Structure and mechanism of bactericidal mammalian perforin-2, an ancient agent of innate immunity

Abstract

Perforin-2 (MPEG1) is thought to enable the killing of invading microbes engulfed by macrophages and other phagocytes, forming pores in their membranes. Loss of perforin-2 renders individual phagocytes and whole organisms significantly more susceptible to bacterial pathogens. Here, we reveal the mechanism of perforin-2 activation and activity using atomic structures of pre-pore and pore assemblies, high-speed atomic force microscopy, and functional assays. Perforin-2 forms a pre-pore assembly in which its pore-forming domain points in the opposite direction to its membrane-targeting domain. Acidification then triggers pore formation, via a 180° conformational change. This novel and unexpected mechanism prevents premature bactericidal attack and may have played a key role in the evolution of all perforin family proteins.

Molecular and Functional Analysis of Pore-Forming Toxin Monalysin From Entomopathogenic Bacterium Pseudomonas entomophila

“…Revealing the structure of native Monalysin in solution and lipid membrane, particularly its dynamic nature, is essential to understand its detailed molecular function and use to evaluate innate immunity mechanisms in flies, as well as to develop biological control agents against insects and biological nanopores. To this end, we employed HS-AFM that enabled dynamic real-time observations of macromolecules at nanometer resolutions, which are not feasible with other methods (42,43), and had recent achievements of revealing the dynamic structures of pore-forming proteins (44)(45)(46)(47).…”

MPEG1/Perforin-2 Haploinsufficiency Associated Polymicrobial Skin Infections and Considerations for Interferon-γ Therapy

“…Given the known bactericidal function of perforin-2 and the location of this variant in the P2 domain and eliminating the Cterminal transmembrane domain, the MPEG1 variant was suspected to be pathogenic ( Figure 1B). If the Tyr430* truncated protein product were stably expressed it would likely be secreted rather than delivered to endophagosomes as its transmembrane domain is essential for the intracellular retention of perforin-2 (6,8). Immunologic assays demonstrated normal neutrophil chemotaxis and extracellular bactericidal activity against Staphylococcus aureus ( Table 2).…”

Perforins Expression by Cutaneous Gamma Delta T Cells

“…The Perforin-2 cytosolic tail directs the endosomes to bacteria-encapsulating phagosomes (46). Acidification of the phagosome stimulates reconfiguration of the MACPF domain, resulting in pore formation on the bacterial cell membrane (46,47). Our group was the first to demonstrate the essential role of Perforin-2 in eliminating intracellular bacterial infections (48,49), confirming the importance of this protein as an antimicrobial effector protein expressed by both phagocytic and tissue forming cells.…”

“…Perforin-2 differs from other MACPF pore formers in that it has a transmembrane domain and localizes to endosomal membranes. The Perforin-2 cytosolic tail directs the endosomes to bacteria-encapsulating phagosomes (46). Acidification of the phagosome stimulates reconfiguration of the MACPF domain, resulting in pore formation on the bacterial cell membrane (46,47).…”

“…Location in the cell Intracellular cytolytic granules (39) Membranes of intracellular endosomes (46,47) Function in GD T cells Lysis of transformed or infected cells (39)(40)(41) Destruction of intracellular bacteria (46,47) Stimulatory factors Surface receptors: -GD TCR 53 suffering from these illnesses. Compounds identified as Perforin inhibitors in NK and AB T cells include diarylthiophenes and benzenesulfonamide based therapeutics (83)(84)(85).…”