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Edited* by Peter N. Devreotes, Johns Hopkins University School of Medicine, Baltimore, MD, and approved February 12, 2013 (received for review November 29, 2012)
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Significance
G-protein–coupled receptors control a variety of important cell behaviors. However, tools are not available to activate these receptors in selected areas of a cell and exert control over cell behavior. Here we recruit unique properties of nonrhodopsin opsins to activate all the major types of G-protein signaling in spatially confined regions of single cells. We show that this approach can be used to optically induce polarized cell behavior and refashion early neuron differentiation. This optical approach can be applied to control other cell behaviors such as immune cell migration and cardiomyocyte contraction.
Abstract
G-protein–coupled receptor (GPCR) activity gradients evoke important cell behavior but there is a dearth of methods to induce such asymmetric signaling in a cell. Here we achieved reversible, rapidly switchable patterns of spatiotemporally restricted GPCR activity in a single cell. We recruited properties of nonrhodopsin opsins—rapid deactivation, distinct spectral tuning, and resistance to bleaching—to activate native Gi, Gq, or Gs signaling in selected regions of a cell. Optical inputs were designed to spatiotemporally control levels of second messengers, IP3, phosphatidylinositol (3,4,5)-triphosphate, and cAMP in a cell. Spectrally selective imaging was accomplished to simultaneously monitor optically evoked molecular and cellular response dynamics. We show that localized optical activation of an opsin-based trigger can induce neurite initiation, phosphatidylinositol (3,4,5)-triphosphate increase, and actin remodeling. Serial optical inputs to neurite tips can refashion early neuron differentiation. Methods here can be widely applied to program GPCR-mediated cell behaviors.
Footnotes
- ↵1To whom correspondence should be addressed. E-mail: gautam{at}wustl.edu.
Author contributions: W.K.A.K. and N.G. designed research; W.K.A.K. performed research; W.K.A.K. and V.K. contributed new reagents/analytic tools; W.K.A.K. and L.G. analyzed data; and W.K.A.K., L.G., and N.G. wrote the paper.
The authors declare no conflict of interest.
↵*This Direct Submission article had a prearranged editor.
This article contains supporting information online at www.pnas.org/lookup/suppl/doi:10.1073/pnas.1220697110/-/DCSupplemental.
Confined optical activation of GPCRs in a cell
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