The protozoan parasite Theileria parva infects and transforms T and B lymphocytes and causes acute, often fatal disease. Cattle can be immunised against the parasite employing an infection and treatment protocol. Infected lymphoblasts express high levels of class I and class II MHC proteins on their surface and immune cattle have been shown to mount strong CD4 and CD8 T cell responses directed against the parasitised cells. Parasite-specific CD8 T cells have been shown to be key mediators of immunity, but the role played by CD4 T cell responses remains unclear. As a first step towards addressing this question, we have undertaken experiments to identify T. parva antigens recognised by specific CD4 T cells and to investigate antigen processing within parasitized cells. CD4 T cell lines generated in vitro from T. parva-immune cattle were used to screen a selected panel of parasite gene products, by testing for recognition of either antigen-presenting cells pulsed with synthetic peptides or cells transfected with parasite cDNAs. Six antigens and epitope regions within them were identified, two of which had previously also been identified as CD8 T cell targets. CD4 T cells from animals of different MHC class II types tended to recognise different parasite proteins and certain epitope specificities were found to be highly dominant. Antigen processing by parasitized cells for recognition by CD4 and CD8 T cells was investigated by testing compounds that inhibit endosomal cathepsins, autophagy, proteasome function or endosomal acidification. Recognition by both T cell subsets was substantially inhibited by a proteasome inhibitor. Cathepsin inhibitors had only a modest inhibitory effect on the CD4 T cells, in contrast to their strong inhibition of ovalbumin-specific CD4 T cells. These results indicate that processing of T. parva antigens recognised CD4 T cells involves a proteasome-dependent pathway.