Granzyme B is recovered by natural killer cells via clathrin-dependent endocytosis

Abstract

When they recognize a target cell, natural killer (NK) cells mount an attack to kill the target by exerting their cytotoxicity via the exocytosis of cytotoxic granules. Although the details of this process (which includes the movement of cytotoxic granules in the immune synapse and their fusion with the plasma membrane, releasing granzymes and perforin into the synaptic cleft) are relatively better understood, the post-exocytosis regulation of the process is still largely unknown. Here we show that a clathrin-dependent endocytosis stimulated by target cell occurs in NK92 cell line, which is closely correlated with granzyme B recovery. Inhibition of the endocytosis significantly attenuates the cytotoxicity of NK92 cells. The NK cell recovery of its released effector molecules, in turn, suggests that endocytosis may well play a key role in the post exocytosis regulation of immune cells.

Classification of human natural killer cells based on migration behavior and cytotoxic response

“…17 However, NK cells could have the potential to kill many more if recycling cytotoxic molecules. 19,20 Eventual depletion of perforin in granules is thought to contribute to "exhaustion" of the NK cells' killing capacity.…”

Surface CD107a/LAMP-1 protects natural killer cells from degranulation-associated damage

“…11 However, once secreted into the synaptic cleft, where Ca 21 concentrations are high and the pH is neutral, other mechanisms are required to facilitate unidirectional killing and to prevent cytotoxic lymphocyte self-destruction. To this end, cytotoxic lymphocytes endocytose granzyme B, reducing the concentration of active granzyme B 17 and express the cytoplasmic serine protease inhibitor serpinB9, which can inactivate granzyme B that has leaked into the cytoplasm. 18,19 Moreover, surface cathepsin B has been suggested to cleave perforin at the cytotoxic cell's membrane, 20 but cytotoxic T lymphocytes (CTLs) from cathepsin B-deficient mice survive normally after target cell encounter.…”

Perforin is recaptured by natural killer cells following target cells stimulation for cytotoxicity

“…Western blotting was done as previously described (Li et al, 2010). In brief, 2 × 10 6 of stimulated NK92 cells were harvested, from which 80 μg of protein was separated by SDS/10% PAGE and transferred to nitrocellulose membranes.…”

“…In our previous study, granzyme B appeared to be recovered after target cell stimulation. This recovery contributes to the cytotoxicity of NK cells (Li et al, 2010). Granzyme B‐induced apoptosis of a target cell is entirely dependent on Pfn (Stepp et al, 1999; Marcenaro et al, 2006; Chowdhury and Lieberman, 2008).…”