Heterologously expressed fungal transient receptor potential channels retain mechanosensitivity in vitro and osmotic response in vivo.
The budding yeast Saccharomyces cerevisiae has a mechanosensitive channel, TrpY1, a member of the Trp superfamily of channels associated with various sensations. Upon a hyperosmotic shift, a yeast cell releases Ca(2+) from the vacuole to the cytoplasm through this channel. The TRPY1 gene has orthologs in other fungal genomes, including TRPY2 of Kluyveromyces lactis and TRPY3 of Candida albicans. We subcloned TRPY2 and TRPY3 and expressed them in the vacuole of S. cerevisiae deleted of TRPY1. The osmotically induced Ca(2+) transient was restored in vivo as reported by transgenic aequorin. Patch-clamp examination showed that the TrpY2 or the TrpY3 channel was similar to TrpY1 in unitary conductance, rectification properties, Ca(2+) sensitivity, and mechanosensitivity. The retention of mechanosensitivity of transient receptor potential channels in a foreign setting, shown here both in vitro and in vivo, implies that these mechanosensitive channels, like voltage-gated or ligand-gated channels, do not discriminate their settings. We discuss various mechanisms, including the possibility that stress from the lipid bilayer by osmotic force transmits forces to the transmembrane domains of these channels.
Zhou
Xin-Liang
XL
Laboratory of Molecular Biology, University of Wisconsin-Madison, Madison, WI 53706, USA.
Loukin
Steven H
SH
Coria
Roberto
R
Kung
Ching
C
Saimi
Yoshiro
Y
D000595:Amino Acid Sequence; D002118:Calcium / Q000378:metabolism; D002176:Candida albicans / Q000378:metabolism; D003001:Cloning, Molecular; D003470:Culture Media / Q000378:metabolism; D003593:Cytoplasm / Q000378:metabolism; D004594:Electrophysiology; D029968:Escherichia coli Proteins / Q000378:metabolism; D005656:Fungal Proteins / Q000378:metabolism; D017930:Genes, Reporter; D016681:Genome, Fungal; D015640:Ion Channel Gating; D007473:Ion Channels / Q000737:chemistry / Q000235:genetics / Q000378:metabolism / Q000502:physiology*; D007700:Kinetics; D007716:Kluyveromyces / Q000378:metabolism; D008024:Ligands; D008051:Lipid Bilayers; D008055:Lipids / Q000737:chemistry; D008969:Molecular Sequence Data; D009995:Osmosis; D018408:Patch-Clamp Techniques; D010802:Phylogeny; D010957:Plasmids / Q000378:metabolism; D016133:Polymerase Chain Reaction; D011312:Pressure; D017434:Protein Structure, Tertiary; D012441:Saccharomyces cerevisiae / Q000378:metabolism*; D013997:Time Factors; D019076:Transgenes
D003470:Culture Media; D029968:Escherichia coli Proteins; D005656:Fungal Proteins; D007473:Ion Channels; D008024:Ligands; D008051:Lipid Bilayers; D008055:Lipids; C469542:MscS protein, E coli; D002118:Calcium