title: A domain-based approach for analyzing the function of aluminum-activated malate transporters from wheat (Triticum aestivum) and Arabidopsis thaliana in Xenopus oocytes.
abstract: Wheat and Arabidopsis plants respond to aluminum (Al) ions by releasing malate from their root apices via Al-activated malate transporter. Malate anions bind with the toxic Al ions and contribute to the Al tolerance of these species. The genes encoding the transporters in wheat and Arabidopsis, TaALMT1 and AtALMT1, respectively, were expressed in Xenopus laevis oocytes and characterized electrophysiologically using the two-electrode voltage clamp system. The Al-activated currents generated by malate efflux were detected for TaALMT1 but not for AtALMT1. Chimeric proteins were generated by swapping the N- and C-terminal halves of TaALMT1 and AtALMT1 (Ta::At and At::Ta). When these chimeras were characterized in oocytes, Al-activated malate efflux was detected for the Ta::At chimera but not for At::Ta, suggesting that the N-terminal half of TaALMT1 is necessary for function in oocytes. An additional chimera, Ta(48)::At, generated by swapping 17 residues from the N-terminus of AtALMT1 with the equivalent 48 residues from TaALMT1, was sufficient to support transport activity. This 48 residue region includes a helical region with a putative transmembrane domain which is absent in AtALMT1. The deletion of this domain from Ta(48)::At led to the complete loss of transport activity. Furthermore, truncations and a deletion at the C-terminal end of TaALMT1 indicated that a putative helical structure in this region was also required for transport function. This study provides insights into the structure-function relationships of Al-activated ALMT proteins by identifying specific domains on the N- and C-termini of TaALMT1 that are critical for basal transport function and Al responsiveness in oocytes.
authors.affiliation: Institute of Plant Science and Resources, Okayama University, Chuo 2-20-1, Kurashiki, Okayama, 710-0046 Japan tsasaki@rib.okayama-u.ac.jp.
authors.forename: Takayuki
authors.initials: T
authors.lastname: Sasaki
authors.affiliation: Institute of Plant Science and Resources, Okayama University, Chuo 2-20-1, Kurashiki, Okayama, 710-0046 Japan.
authors.forename: Yoshiyuki
authors.initials: Y
authors.lastname: Tsuchiya
authors.affiliation: Institute of Plant Science and Resources, Okayama University, Chuo 2-20-1, Kurashiki, Okayama, 710-0046 Japan.
authors.forename: Michiyo
authors.initials: M
authors.lastname: Ariyoshi
authors.affiliation: CSIRO Agriculture, GPO Box 1600, Canberra, ACT 2601, Australia.
authors.forename: Peter R
authors.initials: PR
authors.lastname: Ryan
authors.affiliation: Department of Health and Nutrition, Faculty of Home Economics, Gifu Women's University, Taromaru 80, Gifu, 501-2592 Japan.
authors.forename: Takuya
authors.initials: T
authors.lastname: Furuichi
authors.affiliation: Institute of Plant Science and Resources, Okayama University, Chuo 2-20-1, Kurashiki, Okayama, 710-0046 Japan.
authors.forename: Yoko
authors.initials: Y
authors.lastname: Yamamoto
mesh_terms: D000535:Aluminum / Q000494:pharmacology*; D000595:Amino Acid Sequence; D000818:Animals; D017360:Arabidopsis / Q000187:drug effects / Q000235:genetics / Q000502:physiology*; D029681:Arabidopsis Proteins / Q000235:genetics / Q000378:metabolism*; D001692:Biological Transport; D005260:Female; D008293:Malates / Q000378:metabolism*; D008969:Molecular Sequence Data; D009865:Oocytes; D027361:Organic Anion Transporters / Q000235:genetics / Q000378:metabolism*; D010940:Plant Proteins / Q000235:genetics / Q000378:metabolism; D018517:Plant Roots / Q000235:genetics; D017434:Protein Structure, Tertiary; D011994:Recombinant Proteins; D016415:Sequence Alignment; D017384:Sequence Deletion; D014908:Triticum / Q000187:drug effects / Q000235:genetics / Q000502:physiology*; D014981:Xenopus
chemical_list: C512361:ALMT1 protein, Arabidopsis; D029681:Arabidopsis Proteins; D008293:Malates; D027361:Organic Anion Transporters; D010940:Plant Proteins; D011994:Recombinant Proteins; C030298:malic acid; D000535:Aluminum
keywords: Aluminum-activated malate transporter; Arabidopsis thaliana; Domain structure; Electrophysiology; Wheat (Triticum aestivum); Xenopus laevis oocyte