Cathepsins and their natural inhibitors in CTL and NK cell cytotoxicity

Cystatins (Cys) C and F are potent inhibitors of several cysteine proteases, but their physiological role is still unknown. While Cys C is ubiquitously expressed, the expression of Cys F is highly restricted. Cys F is expressed in dendritic cells, where is upregulated upon activation, and also in natural killer (NK) and CD8+ T cells. Although Cys C has been postulated to be the natural inhibitor of cathepsin (Cat) S for the control of invariant chain degradation in mouse dendritic cells, the distribution and localization of Cys F (in lysosomes/endosomes) points at Cys F as a better candidate. Furthermore, Cys F can also inhibit asparagine endopeptidase, a key molecule in the degradation of myelin basic protein in antigen presenting cells. The expression of Cys F in cells with cytotoxic properties matches the presence of the lysosomal protease Cat W, for which no function has been yet described.
We plan to knock off the expression of these cysteine protease natural inhibitors by introduction of small interference RNA (siRNA). We will conduct functional analysis in dendritic cells, where we will study their effects in protease activity (active site labelling), protein digestion, and also in cytotoxic CD8 and NK cell lines, where we will analyze their cytotoxic capacity. In summary, we should be able to dissect which proteases are sensitive to these inhibitors, and how regulation of the inhibitors affects cell function.

Contact: C.Stöckle, E. Tolosa