Visualization of caspase-3-like activity in cells using a genetically encoded fluorescent biosensor activated by protein cleavage

Jiao Zhang, Xin Wang, Wenjing Cui, Wenwen Wang, Huamei Zhang, Lu Liu, Zicheng Zhang, Zheng Li, Guoguang Ying, Ning Zhang and Binghui Li ()
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Jiao Zhang: Laboratory of Cancer Cell Biology, Key Laboratory of Breast Cancer Prevention and Therapy, Tianjin Medical University Cancer Institute and Hospital
Xin Wang: Key Laboratory of Breast Cancer Prevention and Therapy, Tianjin Medical University Cancer Institute and Hospital
Wenjing Cui: Laboratory of Cancer Cell Biology, Key Laboratory of Breast Cancer Prevention and Therapy, Tianjin Medical University Cancer Institute and Hospital
Wenwen Wang: Laboratory of Cancer Cell Biology, Key Laboratory of Breast Cancer Prevention and Therapy, Tianjin Medical University Cancer Institute and Hospital
Huamei Zhang: Laboratory of Cancer Cell Biology, Key Laboratory of Breast Cancer Prevention and Therapy, Tianjin Medical University Cancer Institute and Hospital
Lu Liu: Laboratory of Cancer Cell Biology, Key Laboratory of Breast Cancer Prevention and Therapy, Tianjin Medical University Cancer Institute and Hospital
Zicheng Zhang: Laboratory of Cancer Cell Biology, Key Laboratory of Breast Cancer Prevention and Therapy, Tianjin Medical University Cancer Institute and Hospital
Zheng Li: Key Laboratory of Breast Cancer Prevention and Therapy, Tianjin Medical University Cancer Institute and Hospital
Guoguang Ying: Laboratory of Cancer Cell Biology, Key Laboratory of Breast Cancer Prevention and Therapy, Tianjin Medical University Cancer Institute and Hospital
Ning Zhang: Laboratory of Cancer Cell Biology, Key Laboratory of Breast Cancer Prevention and Therapy, Tianjin Medical University Cancer Institute and Hospital
Binghui Li: Laboratory of Cancer Cell Biology, Key Laboratory of Breast Cancer Prevention and Therapy, Tianjin Medical University Cancer Institute and Hospital

Nature Communications, 2013, vol. 4, issue 1, 1-13

Abstract: Abstract Cytosolic caspase-3-like proteases, such as caspase-3 and caspase-7, have a central role in mediating the progress of apoptosis. Here to conveniently monitor caspase-3-like activity in the multicellular environment, we have developed genetically encoded switch-on fluorescence-base indicators that are cyclized chimeras containing a caspase-3 cleavage site as a switch. When cleaved by caspase-3-like proteases, the non-fluorescent indicator rapidly becomes fluorescent, and thus detects in real-time the activation of such caspases. We generate cultured cells constitutively expressing these chimeras, and all the healthy cells are non-fluorescent. When these cells are exposed to apoptotic stimuli, dead cells show strong fluorescence depending on caspase activation. With these tools, we monitor in real-time caspase-3-like activity in each cell under various conditions, and show for the first time that the environment of cancer cells affects their sensitivity to chemotherapeutic drugs in a modified soft agar assay. These biosensors should enable better understanding of the biological relevance of caspase-3-like proteases.

Date: 2013
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DOI: 10.1038/ncomms3157

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