title: Fast fluorometric method for measuring pendrin (SLC26A4) Cl-/I- transport activity.
abstract: Malfunction of the SLC26A4 protein leads to Pendred syndrome, characterized by sensorineural hearing loss, often associated with mild thyroid dysfunction and goiter. It is generally assumed that SLC26A4 acts as a chloride/anion exchanger, which in the thyroid gland transports iodide, and in the inner ear contributes to the conditioning of the endolymphatic fluid. Here we describe a fast fluorometric method able to be used to functionally scrutinize SLC26A4 and its mutants described in Pendred syndrome. The validation of the method was done by functionally characterizing the chloride/iodide transport of SLC26A4, and a mutant, i.e. SLC26A4(S28R), which we previously described in a patient with sensorineural hearing loss, hypothyroidism and goiter. Using the fluorometric method we describe here we can continuously monitor and quantify the iodide or chloride amounts transported by the cells, and we found that the transport capability of the SLC26A4(S28R) mutant protein is markedly reduced if compared to wild-type SLC26A4.
authors.affiliation: Department of Biomolecular Sciences and Biotechnology, Universitá degli Studi di Milano.
authors.forename: Silvia
authors.initials: S
authors.lastname: Dossena
authors.forename: Simona
authors.initials: S
authors.lastname: Rodighiero
authors.forename: Valeria
authors.initials: V
authors.lastname: Vezzoli
authors.forename: Claudia
authors.initials: C
authors.lastname: Bazzini
authors.forename: Chiara
authors.initials: C
authors.lastname: Sironi
authors.forename: Giuliano
authors.initials: G
authors.lastname: Meyer
authors.forename: Johannes
authors.initials: J
authors.lastname: Fürst
authors.forename: Markus
authors.initials: M
authors.lastname: Ritter
authors.forename: Maria L
authors.initials: ML
authors.lastname: Garavaglia
authors.forename: Laura
authors.initials: L
authors.lastname: Fugazzola
authors.forename: Luca
authors.initials: L
authors.lastname: Persani
authors.forename: Patrick
authors.initials: P
authors.lastname: Zorowka
authors.forename: Carlo
authors.initials: C
authors.lastname: Storelli
authors.forename: Paolo
authors.initials: P
authors.lastname: Beck-Peccoz
authors.forename: Guido
authors.initials: G
authors.lastname: Bottá
authors.forename: Markus
authors.initials: M
authors.lastname: Paulmichl
mesh_terms: D001692:Biological Transport / Q000187:drug effects; D002460:Cell Line; D002712:Chlorides / Q000378:metabolism*; D005452:Fluoresceins / Q000378:metabolism; D005470:Fluorometry / Q000379:methods*; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007454:Iodides / Q000378:metabolism*; D026901:Membrane Transport Proteins / Q000235:genetics / Q000378:metabolism*; D016297:Mutagenesis, Site-Directed / Q000379:methods; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D000076988:Sulfate Transporters; D014162:Transfection
chemical_list: D002712:Chlorides; D005452:Fluoresceins; D007454:Iodides; D026901:Membrane Transport Proteins; D011993:Recombinant Fusion Proteins; C085609:SLC26A4 protein, human; D000076988:Sulfate Transporters; D049452:Green Fluorescent Proteins; C043829:2',7'-bis(carboxyethyl)-5(6)-carboxyfluorescein