopsis Proteins; D003470:Culture Media; D004269:DNA, Bacterial; C101492:NRT1.1 protein, Arabidopsis; D009566:Nitrates; D010940:Plant Proteins; D011993:Recombinant Fusion Proteins; C036483:T-DNA; D049452:Green Fluorescent Proteins; D006180:Proton-Translocating ATPases