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Quantification of target gene expression by imaging reporter gene expression in living animals

Nature Medicine volume 6, pages 933–937 (2000)Cite this article

At present, most methods to image gene expression in rodents, nonhuman primates and humans are limited to the use of fixed tissue obtained from biopsy or after death. Fixed tissue can be assayed for mRNA levels or a protein product using well-established techniques. Reporter genes sensitive to activation status of endogenous genes have also been used by molecular biologists. Endogenous gene expression is monitored through creation of a chimeric fusion gene in which the promoter of an endogenous gene is coupled to the reporter gene. If the endogenous gene promoter is ‘turned off’, neither the endogenous gene nor the reporter gene is transcribed. Similarly, if the promoter for the endogenous gene is activated, then the reporter gene is transcribed. Although introduction of a chimeric fusion gene into the target organism is required, this allows for monitoring of any endogenous gene of interest where the promoter is sequenced. However, conventional reporter gene methods are generally limited by their inability to determine the location(s), magnitude and time variation of gene expression in living animals.

Newer approaches using green fluorescent protein (GFP) or luciferase as reporter genes support localization of gene expression through video imaging. Approaches using GFP or luciferase as reporter genes work for cells and for living organisms transparent to light, and from a limited depth in small animals1. These techniques are limited because they cannot be generalized (for example, luciferase would not work with most human studies) and because of a lack of tomographic resolution and the inability to quantify gene expression fully. A method harnessing the power of reporter gene technology, which is noninvasive, quantitative and applicable to animals and humans, could advance research in many fields, including monitoring of human gene therapy.

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Figure 1: Measuring target gene expression indirectly by imaging reporter gene expression when the two genes are simultaneously expressed from a bi-cistronic vector containing an IRES.
Figure 2: Attenuation of gene expression in bi-cistronic vectors studied by transient transfection of C6 cells.
Figure 3: Correlation of HSV1-sr39tk and D2R expression studied in transient and stable transfection studies.
Figure 4: Accumulation of 8-3H-PCV and FPCV correlates with HSV1-sr39tk expression in C6 cells stably transfected with pCMV-D2R-IRES-sr39tk.
Figure 5: MicroPET imaging of correlated bi-cistronic reporter gene expression.

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Acknowledgements

We thank M. Black, D. Kaufman, and the UCLA Gene Imaging Consortium for helpful discussions, and A. Green, K. Nguyen, E. Bauer, D. MacLaren, X. Sun, G. Castanedo, V. Dominguez, J. Edwards, W. Ladno, D. J. Liu and R. Sumida and the UCLA cyclotron ‘crew’ for technical assistance. This work was supported in part by funding from the Department of Energy DE-FC03-87ER60615, National Institutes of Health RO1 CA82214-01 and UCLA-Jonsson Comprehensive Cancer Center.

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Authors and Affiliations

  1. The Crump Institute for Molecular Imaging, the UCLA School of Medicine, Los Angeles, 90095–1770, California, USA

    Yajing Yu, Alexander J. Annala, Jorge R. Barrio, Tatsushi Toyokuni, Nagichettiar Satyamurthy, Mohammad Namavari, Simon R. Cherry, Michael E. Phelps, Harvey R. Herschman & Sanjiv S. Gambhir

  2. UCLA/Department of Energy Laboratory of Structural Biology & Molecular Medicine, the UCLA School of Medicine, Los Angeles, 90095–1770, California, USA

    Yajing Yu, Alexander J. Annala, Jorge R. Barrio, Nagichettiar Satyamurthy, Mohammad Namavari, Simon R. Cherry, Michael E. Phelps, Harvey R. Herschman & Sanjiv S. Gambhir

  3. Department of Molecular & Medical Pharmacology, the UCLA School of Medicine, Los Angeles, 90095–1770, California, USA

    Yajing Yu, Alexander J. Annala, Jorge R. Barrio, Tatsushi Toyokuni, Nagichettiar Satyamurthy, Mohammad Namavari, Simon R. Cherry, Michael E. Phelps, Harvey R. Herschman & Sanjiv S. Gambhir

  4. Molecular Biology Institute, the UCLA School of Medicine, Los Angeles, 90095–1770, California, USA

    Yajing Yu, Alexander J. Annala & Harvey R. Herschman

  5. Department of Biomathematics, the UCLA School of Medicine, Los Angeles, 90095–1770, California, USA

    Simon R. Cherry, Michael E. Phelps & Sanjiv S. Gambhir

  6. UCLA-Jonsson Comprehensive Cancer Center, UCLA School of Medicine, Los Angeles, 90095–1770, California, USA

    Harvey R. Herschman & Sanjiv S. Gambhir

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  1. Yajing Yu
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Correspondence to Sanjiv S. Gambhir.

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Yu, Y., Annala, A., Barrio, J. et al. Quantification of target gene expression by imaging reporter gene expression in living animals. Nat Med 6, 933–937 (2000). https://doi.org/10.1038/78704

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