title: Hierarchical nanostructure and synergy of multimolecular signalling complexes.
abstract: Signalling complexes are dynamic, multimolecular structures and sites for intracellular signal transduction. Although they play a crucial role in cellular activation, current research techniques fail to resolve their structure in intact cells. Here we present a multicolour, photoactivated localization microscopy approach for imaging multiple types of single molecules in fixed and live cells and statistical tools to determine the nanoscale organization, topology and synergy of molecular interactions in signalling complexes downstream of the T-cell antigen receptor. We observe that signalling complexes nucleated at the key adapter LAT show a hierarchical topology. The critical enzymes PLCγ1 and VAV1 localize to the centre of LAT-based complexes, and the adapter SLP-76 and actin molecules localize to the periphery. Conditional second-order statistics reveal a hierarchical network of synergic interactions between these molecules. Our results extend our understanding of the nanostructure of signalling complexes and are relevant to studying a wide range of multimolecular complexes.
authors.affiliation: Racah Institute of Physics, The Hebrew University, Jerusalem 91904, Israel.
authors.forename: Eilon
authors.initials: E
authors.lastname: Sherman
authors.affiliation: Laboratory of Cellular and Molecular Biology, CCR, NCI, NIH, Bethesda, Maryland 20892, USA.
authors.forename: Valarie A
authors.initials: VA
authors.lastname: Barr
authors.affiliation: Laboratory of Cellular and Molecular Biology, CCR, NCI, NIH, Bethesda, Maryland 20892, USA.
authors.forename: Robert K
authors.initials: RK
authors.lastname: Merrill
authors.affiliation: Laboratory of Cellular and Molecular Biology, CCR, NCI, NIH, Bethesda, Maryland 20892, USA.
authors.forename: Carole K
authors.initials: CK
authors.lastname: Regan
authors.affiliation: Laboratory of Cellular and Molecular Biology, CCR, NCI, NIH, Bethesda, Maryland 20892, USA.
authors.forename: Connie L
authors.initials: CL
authors.lastname: Sommers
authors.affiliation: Laboratory of Cellular and Molecular Biology, CCR, NCI, NIH, Bethesda, Maryland 20892, USA.
authors.forename: Lawrence E
authors.initials: LE
authors.lastname: Samelson
mesh_terms: D000199:Actins / Q000378:metabolism; D048868:Adaptor Proteins, Signal Transducing / Q000378:metabolism*; D000818:Animals; D006801:Humans; D019169:Jurkat Cells; D008565:Membrane Proteins / Q000378:metabolism* / Q000648:ultrastructure; D051379:Mice; D046912:Multiprotein Complexes / Q000648:ultrastructure*; D049329:Nanostructures; D051966:Phospholipase C gamma / Q000378:metabolism; D010750:Phosphoproteins / Q000378:metabolism; D011948:Receptors, Antigen, T-Cell / Q000378:metabolism; D015398:Signal Transduction; D000072760:Single Molecule Imaging / Q000379:methods*; D013601:T-Lymphocytes / Q000378:metabolism* / Q000648:ultrastructure
chemical_list: D000199:Actins; D048868:Adaptor Proteins, Signal Transducing; C110810:LAT protein, human; D008565:Membrane Proteins; D046912:Multiprotein Complexes; D010750:Phosphoproteins; D011948:Receptors, Antigen, T-Cell; C092947:SLP-76 signal Transducing adaptor proteins; C522157:PLCG1 protein, human; D051966:Phospholipase C gamma