IGR_DNA_DAMAGE_STUDY_FM

Granzyme B plays a key role in cell-mediated programmed cell death. We previously demonstrated that p53 is a functional determinant in the Granzyme B-induced cytotoxic T lymphocyte response. However, the pathways leading to activation of p53 by Granzyme B remain incompletely understood. We now demonstrate that Granzyme B induced DNA damage signaling as revealed by histone H2AX phosphorylation and subsequent activation of the stress kinase CHK2. Confocal microscopy analysis indicates that Granzyme B treatment of tumor cells induced an early translocation of endonuclease caspase-activated DNase. DNA microarray based global transcriptional profiling and RT-PCR indeed revealed genes related to DNA damage. Among these genes, hSMG-1, a genotoxic stress-activated protein, was constantly upregulated in tumor cells following Granzyme B treatment. Knockdown of the hSMG-1 gene in T1 tumor target cell line resulted in a significant inhibition of Granzyme B- and CTL-induced killing. Our data suggest that Granzyme B may exert cell death through DNA damage signaling and uncover a novel molecular link between the DNA damage pathway and Granzyme B-induced cell death.