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  • A novel firefly luciferase biosensor enhances the detection of apoptosis induced by ESAT-6 family proteins of Mycobacterium tuberculosis.

A novel firefly luciferase biosensor enhances the detection of apoptosis induced by ESAT-6 family proteins of Mycobacterium tuberculosis.

Biochemical and biophysical research communications (2014-09-23)
Junwei Shi, Huan Zhang, Liurong Fang, Yongqiang Xi, Yanrong Zhou, Rui Luo, Dang Wang, Shaobo Xiao, Huanchun Chen
ABSTRACT

The activation of caspase-3 is a key surrogate marker for detecting apoptosis. To quantitate caspase-3 activity, we constructed a biosensor comprising a recombinant firefly luciferase containing a caspase-3 cleavage site. When apoptosis was induced, caspase-3 cleavage of the biosensor activated firefly luciferase by a factor greater than 25. The assay conveniently detected apoptosis in real time, indicating that it will facilitate drug discovery. We screened ESAT-6 family proteins of Mycobacterium tuberculosis and found that esxA, esxT and esxL induced apoptosis. Further, activation of nuclear factor-κB (NF-κB) and the NF-κB-regulated genes encoding tumor necrosis factor-α (TNF-α) and tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) participated in esxT-induced apoptosis. We conclude that this assay is useful for high-throughput screening to identify and characterize proteins and drugs that regulate apoptosis.

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