T-CELL-STIMULATORY FRAGMENTS OF FOOT-AND-MOUTH-DISEASE VIRUS RELEASEDBY MILD TREATMENT WITH CATHEPSIN-D

Cathepsin D and cathepsin B are endosomal/lysosomal proteases that are thought to play a role during in vivo antigen processing, releasing f ragments for binding to major histocompatibility complex class II prod ucts and subsequent presentation to T cells. Here we treated purified foot-and-mouth disease virus (FMDV) strain A(10)Holland with both enzy mes. Cathepsin D, but not cathepsin B, was shown to release fragments from reduced or non-reduced FMDV under mild conditions in vitro. Twent y-eight predominant cathepsin D-released fragments were purified by HP LC and identified by amino acid composition analysis and sequencing. T he unseparated set of fragments produced (the digest) was able to stim ulate T cells from eight vaccinated cattle. With respect to the respon se to intact virus the extent of the response to the digest differed b etween animals: four animals could be classified as good responders, t hree as intermediate responders and one as a low responder. Subsequent ly, we investigated the proliferative T cell response to a large set o f synthetic peptides in detail for two animals, one belonging to the g roup of good responders, the other being the low responder. The peptid es covered all 28 cathepsin D-released fragments analysed and also sev eral sequences not recovered from the digest. In this way seven T cell sites could be identified, five of which coincided with cathepsin D-r eleased fragments. The other two T cell sites were VP2[54-72], being a homologue of a T cell site identified for FMDV strain O1K and the N t erminus of VP4. Whether the most dominantly recognized T cell site was recovered from the digest or not was shown to be related to the good or low response to the digest. These findings suggest a role for cathe psin D in the release of some but not all T cell-stimulatory fragments from FMDV.