Activation of caspase-3 during <i>Chlamydia trachomatis</i>-induced apoptosis at a late stage.

Obligate intracellular bacterium <i>Chlamydia trachomatis </i>activates the host cell apoptosis pathway at a late stage of its developmental cycle. However, whether caspase-3, which is a key enzyme of apoptosis, is activated in <i>Chlamydia</i>-infected cells remains unknown. Here, we established HEp-2 cells stably expressing cFluc-DEVD, which is a caspase-3 substrate sequence inserted into cyclic firefly luciferase, and then monitored the dynamics of caspase-3 activity in cells infected with <i>Chlamydia</i>. Transfected cells without infection showed a significant increase in luciferase activity due to stimulation with staurosporine, an inducer of apoptosis. Activation was significantly blocked by addition of caspase inhibitor z-VAD-fmk. Furthermore, as expected, <i>Chlamydia</i> infection caused a significant increase in luciferase activation at 36-48 h post-infection with a contrastive decrease at 24 h post-infection as already well known. Such activation caused by the infection was much stronger with an increase in the amount of bacteria. Thus, caspase-3 activation was accurately monitored by the luciferase activity in HEp-2 cells constitutively expressing the cFluc-DEVD probe. Furthermore, our data showed that <i>C. trachomatis </i>activates caspase-3 in host cells at a late stage of infection.