Rationally designed fluorogenic protease reporter visualizes spatiotemporal dynamics of apoptosis in vivo.
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.
Departments of Pharmaceutical Chemistry and Cardiovascular Research Institute, and Yuhnung.jan@ucsf.edu Xiaokun.Shu@ucsf.edu.
Tsz-Leung
TL
To
Cardiovascular Research Institute, and Physiology, Howard Hughes Medical Institute, University of California, San Francisco, CA 94158.
Beverly J
BJ
Piggott
Departments of Pharmaceutical Chemistry and Cardiovascular Research Institute, and.
Kalpana
K
Makhijani
Departments of Pharmaceutical Chemistry and Cardiovascular Research Institute, and.
Dan
D
Yu
Cardiovascular Research Institute, and Physiology, Howard Hughes Medical Institute, University of California, San Francisco, CA 94158 Yuhnung.jan@ucsf.edu Xiaokun.Shu@ucsf.edu.
Yuh Nung
YN
Jan
Departments of Pharmaceutical Chemistry and Cardiovascular Research Institute, and Yuhnung.jan@ucsf.edu Xiaokun.Shu@ucsf.edu.
Xiaokun
X
Shu
D000818:Animals; D017209:Apoptosis; D063646:Carcinogenesis / Q000473:pathology; D020169:Caspases / Q000378:metabolism; D004331:Drosophila melanogaster / Q000166:cytology* / Q000196:embryology; D004625:Embryo, Nonmammalian / Q000166:cytology; D047108:Embryonic Development; D005456:Fluorescent Dyes / Q000378:metabolism*; D017930:Genes, Reporter; D057809:HEK293 Cells; D006801:Humans; D007259:Infrared Rays; D010447:Peptide Hydrolases / Q000378:metabolism; D013997:Time Factors
D005456:Fluorescent Dyes; D010447:Peptide Hydrolases; D020169:Caspases
apoptosis; bacterial phytochrome; fluorogenic reporters; protease; tumor development