Fig. 2.

DCIR expression leads to sustained type I IFN signaling in DCs. (A) Bone marrow-derived DCs from WT or KO mice were either left uninfected or infected (4 h) with M. tuberculosis H37Rv in the presence or absence of the IFNAR1-blocking monoclonal antibody MAR1-5A3, as indicated. Relative expression of the ISGs Oas2, Irf7, and Isg15 was quantified by RT-qPCR. Red asterisks correspond to comparison analysis between MAR1-5A3–treated and –untreated cells. (B–D) WT or KO DCs were either left uninfected (0 h) or infected for 4 or 18 h with M. tuberculosis. IFNα and IFNβ (B) were quantified in the cell supernatants by ELISA or using the type I IFN-activity reporter cell line B16 Blue IFNα/β (C). The expression of IFNAR1 was quantified by FACS (D). (E–I) WT or KO DCs were left unstimulated (Mock), were infected with M. tuberculosis for 4 h, or were costimulated with the TLR2 agonist lipopeptide Pam₃CSK₄ and IFNβ for 4 h, as indicated. In I, cells were left untreated or were treated with the SHP2 inhibitor GS-493 (24). JAK1 (E), STAT1 (F and I), SHP-2 (G and H), and their phosphorylated forms were quantified in cell lysates by Western blot (WB) (G) or ELISA (E, F, H, and I). G, Right presents quantification of three independent WB analyses (G, Left) by densitometry. In A–I, data are presented as means ± SEM of at least three biological replicates and are representative of at least two independent experiments. Statistical analysis was performed using Student’s t test. *P < 0.05, **P < 0.01.