Cytolysin

A defining property of cytotoxic lymphocytes is their expression and regulated secretion of potent toxins, including the pore-forming protein perforin and serine protease granzymes. Until recently, mechanisms of pore formation and granzyme transfer into the target cell were poorly understood, but advances in structural and cellular biology have now begun to unravel how synergy between perforin and granzymes brings about target cell death. These and other advances are demonstrating the surprisingly broad pathophysiological roles of the perforin-granzyme pathway, and this has important implications for understanding immune homeostasis and for developing immunotherapies for cancer and other diseases. In particular, we are beginning to define and understand a range of human diseases that are associated with a failure to deliver active perforin to target cells. In this Review, we discuss the current understanding of the structural, cellular and clinical aspects of perforin and granzyme biology.

Cytotoxic T lymphocytes (CTLs) and natural killer cells kill virus-infected and tumor cells through the polarized release of perforin and granzymes. Perforin is a pore-forming toxin that creates a lesion in the plasma membrane of the target cell through which granzymes enter the cytosol and initiate apoptosis. Endosomal sorting complexes required for transport (ESCRT) proteins are involved in the repair of small membrane wounds. We found that ESCRT proteins were precisely recruited in target cells to sites of CTL engagement immediately after perforin release. Inhibition of ESCRT machinery in cancer-derived cells enhanced their susceptibility to CTL-mediated killing. Thus, repair of perforin pores by ESCRT machinery limits granzyme entry into the cytosol, potentially enabling target cells to resist cytolytic attack.

Cytolysin A (ClyA) is a pore-forming toxin that is produced by some bacteria from the Enterobacteriaceae family. This review provides an overview of the current state of knowledge regarding ClyA, including the prevalence of the encoding gene and its transcriptional regulation, the secretion pathway used by the protein, and the mechanism of protein assembly, and highlights potential applications of ClyA in biotechnology. ClyA expression is regulated at the transcriptional level, primarily in response to environmental stressors, and ClyA can exist stably both as a soluble monomer and as an oligomeric membrane complex. At high concentrations, ClyA induces cytolysis, whereas at low concentrations ClyA can affect intracellular signaling. ClyA is secreted in outer membrane vesicles (OMVs), which has important implications for biotechnology applications. For example, the native pore-forming ability of ClyA suggests that it could be used as a component of nanopore-based technologies, such as sequencing platforms. ClyA has also been exploited in vaccine development owing to its ability to present antigens on the OMV surface and provoke a robust immune response. In addition, ClyA alone or OMVs carrying ClyA fusion proteins have been investigated for their potential use as anti-tumor agents.