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Plant Biol (Stuttg). 2016 Sep;18(5):750-60. doi: 10.1111/plb.12478. Epub 2016 Jul 12.

Gating of the two-pore cation channel AtTPC1 in the plant vacuole is based on a single voltage-sensing domain.

Author information

1
Institute for Molecular Plant Physiology and Biophysics, University of Würzburg, Würzburg, Germany.
2
Center for Computational and Theoretical Biology, Campus Hubland Nord, Department of Bioinformatics, Biocenter, University Würzburg, Würzburg, Germany.
3
Centro de Bioinformática y Simulación Molecular (CBSM), Universidad de Talca, Talca, Chile.
4
Department of Plant Biology, Ecology, and Evolution, Oklahoma State University, Stillwater, OK, USA.

Abstract

The two-pore cation channel TPC1 operates as a dimeric channel in animal and plant endomembranes. Each subunit consists of two homologous Shaker-like halves, with 12 transmembrane domains in total (S1-S6, S7-S12). In plants, TPC1 channels reside in the vacuolar membrane, and upon voltage stimulation, give rise to the well-known slow-activating SV currents. Here, we combined bioinformatics, structure modelling, site-directed mutagenesis, and in planta patch clamp studies to elucidate the molecular mechanisms of voltage-dependent channel gating in TPC1 in its native plant background. Structure-function analysis of the Arabidopsis TPC1 channel in planta confirmed that helix S10 operates as the major voltage-sensing site, with Glu450 and Glu478 identified as possible ion-pair partners for voltage-sensing Arg537. The contribution of helix S4 to voltage sensing was found to be negligible. Several conserved negative residues on the luminal site contribute to calcium binding, stabilizing the closed channel. During evolution of plant TPC1s from two separate Shaker-like domains, the voltage-sensing function in the N-terminal Shaker-unit (S1-S4) vanished.

KEYWORDS:

Arabidopsis thaliana; gating; plant TPC1; two-pore ion channel; vacuole; voltage sensor

PMID:
27270880
DOI:
10.1111/plb.12478
[Indexed for MEDLINE]

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