Ultrasound Imaging of Gene Expression in Mammalian Cells
The study of cellular processes occurring inside intact organisms requires methods to visualize cellular functions such as gene expression in deep tissues. Ultrasound is a widely used biomedical technology enabling noninvasive imaging with high spatial and temporal resolution. However, no genetically encoded molecular reporters are available to connect ultrasound contrast to gene expression in mammalian cells. To address this limitation, we introduce mammalian acoustic reporter genes. Starting with a gene cluster derived from bacteria, we engineered a eukaryotic genetic program whose introduction into mammalian cells results in the expression of intracellular air-filled protein nanostructures called gas vesicles, which produce ultrasound contrast. Mammalian acoustic reporter genes allow cells to be visualized at volumetric densities below 0.5% and permit high-resolution imaging of gene expression in living animals.
© 2019 American Association for the Advancement of Science. This is an article distributed under the terms of the Science Journals Default License. Received 28 March 2019; accepted 30 August 2019. We thank D. Maresca, B. Ling, and A. Bar-Zion for help with ultrasound imaging; N. Koulena for assistance with tissue histology; M. Abedi, J. Lee, and M. Y. You for assistance with flow cytometry experiments; A. Collazo for confocal microscopy; C. Buser and the Oak Crest Institute of Science for cell sectioning and staining; and E. Jue and W. Chour for assistance with initial experiments. Electron microscopy was performed at the Beckman Institute Resource Center for Transmission Electron Microscopy at Caltech. Fluorescence imaging of tissues was performed in the Biological Imaging Facility of the Caltech Beckman Institute with support from the Arnold and Mabel Beckman Foundation. We appreciate the help of A. Lee-Gosselin and Caltech's Office of Laboratory Animal Research with animal protocols and husbandry. Funding: A.F. was supported by an NSERC graduate fellowship. D.P.S. was supported by an NSF graduate research fellowship (grant no. 1745301). This research was supported by the National Institutes of Health (grant nos. R01EB018975 and U54CA199090 to M.G.S.), the Heritage Medical Research Institute (M.G.S.), the Packard Fellowship for Science and Engineering (M.G.S.), and the Burroughs Welcome Fund Career Award at the Scientific Interface (M.G.S.). Author contributions: A.F. and M.G.S. conceived and planned the research. A.F. and G.H.H. performed the experiments. A.F. and R.W.B. designed the DNA sequences. A.F. and D.P.S. designed and optimized the ultrasound imaging sequences. A.F. analyzed the data. A.F. and M.G.S. wrote the manuscript with input from all authors. M.G.S. supervised the research. Competing interests: The authors declare no competing financial interests. The authors are inventors on patent applications related to ARGs filed by the California Institute of Technology. Data and materials availability: The mARG genetic construct will be deposited with Addgene at the time of manuscript publication. Raw data are available from the authors upon reasonable request.
Submitted - 580647.full.pdf
Supplemental Material - aax4804-Farhadi-SM.pdf