Material Supplementary 8.DC1http://www.jimmunol.org/content/suppl/2010/02/15/jimmunol.090270

Nucleoprotein (N) is an immunodominant antigen in many enveloped virus infections. While the diagnostic value of anti‐N antibodies is clear, their role in immunity is not. This is because while they are non‐neutralising, they somehow clear infection by coronavirus, influenza and LCMV in vivo. Here, we show that anti‐N immune protection is mediated by the cytosolic Fc receptor and E3 ubiquitin ligase TRIM21. Exploiting LCMV as a model system, we demonstrate that TRIM21 uses anti‐N antibodies to target N for cytosolic degradation and generate cytotoxic T cells (CTLs) against N peptide. These CTLs rapidly eliminate N‐peptide‐displaying cells and drive efficient viral clearance. These results reveal a new mechanism of immune synergy between antibodies and T cells and highlights N as an important vaccine target.

In response to primary Ag contact, naive mouse CD8+ T cells undergo clonal expansion and differentiate into effector T cells. After pathogen clearance, most effector T cells die, and only a small number of memory T cell precursors (TMPs) survive to form a pool of long-lived memory T cells (TMs). Although high- and low-affinity CD8+ T cell clones are recruited into the primary response, the TM pool consists mainly of high-affinity clones. It remains unclear whether the more efficient expansion of high-affinity clones and/or cell-intrinsic processes exclude low-affinity T cells from the TM pool. In this article, we show that the lack of IFN-γR signaling in CD8+ T cells promotes TM formation in response to weak, but not strong, TCR agonists. The IFN-γ–sensitive accumulation of TMs correlates with reduced mammalian target of rapamycin activation and the accumulation of long-lived CD62LhiBcl-2hiEomeshi TMPs. Reconstitution of mammalian target of rapamycin or IFN-γR signaling is sufficient to block this process. Hence, our data suggest that IFN-γR signaling actively blocks the formation of TMPs responding to weak TCR agonists, thereby promoting the accumulation of high-affinity T cells finally dominating the TM pool.

T cells sense and respond to their local environment at the nanoscale by forming small actin-rich protrusions, called microvilli, which play critical roles in signaling and antigen recognition, particularly at the interface with the antigen presenting cells. However, the mechanism by which microvilli contribute to cell signaling and activation is largely unknown. Here, we present a tunable engineered system that promotes microvilli formation and T cell signaling via physical stimuli. We discovered that nanoporous surfaces favored microvilli formation and markedly altered gene expression in T cells and promoted their activation. Mechanistically, confinement of microvilli inside of nanopores leads to size-dependent sorting of membrane-anchored proteins, specifically segregating CD45 phosphatases and T cell receptors (TCR) from the tip of the protrusions when microvilli are confined in 200-nm pores but not in 400-nm pores. Consequently, formation of TCR nanoclustered hotspots within 200-nm pores allows sustained and augmented signaling that prompts T cell activation even in the absence of TCR agonists. The synergistic combination of mechanical and biochemical signals on porous surfaces presents a straightforward strategy to investigate the role of microvilli in T cell signaling as well as to boost T cell activation and expansion for application in the growing field of adoptive immunotherapy.