gms | German Medical Science

Objective: One important hallmark of cancer cells is the resistance to apoptosis. Survivin is a structurally and functional unique member of the inhibitor of apoptosis protein (IAP) family that acts at the interface between apoptosis and cell cycle control. In many human cancers aberrant expression of survivin correlates with poor prognosis and resistance to chemotherapy. Moreover, one of the main impacts of Survivin is the inhibition of caspases. Therefore, we analysed the inhibitory potential of the three Survivin splice variants, i.e. Survivin-DEx3, Survivin-2B, and Survivin-3B in vitro.

Methods: Transfection of HEK293 cells (with GFP-Survivin,GFP-Survivin-ΔEx3, GFP-Survivin-2B and GFP-Survivin-3B), Western Blotting, Caspase-Assays with purified recombinant Survivin, XIAP, and Caspase-3 and –9, and cell extracts of transfected HEK293 cells.

Results: 1. Survivin inhibits recombinant Caspase-3 as well as Caspase-9 in a concentration dependent manner. 2. XIAP enhances the ability of Survivin to inhibit Caspase-3 activity.3. In contrast, Survivin does not increase the potential of XIAP to suppress Caspase-9 activity.4. After transfection with GFP-tagged Survivin, HEK293 cells extracts inhibit recombinant Caspase-9 but not recombinant Caspase-3. 5. HEK293 cell extracts transfected with GFP-Survivin-ΔEx3, GFP-Survivin-2B and GFP-Survivin-3B fail to inhibit both Caspase-3 and -9 activity.

Conclusions: IAPs contribute to the lack of ability of many tumors to undergo apoptosis. Our findings show that – in contrast to Survivin and XIAP – the mainly carboxy-terminal modified splice variants Survivin-ΔEx3, Survivin-2B, and Survivin-3B fail to inhibit caspases-3 and -9 activity in vitro. Therefore, there is a striking evidence for the carboxyterminal region of Survivin to be involved in caspase inhibition.