Glycosylation of the osmoresponsive transient receptor potential channel TRPV4 on Asn-651 influences membrane trafficking.
We identified a consensus N-linked glycosylation motif within the pore-forming loop between the fifth and sixth transmembrane segments of the osmoresponsive transient receptor potential (TRP) channel TRPV4. Mutation of this residue from Asn to Gln (i.e., TRPV4(N651Q)) resulted in loss of a slower migrating band on anti-TRPV4 immunoblots and a marked reduction in lectin-precipitable TRPV4 immunoreactivity. HEK293 cells transiently transfected with the mutant TRPV4(N651Q) exhibited increased calcium entry in response to hypotonic stress relative to wild-type TRPV4 transfectants. This increase in hypotonicity responsiveness was associated with an increase in plasma membrane targeting of TRPV4(N651Q) relative to wild-type TRPV4 in both HEK293 and COS-7 cells but had no effect on overall channel abundance in whole cell lysates. Residue N651 of TRPV4 is immediately adjacent to the pore-forming loop. Although glycosylation in this vicinity has not been reported for a TRP channel, the structurally related hexahelical hyperpolarization-activated cyclic nucleotide-gated channel, HCN2, and the voltage-gated potassium channel, human ether-a-go-go-related (HERG), share a nearly identically situated and experimentally confirmed N-linked glycosylation site which promotes rather than limits channel insertion into the plasma membrane. These data point to a potentially conserved structural and functional feature influencing membrane trafficking across diverse members of the voltage-gated-like ion channel superfamily.
Xu
Hongshi
H
Mailcode PP262, Oregon Health and Science Univ., 3314 S.W. US Veterans Hospital Rd., Portland, OR 97239, USA.
Fu
Yi
Y
Tian
Wei
W
Cohen
David M
DM
D000595:Amino Acid Sequence; D000818:Animals; D001692:Biological Transport / Q000502:physiology*; D019556:COS Cells; D048429:Cell Size; D002645:Chickens; D002522:Chlorocebus aethiops; D004594:Electrophysiology; D006031:Glycosylation; D015151:Immunoblotting; D007473:Ion Channels / Q000502:physiology*; D007668:Kidney / Q000166:cytology; D008564:Membrane Potentials; D051379:Mice; D008969:Molecular Sequence Data; D010802:Phylogeny; D051381:Rats; D017385:Sequence Homology; D013329:Structure-Activity Relationship; D050916:TRPV Cation Channels / Q000235:genetics* / Q000502:physiology*; D014162:Transfection; D014882:Water-Electrolyte Balance
D007473:Ion Channels; D050916:TRPV Cation Channels; C416986:TRPV4 protein, human