Two different pathways contribute to cytotoxicity of mouse cytotoxic T lymphocytes (63.25)

Abstract Cytotoxic T lymphocytes (CTLs) are an important part of the immune system and kill cells which are infected by pathogens like viruses or bacteria (named antigen-presenting-cell or APC). Two different killing mechanisms are being used by CTLs - (1) exocytosis of lytic granules which contain cytotoxic substances and (2) the so called Fas/FasL pathway which relies on agonist/receptor interaction and induces apoptosis in the APC. So far the quantitative contribution of either pathway to the total killing capacity of mouse CTL was not known. To address this question we used a knock out mouse strain (Jinx) which lacks the SNARE associated protein Munc13-4. As a result CTLs from these mice display a severe defect in the secretion of cytotoxic substances, as we could demonstrate by a Granzyme B release assay (reduction of 85% ± 4%) and a FACS based degranulation assay (reduction of 70% ± 9%). Interestingly we found that despite the lytic granule release failure CTLs from Munc13-4 -/- have a remaining killing capacity of about 43%, arguing for a significant contribution of the Fas/FasL pathway. To strengthen this finding we established a single cell assay in which the increase of intracellular Ca2+ in the APC was used as a measure of apoptosis and found that about 50% of killing was induced by the Fas/FasL pathway alone. We conclude from these data that both killing mechanisms are equally important and function synergistically to maintain the cytoxic activity of a mouse CTL.