Notes: These authors developed luciferase-based biosensors to monitor target cell-specific apoptosis in real time. They used GloSensor™  technology to develop biosensors containing a circularly permuted luciferase linked internally by either caspase 3/7 or granzyme B/caspase 8 cleavage sites.  Cleavage by the respective proteases causes activation of bioluminescence, providing a convenient way to study patterns of caspase activation by different cells of the immune system. (4527)

Notes: These authors generated luciferase biosensors and used them to screen for potential inhibitors of Middle East respiratory syndrome coronavirus (MERS-CoV) replication. The biosensors were developed using the pGloSensor™-30F plasmid backbone and were engineered to include cleavage sites for the viral papain-like protease (PLpro) or 3-chymotrypsin-like protease (3CLpro). Lytic, endpoint assays and live-cell assays were performed to detect protease activity. The authors identified a small-molecule inhibitor that blocked the activity of MERS-CoV 3CLpro. These biosensor-based assays allowed rapid evaluation of viral protease activity and screening for protease inhibitors. (4526)

Notes:

This paper describes use of GloSenso™ technology to develop a luciferase biosensor for detection of caspase 3/7 activity. The authors stably expressed a caspase-3/7 biosensor (pGloSensor™-30F DEVDG Vector, available as a custom product from Promega) in 1833 breast cells and D54 glioma cells, which were then used to monitor caspase activation in cell culture following addition of the cell-permeable substrate (GloSensor™ cAMP Reagent). The paper demonstrates use of this  luciferase biosensor to detect apoptosis in vivo in cultured cells and in a mouse model, and also demonstrates the utility of the technology for high-throughput screening for pharmacologically active compounds. The regulation of apoptosis by caspases is used as an example in this study, biosensors to study other proteases involved in the regulation of cellular processes can be designed using the concepts described in the paper. (4525)