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Acoustic biosensors for ultrasound imaging of enzyme activity

Lakshmanan, Anupama and Jin, Zhiyang and Nety, Suchita P. and Sawyer, Daniel P. and Lee-Gosselin, Audrey and Malounda, Dina and Swift, Mararet B. and Maresca, David and Shapiro, Mikhail G. (2020) Acoustic biosensors for ultrasound imaging of enzyme activity. Nature Chemical Biology, 16 (9). pp. 988-996. ISSN 1552-4450. https://resolver.caltech.edu/CaltechAUTHORS:20200713-133644814

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[img] Image (JPEG) (Extended Data Fig. 1: Engineering an acoustic sensor of TEV endopeptidase activity) - Supplemental Material
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[img] Image (JPEG) (Extended Data Fig. 2: Engineering an acoustic sensor of calpain activity) - Supplemental Material
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[img] Image (JPEG) (Extended Data Fig. 3: Characterization of GVWT sample with calpain protease) - Supplemental Material
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[img] Image (JPEG) (Extended Data Fig. 4: Engineering an acoustic sensor of ClpXP proteolytic activity -- corrected) - Supplemental Material
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[img] Image (JPEG) (Extended Data Fig. 5: Constructing intracellular acoustic sensor genes for dynamic monitoring of protease activity and circuit-driven gene expression) - Supplemental Material
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[img] Image (JPEG) (Extended Data Fig. 6: Schematic illustrating the in vivo ultrasound imaging experiment) - Supplemental Material
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[img] Image (JPEG) (Extended Data Fig. 7: Ultrasound imaging of bacteria expressing acoustic sensor genes in the gastrointestinal tract of mice. -- corrected) - Supplemental Material
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[img] Image (JPEG) (Extended Data Fig. 8: ASGClpXP -expressing cells showed higher contrast to tissue with nonlinear imaging) - Supplemental Material
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Abstract

Visualizing biomolecular and cellular processes inside intact living organisms is a major goal of chemical biology. However, existing molecular biosensors, based primarily on fluorescent emission, have limited utility in this context due to the scattering of light by tissue. In contrast, ultrasound can easily image deep tissue with high spatiotemporal resolution, but lacks the biosensors needed to connect its contrast to the activity of specific biomolecules such as enzymes. To overcome this limitation, we introduce the first genetically encodable acoustic biosensors—molecules that ‘light up’ in ultrasound imaging in response to protease activity. These biosensors are based on a unique class of air-filled protein nanostructures called gas vesicles, which we engineered to produce nonlinear ultrasound signals in response to the activity of three different protease enzymes. We demonstrate the ability of these biosensors to be imaged in vitro, inside engineered probiotic bacteria, and in vivo in the mouse gastrointestinal tract.


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https://doi.org/10.1038/s41589-020-0591-0DOIArticle
https://rdcu.be/b5ACsPublisherFree ReadCube access
https://doi.org/10.1038/s41589-020-0630-xDOIPublisher Correction
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https://pubs.acs.org/doi/10.1021/cen-09828-scicon3Featured InC&EN -- Science Concentrates
ORCID:
AuthorORCID
Lakshmanan, Anupama0000-0002-6702-837X
Jin, Zhiyang0000-0002-4411-6991
Nety, Suchita P.0000-0002-4201-1061
Sawyer, Daniel P.0000-0003-2926-191X
Lee-Gosselin, Audrey0000-0002-2431-2741
Malounda, Dina0000-0001-7086-9877
Maresca, David0000-0002-4921-6406
Shapiro, Mikhail G.0000-0002-0291-4215
Additional Information:© 2020 Springer Nature Limited. Received 18 August 2019; Accepted 12 June 2020; Published 13 July 2020. The authors thank Z. Sun, A. Shur and R. Murray for sharing the protocols and reagents used for the cell-free TX–TL system. TEM was done at the Beckman Institute Resource Center for Transmission Electron Microscopy at Caltech. This research was supported by the National Institutes of Health (NIH) (no. R01-EB018975) and Defense Advanced Research Projects Agency (no. W911NF-14-1-0111). A.L. was supported by a National Science Foundation (NSF) Graduate Research Fellowship (no. 1144469) and the Biotechnology Leadership Pre-doctoral Training Program in Micro/Nanomedicine (Rosen Bioengineering Center and NIH Training Grant no. 5T32GM112592-03/04). D.P.S. was supported by an NSF Graduate Research Fellowship (no. 1745301). D.M. was supported by the Human Frontier Science Program (no. LT000637/2016). Related research in the Shapiro Laboratory is supported by the Heritage Medical Research Institute, Burroughs Wellcome Career Award at the Scientific Interface, Pew Scholarship in the Biomedical Sciences and Packard Fellowship for Science and Engineering. Author Contributions: A.L. and M.G.S. conceived the study. A.L., Z.J. and S.P.N. designed and planned the experiments. A.L., Z.J., S.P.N., D.P.S., A.L-G., M.B.S. and D. Malounda conducted the experiments. Z.J., D.P.S. and D. Maresca wrote the MATLAB scripts for ultrasound imaging and data processing. A.L., Z.J. and M.G.S. analyzed the data. A.L., Z.J. and M.G.S. wrote the manuscript with input from all authors. All authors gave approval to the final version of the manuscript. The authors declare no competing interests.
Errata:Correction to: Nature Chemical Biology https://doi.org/10.1038/s41589-020-0591-0, published online 13 July 2020 In the version of this article originally published, there were errors in Fig. 4 and Extended Data Figs. 5 and 7. In Fig. 4f, labels “+ aTc” and “− atc” were on the wrong sides. “+ aTc” should be placed over the blue bar on the right-hand column, and “− aTc” should be on the left. In Extended Data Fig. 5b, the colormap accompanying the nonlinear ultrasound image was blank. Additionally, the label "ASGWT" on the left should be "ARGWT", and the orange and blue overbar colors should be swapped, with orange on the right and blue on the left. In Extended Data Fig. 7, the colormaps accompanying the nonlinear ultrasound images in b and c were blank, and the significance asterisks above the nonlinear data in d and e were shown as crossed squares. The errors have been corrected in the HTML and PDF versions of the paper. Published: 23 July 2020.
Group:Heritage Medical Research Institute
Funders:
Funding AgencyGrant Number
NIHR01-EB018975
Defense Advanced Research Projects Agency (DARPA)W911NF-14-1-0111
NSF Graduate Research FellowshipDGE-1144469
NIH5T32GM112592-03/04
NSF Graduate Research FellowshipDGE-1745301
Human Frontier Science ProgramLT000637/2016
Heritage Medical Research InstituteUNSPECIFIED
Burroughs Wellcome FundUNSPECIFIED
Pew Charitable TrustUNSPECIFIED
David and Lucile Packard FoundationUNSPECIFIED
Subject Keywords:Imaging; Proteases; Protein design; Synthetic biology
Issue or Number:9
Record Number:CaltechAUTHORS:20200713-133644814
Persistent URL:https://resolver.caltech.edu/CaltechAUTHORS:20200713-133644814
Official Citation:Lakshmanan, A., Jin, Z., Nety, S.P. et al. Acoustic biosensors for ultrasound imaging of enzyme activity. Nat Chem Biol 16, 988–996 (2020). https://doi.org/10.1038/s41589-020-0591-0
Usage Policy:No commercial reproduction, distribution, display or performance rights in this work are provided.
ID Code:104360
Collection:CaltechAUTHORS
Deposited By: Tony Diaz
Deposited On:13 Jul 2020 21:01
Last Modified:20 Aug 2020 20:02

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