The I-Ab-restricted alloresponse of D10.G4.1 T cells is based on the recognition of an endogenous peptide.

The alloreactivity of T cells is thought to be based on the cross-reactive recognition of allogeneic major histocompatibility complex (MHC) molecules which have bound peptides derived from self antigens or, in the case of cultured T cells, from serum components. While studying the processing requirements of conalbumin (CA) that is recognized by D10.G4.1 T cells in combination with I-Ak molecules we also analysed the cross-reactive stimulation of clone D10.G4.1 T cells by allogeneic, I-Ab expressing stimulator cells which is shown here to be CD4 dependent. In order to distinguish between an endogenous or exogenous origin of a peptide that is presumably co-recognized with I-Ab different types of stimulatory/antigen-presenting cells (APC) were treated with drugs that are known to influence the processing and/or presentation of antigens. It was found that the alloreactive response of D10.G4.1 cells was abolished if the APC were treated with brefeldin A or inhibitors of protein biosynthesis. Under the same conditions neither the CA-specific response of D10.G4.1 cells nor the activation of control T cells by ovalbumin (OVA) or insulin was affected. On the other hand, the use of lysosomotropic agents or inhibitors of glycoprotein trimming had no influence on the ability of the APC to induce the alloresponse of D10.G4.1 cells, whilst the presentation of CA and other protein antigens by the APC was prevented. In addition, treatment of APC with pronase to remove surface MHC molecules or acidic buffer to remove peptides from the binding groove of MHC class II molecules at the surface of APC strongly diminished their ability to induce an alloresponse. However, this capacity was restored by incubating the APC for 2 hr in serum-free medium. These data indicate that the alloreactive response of D10.G4.1 cells is based on the recognition of newly synthesized endogenous peptide(s) in combination with I-Ab.