Identification of a tetrameric assembly domain in the C terminus of heat-activated TRPV1 channels

Feng Zhang, Shuang Liu, Fan Yang, Jie Zheng, Kewei Wang

Research output: Contribution to journalArticle

24 Citations (Scopus)

Abstract

Transient receptor potential (TRP) channels as cellular sensors are thought to function as tetramers. Yet, the molecular determinants governing channel multimerization remain largely elusive. Here we report the identification of a segment comprising21amino acids(residues752-772 of mouse TRPV1) after the known TRP-like domaininthe channel Cterminus that functions as a tetrameric assembly domain (TAD). Purified recombinant C-terminal proteins of TRPV1-4, but not the N terminus, mediated the protein-protein interaction in an in vitro pulldown assay. Western blot analysis combined with electrophysiology and calcium imaging demonstrated that TAD exerted a robust dominant-negative effect on wild-type TRPV1. When fused with the membrane-tethered peptide Gap43, the TAD blocked the formation of stable homomultimers. Calcium imaging and current recordings showed that deletion of the TAD in a poreless TRPV1 mutant subunit suppressed its dominant-negative phenotype, confirming the involvement of the TAD in assembly of functional channels. Our findings suggest that the C-terminal TAD in TRPV1 channels functions as a domain that is conserved among TRPV1-4 and mediates a direct subunit-subunit interaction for tetrameric assembly.

Original languageEnglish (US)
Pages (from-to)15308-15316
Number of pages9
JournalJournal of Biological Chemistry
Volume286
Issue number17
DOIs
StatePublished - Apr 29 2011

Fingerprint

Hot Temperature
Calcium
Transient Receptor Potential Channels
Electrophysiology
Protein C
Proteins
Western Blotting
Phenotype
Peptides
Acids
Membranes
Imaging techniques
Assays
In Vitro Techniques
Sensors

ASJC Scopus subject areas

  • Biochemistry
  • Cell Biology
  • Molecular Biology

Cite this

Identification of a tetrameric assembly domain in the C terminus of heat-activated TRPV1 channels. / Zhang, Feng; Liu, Shuang; Yang, Fan; Zheng, Jie; Wang, Kewei.

In: Journal of Biological Chemistry, Vol. 286, No. 17, 29.04.2011, p. 15308-15316.

Research output: Contribution to journalArticle

Zhang, Feng ; Liu, Shuang ; Yang, Fan ; Zheng, Jie ; Wang, Kewei. / Identification of a tetrameric assembly domain in the C terminus of heat-activated TRPV1 channels. In: Journal of Biological Chemistry. 2011 ; Vol. 286, No. 17. pp. 15308-15316.
@article{292408897f624b09aaeaff862b012131,
title = "Identification of a tetrameric assembly domain in the C terminus of heat-activated TRPV1 channels",
abstract = "Transient receptor potential (TRP) channels as cellular sensors are thought to function as tetramers. Yet, the molecular determinants governing channel multimerization remain largely elusive. Here we report the identification of a segment comprising21amino acids(residues752-772 of mouse TRPV1) after the known TRP-like domaininthe channel Cterminus that functions as a tetrameric assembly domain (TAD). Purified recombinant C-terminal proteins of TRPV1-4, but not the N terminus, mediated the protein-protein interaction in an in vitro pulldown assay. Western blot analysis combined with electrophysiology and calcium imaging demonstrated that TAD exerted a robust dominant-negative effect on wild-type TRPV1. When fused with the membrane-tethered peptide Gap43, the TAD blocked the formation of stable homomultimers. Calcium imaging and current recordings showed that deletion of the TAD in a poreless TRPV1 mutant subunit suppressed its dominant-negative phenotype, confirming the involvement of the TAD in assembly of functional channels. Our findings suggest that the C-terminal TAD in TRPV1 channels functions as a domain that is conserved among TRPV1-4 and mediates a direct subunit-subunit interaction for tetrameric assembly.",
author = "Feng Zhang and Shuang Liu and Fan Yang and Jie Zheng and Kewei Wang",
year = "2011",
month = "4",
day = "29",
doi = "10.1074/jbc.M111.223941",
language = "English (US)",
volume = "286",
pages = "15308--15316",
journal = "Journal of Biological Chemistry",
issn = "0021-9258",
publisher = "American Society for Biochemistry and Molecular Biology Inc.",
number = "17",

}

TY - JOUR

T1 - Identification of a tetrameric assembly domain in the C terminus of heat-activated TRPV1 channels

AU - Zhang, Feng

AU - Liu, Shuang

AU - Yang, Fan

AU - Zheng, Jie

AU - Wang, Kewei

PY - 2011/4/29

Y1 - 2011/4/29

N2 - Transient receptor potential (TRP) channels as cellular sensors are thought to function as tetramers. Yet, the molecular determinants governing channel multimerization remain largely elusive. Here we report the identification of a segment comprising21amino acids(residues752-772 of mouse TRPV1) after the known TRP-like domaininthe channel Cterminus that functions as a tetrameric assembly domain (TAD). Purified recombinant C-terminal proteins of TRPV1-4, but not the N terminus, mediated the protein-protein interaction in an in vitro pulldown assay. Western blot analysis combined with electrophysiology and calcium imaging demonstrated that TAD exerted a robust dominant-negative effect on wild-type TRPV1. When fused with the membrane-tethered peptide Gap43, the TAD blocked the formation of stable homomultimers. Calcium imaging and current recordings showed that deletion of the TAD in a poreless TRPV1 mutant subunit suppressed its dominant-negative phenotype, confirming the involvement of the TAD in assembly of functional channels. Our findings suggest that the C-terminal TAD in TRPV1 channels functions as a domain that is conserved among TRPV1-4 and mediates a direct subunit-subunit interaction for tetrameric assembly.

AB - Transient receptor potential (TRP) channels as cellular sensors are thought to function as tetramers. Yet, the molecular determinants governing channel multimerization remain largely elusive. Here we report the identification of a segment comprising21amino acids(residues752-772 of mouse TRPV1) after the known TRP-like domaininthe channel Cterminus that functions as a tetrameric assembly domain (TAD). Purified recombinant C-terminal proteins of TRPV1-4, but not the N terminus, mediated the protein-protein interaction in an in vitro pulldown assay. Western blot analysis combined with electrophysiology and calcium imaging demonstrated that TAD exerted a robust dominant-negative effect on wild-type TRPV1. When fused with the membrane-tethered peptide Gap43, the TAD blocked the formation of stable homomultimers. Calcium imaging and current recordings showed that deletion of the TAD in a poreless TRPV1 mutant subunit suppressed its dominant-negative phenotype, confirming the involvement of the TAD in assembly of functional channels. Our findings suggest that the C-terminal TAD in TRPV1 channels functions as a domain that is conserved among TRPV1-4 and mediates a direct subunit-subunit interaction for tetrameric assembly.

UR - http://www.scopus.com/inward/record.url?scp=79955439687&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=79955439687&partnerID=8YFLogxK

U2 - 10.1074/jbc.M111.223941

DO - 10.1074/jbc.M111.223941

M3 - Article

C2 - 21357419

AN - SCOPUS:79955439687

VL - 286

SP - 15308

EP - 15316

JO - Journal of Biological Chemistry

JF - Journal of Biological Chemistry

SN - 0021-9258

IS - 17

ER -