Stoichiometry of Scaffold Complexes in Living Neurons - DLC2 as a Dimerization Engine for GKAP

Enora Moutin, Vincent Compan, Fabrice Raynaud, Caroline Clerté, Nathalie Bouquier, Gilles Labesse, Matthew L. Ferguson, Laurent Fagni, Catherine A. Royer, Julie Perroy

Research output: Contribution to journalArticlepeer-review

10 Scopus citations

Abstract

Quantitative spatio-temporal characterization of protein interactions in living cells remains a major challenge facing modern biology. We have investigated in living neurons the spatial dependence of the stoichiometry of interactions between two core proteins of the NMDA receptor-associated scaffolding complex, GKAP and DLC2, using a novel variation of Fluorescence Fluctuation Microscopy called two-photon scanning Number and Brightness (sN&B). We found that dimerization of DLC2 was required for its interaction with GKAP, which in turn potentiated GKAP self-association. In dendritic shaft, the DLC2-GKAP hetero-oligomeric complexes were composed mainly of 2 DLC2 and 2 GKAP monomers, while in spines, the hetero-complexes were much larger, with an average of ∼16 DLC2 and ∼13 GKAP. Disruption of the GKAP-DLC2 interaction strongly destabilized the oligomers, decreasing the spine-preferential localization of GKAP and inhibiting NMDA receptor activity. Hence, DLC2 serves a hub function in the control of glutamatergic transmission via ordering of GKAP-containing complexes in dendritic spines. Beyond illuminating the role of DLC2–GKAP interactions in glutamergic signalling, these data underscore the power of the sN&B approach for quantitative spatio-temporal imaging of other important protein complexes.

Original languageAmerican English
Pages (from-to)3451-3462
Number of pages12
JournalPhysics Faculty Publications and Presentations
Volume127
Issue number16
DOIs
StatePublished - 15 Aug 2014

Keywords

  • Bioluminescence resonance energy transfer
  • Dynein light chain
  • Guanylate kinase-associated protein
  • Oligomerization
  • Scaffold
  • Scanning number and brightness

EGS Disciplines

  • Physics

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