USP18 expression in tumor modulates immune cell population and phenotype. C57/BL6 mice received intravenous inoculation of 3 × 105 B16-OVA-USP18 or B16-OVA-GFP tumor cells. 2 weeks later, T cell, CD11b+ myeloid cell and NK cell infiltration in the lung was analyzed by flow cytometry (A). Antigen-specific OT-1 tetramer-positive CD8+ T cells in the lung draining lymph nodes (DLN), lung, and spleen were analyzed (B). PD-1 expression on CD8+ T cells (C) was analyzed. NKG2D expression on NK cells in tumor microenvironment was monitored by flow cytometry (D). ISG15 levels in B16-OVA-GFP, B16-OVA-USP18 or B16-OVA-shUSP18 tumor lysate was analyzed by ELISA (E). CD11c+ dendritic cells were isolated from B16-OVA-GFP, or B16-OVA-USP18, and B16-OVA-shUSP18 lung melanoma and cultured in vitro with or without LPS. Dendritic cell maturation and cytokine were assayed by flow cytometry and ELISA, respectively (F-G).