Mechanisms regulating the CD8+ T cell response to Mycobacterium tuberculosis

Abstract

CD8+ T cells that recognize peptide antigens presented by major histocompatibility complex (MHC) class I molecules are an important component of the cellular immune response to mycobacterial pathogens, including Mycobacterium tuberculosis. Following uptake by macrophages, mycobacterial antigens are processed for presentation on MHC class I molecules through a process that is referred to generally as "cross-presentation". There are multiple proposed mechanisms for the cross-presentation of mycobacterial antigens, and published studies have found apparently conflicting evidence with regard to which mechanisms are used by macrophages infected with M. tuberculosis. Using recombinant strains of M. smegmatis engineered to express different M. tuberculosis antigens tagged with an MHC class I binding peptide sequence from ovalbumin (ova), we show that the mechanism of cross presentation by macrophages can vary depending on which particular mycobacterial antigen is studied. Fusion proteins in which the ova epitope was appended to the C-termini of either M. tuberculosis Antigen 85B or 19-kDa lipoprotein were processed in the phagolysosomes of macrophages by a mechanism requiring the cysteine protease cathepsin-S, and loaded onto MHC class I molecules that recycled to endosomes from the cell surface. In contrast, the presentation of a fusion protein in which the ova epitope was appended to ESAT-6 was not dependent on cathepsin-S expression or MHC class I recycling. Instead, this required the transporter associated with antigen processing (TAP) complex, which is a key component of the classical processing pathway by which newly synthesized MHC class I molecules acquire peptide antigens in the endoplasmic reticulum. Whereas these in vitro studies showed that at least two separate pathways are used in macrophages for MHC class I presentation of mycobacterial antigens, we found that efficient CD8+ T cell priming in vivo in mice was dependent on TAP for all antigens tested.;A property of M. tuberculosis that has been linked to evasion of host CD8+ T cell immunity is the ability of these bacilli to prevent the death by apoptosis that is typically triggered upon infection of phagocytic cells by vacuolar bacterial pathogens. Infection of macrophages with nonpathogenic strains of mycobacteria, including M. smegmatis and the avirulent strain of M. tuberculosis , H37Ra, leads to the rapid induction of apoptosis. In contrast, pathogenic mycobacteria including M. tuberculosis have evolved mechanisms to inhibit this response. Little is known about the molecular mechanisms underlying this inhibition or its consequences for host immunity. We demonstrated that inactivation of the virulence-associated secA2 secretion apparatus of M. tuberculosis caused marked enhancement of apoptosis of infected macrophages due to loss of secretion of mycobacterial superoxide dismutase. Deletion of secA2 was associated with markedly increased priming of antigen-specific CD8+ T cells, and augmented effector and memory CD8+ T cell responses in vivo. Compared to standard vaccination with attenuated M. bovis BCG, vaccination with DeltasecA2 generated significantly enhanced protective immunity against M. tuberculosis challenge in mouse and guinea pig models. Our results define a mechanism for a key immune evasion strategy of M. tuberculosis, and provide a novel strategy for improving live attenuated mycobacterial vaccines.