title: Activity of tonoplast proton pumps and Na+/H+ exchange in potato cell cultures is modulated by salt.
abstract: The efficient exclusion of excess Na from the cytoplasm and vacuolar Na(+) accumulation are the main mechanisms for the adaptation of plants to salt stress. This is typically carried out by transmembrane transport proteins that exclude Na(+) from the cytosol in exchange for H(+), a secondary transport process which is energy-dependent and driven by the proton-motive force generated by plasma-membrane and tonoplast proton pumps. Tonoplast enriched-vesicles from control and 150 mM NaCl-tolerant calli lines were used as a model system to study the activity of V-H(+)-PPase and V-H(+)-ATPase and the involvement of Na(+) compartmentalization into the vacuole as a mechanism of salt tolerance in Solanum tuberosum. Both ATP- and pyrophosphate (PP(i))-dependent H(+)-transport were higher in tonoplast vesicles from the salt-tolerant line than in vesicles from control cells. Western blotting of tonoplast proteins confirmed that changes in V-H(+)-PPase activity are correlated with increased protein amount. Conversely, immunodetection of the A-subunit of V-H(+)-ATPase revealed that a mechanism of post-translational regulation is probably involved. Na(+)-dependent dissipation of a pre-established pH gradient was used to measure Na(+)/H(+) exchange in tonoplast vesicles. The initial rates of proton efflux followed Michaelis-Menten kinetics and the V(max) of proton dissipation was 2-fold higher in NaCl-tolerant calli when compared to the control. H(+)-coupled exchange was specific for Na(+) and Li(+) and not for K(+). The increase of both the pH gradient across the tonoplast and the Na(+)/H(+) antiport activity in response to salt strongly suggests that Na(+) sequestration into the vacuole contributes to salt tolerance in potato.
authors.affiliation: Departamento de Botânica, Faculdade de Ciências, Universidade do Porto, Ed. FC4, Rua do Campo Alegre, s/n masculine, 4169-007 Porto, Portugal.
authors.forename: Filipa
authors.initials: F
authors.lastname: Queirós
authors.forename: Natacha
authors.initials: N
authors.lastname: Fontes
authors.forename: Paulo
authors.initials: P
authors.lastname: Silva
authors.forename: Domingos
authors.initials: D
authors.lastname: Almeida
authors.forename: Masayoshi
authors.initials: M
authors.lastname: Maeshima
authors.forename: Hernâni
authors.initials: H
authors.lastname: Gerós
authors.forename: Fernanda
authors.initials: F
authors.lastname: Fidalgo
mesh_terms: D015153:Blotting, Western; D050260:Carbohydrate Metabolism / Q000187:drug effects; D018929:Cell Culture Techniques; D007477:Ions; D010455:Peptides / Q000378:metabolism; D017494:Proton Pumps / Q000378:metabolism*; D012965:Sodium Chloride / Q000494:pharmacology*; D017923:Sodium-Hydrogen Exchangers / Q000378:metabolism*; D011198:Solanum tuberosum / Q000166:cytology* / Q000201:enzymology / Q000254:growth & development / Q000378:metabolism*; D025262:Vacuolar Proton-Translocating ATPases / Q000378:metabolism; D014617:Vacuoles / Q000187:drug effects* / Q000201:enzymology / Q000378:metabolism*
chemical_list: D007477:Ions; D010455:Peptides; D017494:Proton Pumps; D017923:Sodium-Hydrogen Exchangers; D012965:Sodium Chloride; D025262:Vacuolar Proton-Translocating ATPases