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Proc Natl Acad Sci U S A. 2015 Mar 17;112(11):3338-43. doi: 10.1073/pnas.1502857112. Epub 2015 Mar 2.

Rationally designed fluorogenic protease reporter visualizes spatiotemporal dynamics of apoptosis in vivo.

Author information

1
Departments of Pharmaceutical Chemistry and Cardiovascular Research Institute, and Yuhnung.jan@ucsf.edu Xiaokun.Shu@ucsf.edu.
2
Cardiovascular Research Institute, and Physiology, Howard Hughes Medical Institute, University of California, San Francisco, CA 94158.
3
Departments of Pharmaceutical Chemistry and Cardiovascular Research Institute, and.
4
Cardiovascular Research Institute, and Physiology, Howard Hughes Medical Institute, University of California, San Francisco, CA 94158 Yuhnung.jan@ucsf.edu Xiaokun.Shu@ucsf.edu.

Abstract

Fluorescence resonance energy transfer-based reporters have been widely used in imaging cell signaling; however, their in vivo application has been handicapped because of poor signal. Although fluorogenic reporters overcome this problem, no such reporter of proteases has been demonstrated for in vivo imaging. Now we have redesigned an infrared fluorescent protein so that its chromophore incorporation is regulated by protease activity. Upon protease activation, the infrared fluorogenic protease reporter becomes fluorescent with no requirement of exogenous cofactor. To demonstrate biological applications, we have designed an infrared fluorogenic executioner-caspase reporter, which reveals spatiotemporal coordination between cell apoptosis and embryonic morphogenesis, as well as dynamics of apoptosis during tumorigenesis in Drosophila. The designed scaffold may be used to engineer reporters of other proteases with specific cleavage sequence.

KEYWORDS:

apoptosis; bacterial phytochrome; fluorogenic reporters; protease; tumor development

PMID:
25733847
PMCID:
PMC4371907
DOI:
10.1073/pnas.1502857112
[Indexed for MEDLINE]
Free PMC Article

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