A TRPV4 channel C-terminal folding recognition domain critical for trafficking and function

Lei Lei, Xu Cao, Fan Yang, Di Jing Shi, Yi Quan Tang, Jie Zheng, Kewei Wang

Research output: Contribution to journalArticle

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Abstract

The Ca2 -permeable transient receptor potential vanilloid subtype 4 (TRPV4) channel mediates crucial physiological functions, such as calcium signaling, temperature sensing, and maintaining cell volume and energy homeostasis. Noticeably, most disease-causing genetic mutations are concentrated in the cytoplasmic domains. In the present study, we focused on the role of the TRPV4 C terminus in modulating protein folding, trafficking, and activity. By examining a series of C-terminal deletions, we identified a 20-amino acid distal region covering residues 838-857 that is critical for channel folding, maturation, and trafficking. Surface biotinylation, confocal imaging, and fluorescence- based calcium influx assay demonstrated that mutant proteins missing this region were trapped in the endoplasmic reticulum and unglycosylated, leading to accelerated degradation and loss of channel activity. Rosetta de novo structural modeling indicated that residues 838-857 assume a defined conformation, with Gly849 and Pro851 located at critical positions. Patch clamp recordings confirmed that lowering the temperature from 37 to 30 °C rescued channel activity of folding-defective mutants. Moreover, biochemical tests demonstrated that, in addition to participating in C-C interaction, the C terminus also interacts with the N terminus. Taken together, our findings indicate that the C-terminal region of TRPV4 is critical for channel protein folding and maturation, and the short distal segment plays an essential role in this process. Therefore, selectively disrupting the folding-sensitive region may present therapeutic potential for treating overactive TRPV4-mediated diseases, such as pain and skeletal dysplasias.

Original languageEnglish (US)
Pages (from-to)10427-10439
Number of pages13
JournalJournal of Biological Chemistry
Volume288
Issue number15
DOIs
StatePublished - Apr 12 2013

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TRPV Cation Channels
Protein folding
Protein Folding
Biotinylation
Calcium
Inborn Genetic Diseases
Temperature
Calcium Signaling
Optical Imaging
Clamping devices
Protein Transport
Mutant Proteins
Cell Size
Endoplasmic Reticulum
Conformations
Assays
Homeostasis
Fluorescence
Imaging techniques
Amino Acids

ASJC Scopus subject areas

  • Biochemistry
  • Cell Biology
  • Molecular Biology

Cite this

A TRPV4 channel C-terminal folding recognition domain critical for trafficking and function. / Lei, Lei; Cao, Xu; Yang, Fan; Shi, Di Jing; Tang, Yi Quan; Zheng, Jie; Wang, Kewei.

In: Journal of Biological Chemistry, Vol. 288, No. 15, 12.04.2013, p. 10427-10439.

Research output: Contribution to journalArticle

Lei, Lei ; Cao, Xu ; Yang, Fan ; Shi, Di Jing ; Tang, Yi Quan ; Zheng, Jie ; Wang, Kewei. / A TRPV4 channel C-terminal folding recognition domain critical for trafficking and function. In: Journal of Biological Chemistry. 2013 ; Vol. 288, No. 15. pp. 10427-10439.
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