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Identification and characterization of novel nicotinic receptor‐associated proteins in Caenorhabditis elegans
Author(s) -
Gottschalk Alexander,
Almedom Ruta B,
Schedletzky Thorsten,
Anderson Scott D,
Yates John R,
Schafer William R
Publication year - 2005
Publication title -
the embo journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 7.484
H-Index - 392
eISSN - 1460-2075
pISSN - 0261-4189
DOI - 10.1038/sj.emboj.7600741
Subject(s) - biology , nicotinic acetylcholine receptor , caenorhabditis elegans , microbiology and biotechnology , acetylcholine receptor , receptor , neurotransmission , biochemistry , gene
Nicotinic acetylcholine receptors (nAChRs) mediate fast excitatory neurotransmission in neurons and muscles. To identify nAChR accessory proteins, which may regulate their expression or function, we performed tandem affinity purification of the levamisole‐sensitive nAChR from Caenorhabditis elegans , mass spectrometry of associated components, and RNAi‐based screening for effects on in vivo nicotine sensitivity. Among the proteins identified was the calcineurin A subunit TAX‐6, which appeared to function as a negative regulator of nAChR activity. We also identified five proteins not previously linked to nAChR function, whose inactivation conferred nicotine resistance, implicating them as positive regulators of nAChR activity. Of these, the copine NRA‐1 colocalized with the levamisole receptor at neuronal and muscle plasma membranes, and, when mutated, caused reduced synaptic nAChR expression. Loss of SOC‐1, which acts in receptor tyrosine kinase (RTK) signaling, also reduced synaptic levamisole receptor levels, as did mutations in the fibroblast growth factor receptor EGL‐15, and another RTK, CAM‐1. Thus, tandem affinity purification is a viable approach to identify novel proteins regulating neurotransmitter receptor activity or expression in model systems like C. elegans .