Assessing the cleavage of gasdermin-family cell death effectors by microbial proteases

Gasdermin-family proteins initiate the lytic cell death program pyroptosis upon proteolytic removal of an autoinhibitory domain. For each gasdermin, proteases capable of mediating an activating cleavage event have been clearly defined. However, for new proteases, there are unique challenges in evaluating whether a specific cleavage event results in an active or inactive gasdermin. This chapter outlines methods and key controls suitable for the initial discovery and evaluation of protease activation of gasdermins that are particularly suitable for the study of alternative pathways for activation, such as by the proteases of bacteria and viruses.