title: Quantification of receptor tyrosine kinase transactivation through direct dimerization and surface density measurements in single cells.
abstract: Cell signaling involves dynamic changes in protein oligomerization leading to the formation of different signaling complexes and modulation of activity. Spatial intensity distribution analysis (SpIDA) is an image analysis method that can directly measure oligomerization and trafficking of endogenous proteins in single cells. Here, we show the use of SpIDA to quantify dimerization/activation and surface transport of receptor protein kinases--EGF receptor and TrkB--at early stages of their transactivation by several G protein-coupled receptors (GPCRs). Transactivation occurred on the same timescale and was directly limited by GPCR activation but independent of G-protein coupling types. Early receptor protein kinase transactivation and internalization were not interdependent for all receptor pairs tested, revealing heterogeneity between groups of GPCRs. SpIDA also detected transactivation of TrkB by dopamine receptors in intact neurons. By allowing for time and space resolved quantification of protein populations with heterogeneous oligomeric states, SpIDA provides a unique approach to undertake single cell multivariate quantification of signaling processes involving changes in protein interactions, trafficking, and activity.
authors.affiliation: Departments of Chemistry and Physics, McGill University, Montreal, QC, Canada H3G 0B1.
authors.forename: Jody L
authors.initials: JL
authors.lastname: Swift
authors.forename: Antoine G
authors.initials: AG
authors.lastname: Godin
authors.forename: Kim
authors.initials: K
authors.lastname: Doré
authors.forename: Laure
authors.initials: L
authors.lastname: Freland
authors.forename: Nathalie
authors.initials: N
authors.lastname: Bouchard
authors.forename: Chelsea
authors.initials: C
authors.lastname: Nimmo
authors.forename: Mikhail
authors.initials: M
authors.lastname: Sergeev
authors.forename: Yves
authors.initials: Y
authors.lastname: De Koninck
authors.forename: Paul W
authors.initials: PW
authors.lastname: Wiseman
authors.forename: Jean-Martin
authors.initials: JM
authors.lastname: Beaulieu
mesh_terms: D000818:Animals; D016466:CHO Cells; D006224:Cricetinae; D003412:Cricetulus; D066246:ErbB Receptors / Q000235:genetics / Q000378:metabolism*; D051379:Mice; D008822:Mice, Transgenic; D008856:Microscopy, Fluorescence; D009474:Neurons / Q000166:cytology / Q000378:metabolism*; D055503:Protein Multimerization / Q000502:physiology*; D020813:Receptor, trkB / Q000235:genetics / Q000378:metabolism*; D011954:Receptors, Dopamine / Q000235:genetics / Q000378:metabolism*; D015533:Transcriptional Activation / Q000502:physiology*
chemical_list: D011954:Receptors, Dopamine; D066246:ErbB Receptors; D020813:Receptor, trkB