Distinct amino acids in the C-linker domain of the Arabidopsis K+ channel KAT2 determine its subcellular localization and activity at the plasma membrane

Manuel Nieves-Cordones, Alain Chavanieu, Linda Jeanguenin, Carine Alcon, Wojciech Szponarski, Sebastien Estaran, Isabelle Chérel, Sabine Zimmermann, Hervé Sentenac, Isabelle Gaillard

Research output: Contribution to journalArticlepeer-review

19 Scopus citations

Abstract

ShakerK+ channels form the majorK+ conductance of the plasma membrane in plants. They are composed of four subunits arranged around a central ion-conducting pore. The intracellular carboxy-terminal region of each subunit contains several regulatory elements, including a C-linker region and a cyclic nucleotide-binding domain (CNBD). The C-linker is the first domain present downstream of the sixth transmembrane segment and connects the CNBD to the transmembrane core. With the aim of identifying the role of the C-linker in the Shaker channel properties, we performed subdomain swapping between the C-linker of two Arabidopsis (Arabidopsis thaliana) Shaker subunits, K+ channel in Arabidopsis thaliana2 (KAT2) and Arabidopsis thaliana K+ rectifying channel1 (AtKC1). These two subunits contribute to K+ transport in planta by forming heteromeric channels with other Shaker subunits. However, they display contrasting behavior when expressed in tobacco mesophyll protoplasts: KAT2 forms homotetrameric channels active at the plasma membrane, whereas AtKC1 is retained in the endoplasmic reticulum when expressed alone. The resulting chimeric/mutated constructs were analyzed for subcellular localization and functionally characterized. We identified two contiguous amino acids, valine-381 and serine-382, located in the C-linker carboxy-terminal end, which preventKAT2 surface expressionwhenmutated into the equivalent residues fromAtKC1.Moreover,we demonstrated that the nine-amino acid stretch 312TVRAASEFA320 that composes the first C-linker a-helix located just below the pore is a crucial determinant of KAT2 channel activity. A KAT2 C-linker/CNBD three-dimensional model, based on animal HCN (for Hyperpolarization-activated, cyclic nucleotide-gated K+) channels as structure templates, has been built and used to discuss the role of the C-linker in plant Shaker inward channel structure and function.

Original languageEnglish (US)
Pages (from-to)1415-1429
Number of pages15
JournalPlant physiology
Volume164
Issue number3
DOIs
StatePublished - Mar 1 2014

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