Orientation of Arabidopsis thaliana KAT1 channel in the plasma membrane

• Published in: The Journal of membrane biology Vol. 201; no. 3; pp. 157 - 165
• Main Authors: Mura, C V, Cosmelli, D, Muñoz, F, Delgado, R
• Format: Journal Article
• Language: English
• Published: United States Springer Nature B.V 01.11.2004
• Subjects: Animals; Arabidopsis - enzymology; Arabidopsis - genetics; Arabidopsis Proteins; Cell Membrane - chemistry; Cell Membrane - enzymology; Cell Membrane - genetics; COS Cells; Cricetinae; Oocytes; Plant Proteins - chemistry; Plant Proteins - genetics; Plant Proteins - metabolism; Potassium Channels - metabolism; Potassium Channels, Inwardly Rectifying - genetics; Potassium Channels, Inwardly Rectifying - metabolism; Recombinant Proteins - genetics; Recombinant Proteins - metabolism; Shaker Superfamily of Potassium Channels; Xenopus
• ISSN: 0022-2631; 1432-1424
• DOI: 10.1007/s00232-004-0713-8

The Arabidopsis thaliana KAT1, an inward-rectifying potassium channel, shares molecular features with the Shaker family of outward rectifier K(+) channels. The KAT1 amino-acid sequence reveals the presence of a positively charged S4 and a segment containing the TXGYGD signature sequence in the pore (P) region. To test whether the inward-rectifying properties of KAT1 are due to reverse orientation in the membrane, such that the voltage sensor is oriented in the opposite direction of the electric field compared with the Shaker K(+) channel, we have inserted a flag epitope in the NH(2) terminus or the S3-S4 loop. The KAT1 and tagged constructs expressed functional channels in whole cells, Xenopus oocytes and COS-7. The electrophysiological properties of both tagged constructs were similar to those of the wild type. Immunofluorescence with an antibody against the flag epitope and an anti-C terminal KAT1 determined the membrane localization of these epitopes and the orientation of the KAT1 channel in the membrane. Our data confirm that KAT1 in eukaryotic cells has an orientation similar to the Shaker K(+) channel.