The role of serpinB9 in the cellular immune response

Cytotoxic lymphocytes, comprising CD8+ T cells and NK cells, are vital for the elimination of virally infected or abnormal cells. One mechanism utilized by CLs to initiate death in target cells is the intracellular delivery of granzymes; a family of serine proteases stored in lysosome related organelles (LRO) in the CLs. Of the granzyme family, granzyme B (grB) is the most cytotoxic. CLs and dendritic cells (DCs) express PI-9, an intracellular inhibitor of grB. PI-9 is suggested to protect these cells from grB, which may access the cytosol via either cell-cell contact by degranulation or by translocation from the LRO to the cytosol of these cells. To better understand the expression and role of PI-9 in the immune system, mice deficient in the murine orthologue of PI-9, SPI6, were generated. SPI6 was found to have a similar expression pattern to PI-9, and is most highly expressed in effector CD8+ T cells, NK cells and DCs. Infection of SPI6 KO mice with the virulent Moscow strain of the mouse pathogen Ectromelia virus (ECTV) results in a grB-dependent decrease in the proportion of effector CD8+ T cells and NK cells, leading to increased mortality induced by ECTV. This demonstrates that SPI6 is highly important in preventing excessive or premature grB mediated death of CLs. GrB has been demonstrated previously to play a role in limiting the viability of CLs, an important process in preventing immune related pathology. GrB mediated death of CLs is potentially facilitated by a novel mechanism of serpin regulation described in this thesis, in which reactive oxygen species (ROS) generated by CLs during activation inactivate PI-9/SPI6. PI-9/SPI6 function is reversibly inhibited by oxidants, preventing it forming a complex with, and inactivating, grB. This is due to a vicinal disulfide bond formed between two conserved cysteine residues in the reactive centre loop. This suggests that modulation of both cytosolic grB and functional PI-9 may reflect a mechanism that limits the lifespan of CLs.