Characterization of the role of caspase-11 in regulating actin dynamics and cell migration

Caspase-11 is a member of mammalian caspase family of cysteine proteases. Previous studies showed that under inflammation conditions (e.g. LPS stimulation), caspase-11 activates caspase-1, which is the enzyme that cleaves pro-interleukin-1[beta] (pro-IL-1[beta]) into mature IL-1[beta]. Caspase-11 can also directly process caspase-3 to induce apoptosis. In this thesis work, I searched for proteins that interact with caspase-11 and I identified two actin associated proteins, Aip1 ( a ctin i nteracting p rotein 1) and Flightless-I. Aip1 is know to regulate cofilin-mediated actin depolymerization. My study shows that caspase-11 interacts physically and functionally with Aip1. Binding domains between these two proteins are mapped to the CARD domain of caspase-11 and the C-terminal WD40 propeller of Aip1. Inhibition of Aip1 or caspase-11 expression reduces mammalian cell motility. Actin dynamics is altered in caspase-11-/- cells. Recombinant caspase-11 promotes cofilin- and Aip1-dependent actin depolymerization in vitro. These results reveal a novel function of caspase-11 in regulating actin dynamics and cell migration and suggest caspase-11 might be able to regulate inflammatory response at multiple levels. The second protein, Flightless-I, has a N-terminal leucine-rich repeat (LRR) region and a C-terminal gelsolin-like domain (GLD). The GLD is known for actin binding. Different from the interaction between caspase-11 and Aip1, which is mediated through the CARD domain of caspase-11, Flightless-I interacts with the p30 enzymatic domain of caspase-11. The enzymatic activity domain of caspase-11 can bind to both LRR and GLD of Flightless-I. Co-localization of caspase-11 and Flightless-I is found at the cell leading edge in caspase-11 and Flightless-I co-expressing cells. Overexpression of Flightless-I but not Aip1 makes more caspase-11 localize to the highly cross-linked actin filament fraction. These results suggest Flightless-I, as an actin binding protein, is able to modify the localization of caspase-11. Caspase-1 can also interact with Flightless-I. Overexpression of Flightless-I inhibits the enzymatic activity of caspase-1 in both self-processing and processing pro-IL-1[beta] into mature IL-1[beta]. The inhibitory property of Flightless-I is preserved in both LRR and GLD domain alone, although less efficient than the full-length Flightless-I, suggesting Flightless-I might work like an inhibitory clamp on caspase-1 or caspase-11. This thesis work expands our current understanding of the function and regulation of mammalian caspases under physiological conditions.