An extracellular hydrophilic carboxy‐terminal domain regulates the activity of TaALMT1, the aluminum‐activated malate transport protein of wheat |
| |
Authors: | Takuya Furuichi Takayuki Sasaki Yoshiyuki Tsuchiya Peter R Ryan Emmanuel Delhaize Yoko Yamamoto |
| |
Institution: | 1. Institute of Plant Science and Resources, Okayama University, Chuo 2‐20‐1, Kurashiki, Okayama 710‐0046, Japan;2. CSIRO Plant Industry, GPO Box 1600, Canberra, ACT 2601, Australia |
| |
Abstract: | Al3+‐resistant cultivars of wheat (Triticum aestivum L.) release malate through the Al3+‐activated anion transport protein Triticum aestivum aluminum‐activated malate transporter 1 (TaALMT1). Expression of TaALMT1 in Xenopus oocytes and tobacco suspension cells enhances the basal transport activity (inward and outward currents present in the absence of external Al3+), and generates the same Al3+‐activated currents (reflecting the Al3+‐dependent transport function) as observed in wheat cells. We investigated the amino acid residues involved in this Al3+‐dependent transport activity by generating a series of mutations to the TaALMT1 protein. We targeted the acidic residues on the hydrophilic C‐terminal domain of TaALMT1 and changed them to uncharged residues by site‐directed mutagenesis. These mutant proteins were expressed in Xenopus oocytes and their transport activity was measured before and after Al3+ addition. Three mutations (E274Q, D275N and E284Q) abolished the Al3+‐activated transport activity without affecting the basal transport activity. Truncation of the hydrophilic C‐terminal domain abolished both basal and Al3+‐activated transport activities. Al3+‐dependent transport activity was recovered by fusing the N‐terminal region of TaALMT1 with the C‐terminal region of AtALMT1, a homolog from Arabidopsis. These findings demonstrate that the extracellular C‐terminal domain is required for both basal and Al3+‐dependent TaALMT1 activity. Furthermore, we identified three acidic amino acids within this domain that are specifically required for the activation of transport function by external Al3+. |
| |
Keywords: | acidic amino acids aluminum activation malate transport Triticum aestivum aluminum‐activated malate transporter 1 two‐electrode voltage clamp Xenopus oocyte |
|
|