α-Galactosylceramide-expanded virtual memory CD8⁺ T cells confer protection against a broad range of pathogens

α-Galactosylceramide (α-GalCer) treatment of animals is a well-established model to investigate the beneficial roles of invariant natural killer T (iNKT) cells in anti-infection and anti-tumor immunity. We previously found that α-GalCer treatment expands not only iNKT cells but also a group of antigen-inexperienced memory-like innate virtual memory CD8⁺ T (CD8⁺ T_VM) cells. With the bystander protective potential of CD8⁺ T_VM cells, their roles in α-GalCer-mediated immunity against infection remain largely unexplored. Here, we reported that α-GalCer treatment conferred protection against systemic, enteric, and pulmonary bacterial infections, as well as pulmonary viral infection. This protection persisted after iNKT cells had diminished and coincided with sustained expansion of CD8⁺ T_VM cells. Adoptive transfer of α-GalCer-expanded CD8⁺ T_VM cells one day before infection significantly decreased bacterial and viral burdens. Mechanistically, α-GalCer treatment activated iNKT cells to produce IL-4, which subsequently expanded the number of CD8⁺ T_VM cells that were capable of producing TNF-α and IFN-γ. But the ability of each individual CD8⁺ T_VM cell to produce these cytokines was not enhanced. Lung-localized CD8⁺ T_VM cells were the primary contributors to α-GalCer-expanded CD8⁺ T_VM cells in the lung. α-GalCer-expanded CD8⁺ T_VM cells can reduce pulmonary viral titer in the host in a TNF-α-dependent manner in the absence of IFN-γ signaling. Together, these findings describe an immune cascade in which α-GalCer-activated iNKT cells orchestrate the expansion and function of CD8⁺ T_VM cells through IL-4 production, reveal previously unrecognized roles of CD8⁺ T_VM cells as downstream effectors in α-GalCer-mediated immunotherapeutic effects, and highlight the protective potential of CD8⁺ T_VM cells in host defense against diverse pathogens, thus providing a novel approach to developing new therapeutics for combating not only various infectious diseases but also tumors.