title: Phase-locked signals elucidate circuit architecture of an oscillatory pathway.
abstract: This paper introduces the concept of phase-locking analysis of oscillatory cellular signaling systems to elucidate biochemical circuit architecture. Phase-locking is a physical phenomenon that refers to a response mode in which system output is synchronized to a periodic stimulus; in some instances, the number of responses can be fewer than the number of inputs, indicative of skipped beats. While the observation of phase-locking alone is largely independent of detailed mechanism, we find that the properties of phase-locking are useful for discriminating circuit architectures because they reflect not only the activation but also the recovery characteristics of biochemical circuits. Here, this principle is demonstrated for analysis of a G-protein coupled receptor system, the M3 muscarinic receptor-calcium signaling pathway, using microfluidic-mediated periodic chemical stimulation of the M3 receptor with carbachol and real-time imaging of resulting calcium transients. Using this approach we uncovered the potential importance of basal IP3 production, a finding that has important implications on calcium response fidelity to periodic stimulation. Based upon our analysis, we also negated the notion that the Gq-PLC interaction is switch-like, which has a strong influence upon how extracellular signals are filtered and interpreted downstream. Phase-locking analysis is a new and useful tool for model revision and mechanism elucidation; the method complements conventional genetic and chemical tools for analysis of cellular signaling circuitry and should be broadly applicable to other oscillatory pathways.
authors.affiliation: Biomedical Engineering Department, University of Michigan, Ann Arbor, Michigan, United States of America.
authors.forename: Andreja
authors.initials: A
authors.lastname: Jovic
authors.forename: Bryan
authors.initials: B
authors.lastname: Howell
authors.forename: Michelle
authors.initials: M
authors.lastname: Cote
authors.forename: Susan M
authors.initials: SM
authors.lastname: Wade
authors.forename: Khamir
authors.initials: K
authors.lastname: Mehta
authors.forename: Atsushi
authors.initials: A
authors.lastname: Miyawaki
authors.forename: Richard R
authors.initials: RR
authors.lastname: Neubig
authors.forename: Jennifer J
authors.initials: JJ
authors.lastname: Linderman
authors.forename: Shuichi
authors.initials: S
authors.lastname: Takayama
mesh_terms: D020013:Calcium Signaling / Q000502:physiology*; D002217:Carbachol; D019295:Computational Biology / Q000379:methods*; D057809:HEK293 Cells; D006801:Humans; D008954:Models, Biological; D043587:Receptor, Muscarinic M3; D015203:Reproducibility of Results
chemical_list: D043587:Receptor, Muscarinic M3; D002217:Carbachol