Functional role of granzymes in epidermal γδ T cell-mediated killing of tumor targets

Granzymes, a family of serine proteases contained in cytoplasmic granules, play a critical role in killing tumor targets, by triggering the rapid breakdown of DNA and subsequent apoptosis. Dendritic epidermal T cells (DETC), skin-specific members of the tissue-type γδ T cell family in mice, have been shown to kill selected tumor cell lines. Our purpose was, therefore, to elucidate the functional role of granzymes in DETC-mediated killing. The short-term DETC line (C25) established from CBA mice: a) lysed YAC-1 lymphoma targets effectively (51Cr release assay), b) contained significant BLT esterase activity (colorimetric enzyme analysis), c) expressed constitutively mRNA for granzymes A and B (Northern blotting), and d) produced relatively large amounts of granzyme A (immunoblotting). mRNA expression for granzyme B was confirmed by RT-PCR analysis of Thy-l+ epidermal cells (i.e., DETC) that were freshly isolated from CBA mice. C25 DETC also induced apoptosis of the YAC-1 targets as measured by their capacity to cause breakdown of 3H-thymidine-labeled DNA. Importantly, this ability was blocked completely by preincubation of DETC with the granzyme inhibitor, dichloroisocoumarin (DCI), but not with the cysteine protease inhibitor, E-64. Thus, we conclude that granzymes play an essential role in DETC-mediated apoptosis of tumor targets.