The phosphoinositide PI(3,5)P2 mediates activation of mammalian but not plant TPC proteins: Functional expression of endolysosomal channels in yeast and plant cells

Anna Boccaccio, Joachim Scholz-Starke, Shin Hamamoto, Nina Larisch, Margherita Festa, Paul Vijay Kanth Gutla, Alex Costa, Petra Dietrich, Nobuyuki Uozumi, Armando Carpaneto

Research output: Contribution to journalArticlepeer-review

30 Citations (Scopus)

Abstract

Two-pore channel proteins (TPC) encode intracellular ion channels in both animals and plants. In mammalian cells, the two isoforms (TPC1 and TPC2) localize to the endo-lysosomal compartment, whereas the plant TPC1 protein is targeted to the membrane surrounding the large lytic vacuole. Although it is well established that plant TPC1 channels activate in a voltage- and calcium-dependent manner in vitro, there is still debate on their activation under physiological conditions. Likewise, the mode of animal TPC activation is heavily disputed between two camps favoring as activator either nicotinic acid adenine dinucleotide phosphate (NAADP) or the phosphoinositide PI(3,5)P2. Here, we investigated TPC current responses to either of these second messengers by whole-vacuole patch-clamp experiments on isolated vacuoles of Arabidopsis thaliana. After expression in mesophyll protoplasts from Arabidopsis tpc1 knock-out plants, we detected the Arabidopsis TPC1-EGFP and human TPC2-EGFP fusion proteins at the membrane of the large central vacuole. Bath (cytosolic) application of either NAADP or PI(3,5)P2 did not affect the voltage- and calcium-dependent characteristics of AtTPC1-EGFP. By contrast, PI(3,5)P2 elicited large sodium currents in hTPC2-EGFP-containing vacuoles, while NAADP had no such effect. Analogous results were obtained when PI(3,5)P2 was applied to hTPC2 expressed in baker's yeast giant vacuoles. Our results underscore the fundamental differences in the mode of current activation and ion selectivity between animal and plant TPC proteins and corroborate the PI(3,5)P2-mediated activation and Na+ selectivity of mammalian TPC2.

Original languageEnglish
Pages (from-to)4275-4283
Number of pages9
JournalCellular and Molecular Life Sciences
Volume71
Issue number21
DOIs
Publication statusPublished - 2014 Nov

Keywords

  • Lysosome
  • Patch clamp
  • Plant vacuole
  • TPC channels
  • Yeast vacuole

ASJC Scopus subject areas

  • Molecular Medicine
  • Molecular Biology
  • Pharmacology
  • Cellular and Molecular Neuroscience
  • Cell Biology

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