The human anti-apoptotic proteins cIAP1 and cIAP2 bind but do not inhibit caspases

Area of Science:

• Cellular biology
• Molecular biology
• Biochemistry

Background:

• Cellular inhibitor of apoptosis proteins (cIAPs) are known to regulate apoptosis.
• Previous research suggested cIAPs inhibit apoptosis through direct caspase inhibition via their baculovirus IAP repeat (BIR) domains.
• cIAPs contain three BIR domains, with domains 2 and 3 implicated in potential caspase interactions.

Purpose of the Study:

• To investigate the direct caspase inhibitory function of cIAP BIR domains.
• To determine if cIAP BIR domains 2 and 3 can bind and inhibit caspases-7 and -9.
• To explore the structural basis for caspase binding and inhibition by cIAPs.

Main Methods:

• Expressing and purifying BIR domains 2 and 3 of cIAPs.
• Performing binding assays to assess interactions between cIAP BIR domains and caspases-7 and -9.
• Analyzing the inhibitory activity of cIAP BIR domains against caspases.
• Mutagenesis studies to convert cIAP BIR domains into functional caspase inhibitors by incorporating XIAP residues.

Main Results:

• cIAP BIR domains 2 and 3 were shown to bind caspases-7 and -9.
• Neither cIAP BIR domain 2 nor 3 demonstrated caspase inhibitory activity.
• Critical substitutions in cIAP BIR domains, distinct from those in the X-linked inhibitor of apoptosis protein (XIAP), were identified as the reason for lack of inhibition.
• Conversion of cIAP BIR domains into potent caspase inhibitors was achieved by introducing XIAP-specific residues.

Conclusions:

• cIAPs possess protein scaffolds capable of binding caspases but have lost or never acquired the specific interaction sites for direct caspase inhibition.
• The binding function of cIAP BIR domains is likely crucial for their physiological roles, but direct caspase inhibition is not one of them.
• Structural modifications distinguish cIAPs from potent caspase inhibitors like XIAP, highlighting divergent functional evolution within the IAP family.