Spatial Multiplexing of Fluorescent Reporters for Imaging Signaling Network Dynamics

Changyang Linghu, Shannon L. Johnson, Pablo A. Valdes, Or A. Shemesh, Won Min Park, Demian Park, Kiryl D. Piatkevich, Asmamaw T. Wassie, Yixi Liu, Bobae An, Stephanie A. Barnes, Orhan T. Celiker, Chun Chen Yao, Chih Chieh (Jay) Yu, Ru Wang, Katarzyna P. Adamala, Mark F. Bear, Amy E. Keating, Edward S. Boyden

Research output: Contribution to journalArticlepeer-review


In order to analyze how a signal transduction network converts cellular inputs into cellular outputs, ideally one would measure the dynamics of many signals within the network simultaneously. We found that, by fusing a fluorescent reporter to a pair of self-assembling peptides, it could be stably clustered within cells at random points, distant enough to be resolved by a microscope but close enough to spatially sample the relevant biology. Because such clusters, which we call signaling reporter islands (SiRIs), can be modularly designed, they permit a set of fluorescent reporters to be efficiently adapted for simultaneous measurement of multiple nodes of a signal transduction network within single cells. We created SiRIs for indicators of second messengers and kinases and used them, in hippocampal neurons in culture and intact brain slices, to discover relationships between the speed of calcium signaling, and the amplitude of PKA signaling, upon receiving a cAMP-driving stimulus.

Original languageEnglish (US)
Pages (from-to)1682-1698.e24
Issue number6
StatePublished - Dec 10 2020
Externally publishedYes


  • cAMP
  • calcium imaging
  • fluorescent reporters
  • live-cell imaging
  • protein kinase
  • protein scaffold
  • signal transduction
  • signaling pathway
  • signaling reporter islands
  • spatial multiplexing

ASJC Scopus subject areas

  • General Biochemistry, Genetics and Molecular Biology


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