Toll-like receptor-3 (TLR3), a member of an immune recognition receptor family, is widely expressed in tumour cells and has been shown previously to have the capacity to not only activate immune signalling pathways, but also to exert proapoptotic activity in some cells. We show here that HaCaT human keratinocytes are susceptible to apoptosis induction by the TLR3
ligand poly I:C, and use these cells as a model to analyse the apoptotic signalling pathway. Although the BH3-only
protein Noxa was transcriptionally induced by
poly I:C and translocated to mitochondria, RNAi experiments showed that the BH3-only
proteins Noxa, Bim and Puma were individually dispensable for
poly I:C-induced apoptosis. Instead,
poly I:C-induced activation of
caspase-8 via TLR3 and its adapter TRIF was required for apoptosis. In human
melanoma cell lines
poly I:C failed to induce apoptosis unless
protein synthesis was blocked. Significantly, sensitisation towards
poly I:C-dependent
caspase-8 activation and apoptosis in
melanoma cells was also achieved by the synthetic Smac mimetic/
inhibitor of apoptosis protein (IAP) antagonist,
LBW242, or by specific downregulation of cIAP1 by
siRNA. Inactivation of
caspase-8 by CrmA overexpression reduced
poly I:C/
LBW242-induced apoptosis. These results indicate that the proapoptotic activity of TLR3/TRIF/
caspase-8 in
melanoma cells is under the control of IAPs, and the use of novel Smac mimetics might be a feasible approach to target
melanoma.