Cystatin F as a regulator of immune cell cytotoxicity

Abstract

Cysteine cathepsins are lysosomal peptidases involved in the regulation of innate and adaptive immune responses. Among the diverse processes, regulation of granule-dependent cytotoxicity of cytotoxic T-lymphocytes (CTLs) and natural killer (NK) cells during cancer progression has recently gained significant attention. The function of cysteine cathepsins is regulated by endogenous cysteine protease inhibitors-cystatins. Whereas other cystatins are generally cytosolic or extracellular proteins, cystatin F is present in endosomes and lysosomes and is thus able to regulate the activity of its target directly. It is delivered to endosomal/lysosomal vesicles as an inactive, disulphide-linked dimer. Proteolytic cleavage of its N-terminal part leads to the monomer, the only form that is a potent inhibitor of cathepsins C, H and L, involved in the activation of granzymes and perforin. In NK cells and CTLs the levels of active cathepsin C and of granzyme B are dependent on the concentration of monomeric, active cystatin F. In tumour microenvironment, inactive dimeric cystatin F can be secreted from tumour cells or immune cells and further taken up by the cytotoxic cells. Subsequent monomerization and inhibition of cysteine cathepsins within the endosomal/lysosomal vesicles impairs granzyme and perforin activation, and provokes cell anergy. Further, the glycosylation pattern has been shown to be important in controlling secretion of cystatin F from target cells, as well as internalization by cytotoxic cells and trafficking to endosomal/lysosomal vesicles. Cystatin F is therefore an important mediator used by bystander cells to reduce NK and T-cell cytotoxicity.

Keywords: CITIM 2017; Cathepsins; Cell cytotoxicity; Cystatin F; Natural killer cells; T cells.

Fig. 1

Co-localization/interaction of cystatin F and cathepsin C in TALL-104 cells determined by proximity ligation assay. Blue: DAPI stained nuclei; red: proximity ligation assay signals. The method was carried out as described in Soderberg and collaborators [57]. Images were taken on a Carl Zeiss LSM 710 confocal microscope. DAPI 4′,6′-Diamidino-2-phenylindole, LSM laser scanning microscope

Fig. 2

Increasing extracellular concentration of cystatin F negatively affects the granule-mediated cytotoxicity of NK cells and CTLs. Cystatin F originating from macrophages, mast cells, dendritic cells and/or tumour cells can be taken up by NK cells and CTLs and transported to secretory lysosomes where it is activated. In secretory lysosomes, cystatin F inhibits the cathepsins C and H needed for proteolytic activation of granzymes—the major cytotoxic mediators of NK cells and CTLs