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Förster Resonance Energy Transfer (FRET) Correlates of Altered Subunit Stoichiometry in Cys-Loop Receptors, Exemplified by Nicotinic α4β2

Srinivasan, Rahul and Richards, Christopher I. and Dilworth, Crystal and Moss, Fraser J. and Dougherty, Dennis A. and Lester, Henry A. (2012) Förster Resonance Energy Transfer (FRET) Correlates of Altered Subunit Stoichiometry in Cys-Loop Receptors, Exemplified by Nicotinic α4β2. International Journal of Molecular Sciences, 13 (8). pp. 10022-10040. ISSN 1422-0067. PMCID PMC3431844. doi:10.3390/ijms130810022. https://resolver.caltech.edu/CaltechAUTHORS:20120907-141513430

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Abstract

We provide a theory for employing Förster resonance energy transfer (FRET) measurements to determine altered heteropentameric ion channel stoichiometries in intracellular compartments of living cells. We simulate FRET within nicotinic receptors (nAChRs) whose α4 and β2 subunits contain acceptor and donor fluorescent protein moieties, respectively, within the cytoplasmic loops. We predict FRET and normalized FRET (NFRET) for the two predominant stoichiometries, (α4)3(β2)2 vs. (α4)2(β2)3. Studying the ratio between FRET or NFRET for the two stoichiometries, minimizes distortions due to various photophysical uncertainties. Within a range of assumptions concerning the distance between fluorophores, deviations from plane pentameric geometry, and other asymmetries, the predicted FRET and NFRET for (α4)3(β2)2 exceeds that of (α4)2(β2)3. The simulations account for published data on transfected Neuro2a cells in which α4β2 stoichiometries were manipulated by varying fluorescent subunit cDNA ratios: NFRET decreased monotonically from (α4)3(β2)2 stoichiometry to mostly (α4)2(β2)3. The simulations also account for previous macroscopic and single-channel observations that pharmacological chaperoning by nicotine and cytisine increase the (α4)2(β2)3 and (α4)3(β2)2 populations, respectively. We also analyze sources of variability. NFRET-based monitoring of changes in subunit stoichiometry can contribute usefully to studies on Cys-loop receptors.


Item Type:Article
Related URLs:
URLURL TypeDescription
http://dx.doi.org/10.3390/ijms130810022DOIArticle
http://www.mdpi.com/1422-0067/13/8/10022PublisherArticle
https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3431844/PubMed CentralArticle
ORCID:
AuthorORCID
Moss, Fraser J.0000-0002-8519-6991
Dougherty, Dennis A.0000-0003-1464-2461
Lester, Henry A.0000-0002-5470-5255
Additional Information:© 2012 by the authors; licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution license (http://creativecommons.org/licenses/by/3.0/). Received: 30 May 2012; in revised form: 2 July 2012; Accepted: 5 July 2012; Published: 10 August 2012. We thank Kimberly Scott for discussions. Supported by grants from the U.S. National Institutes of Health (NS-11756, NS-34407, AG-033954), Targacept Inc., Louis and Janet Fletcher, the Michael J. Fox Foundation, the California Tobacco-Related Disease Research Program postdoctoral fellowship (18FT-0066 to R.S.), a Beckman Institute Fellowship (C.I.R) and an NIH Kirschstein National Research Service Award (DA030877 to C.I.R).
Funders:
Funding AgencyGrant Number
NIHNS-11756
NIHNS-34407
NIHAG-033954
Targacept Inc.UNSPECIFIED
Louis and Janet FletcherUNSPECIFIED
Michael J. Fox FoundationUNSPECIFIED
California Tobacco-Related Disease Research Program18FT-0066
Caltech Beckman InstituteUNSPECIFIED
NIH Predoctoral FellowshipDA030877
Subject Keywords:nicotine; cytisine; NFRET; nicotine addiction; Parkinson’s disease; ion channels
Issue or Number:8
PubMed Central ID:PMC3431844
DOI:10.3390/ijms130810022
Record Number:CaltechAUTHORS:20120907-141513430
Persistent URL:https://resolver.caltech.edu/CaltechAUTHORS:20120907-141513430
Official Citation:Srinivasan, R.; Richards, C.I.; Dilworth, C.; Moss, F.J.; Dougherty, D.A.; Lester, H.A. Förster Resonance Energy Transfer (FRET) Correlates of Altered Subunit Stoichiometry in Cys-Loop Receptors, Exemplified by Nicotinic α4β2. Int. J. Mol. Sci. 2012, 13, 10022-10040
Usage Policy:This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution license (http://creativecommons.org/licenses/by/3.0/).
ID Code:33941
Collection:CaltechAUTHORS
Deposited By: Tony Diaz
Deposited On:07 Sep 2012 21:31
Last Modified:09 Nov 2021 23:05

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