Supplementary Figure 4: Ectopic expression of GBPs in Ifnar1–/– cells and efficiency of Gbp2 and Gbp5 knockdown. | Nature Immunology

Supplementary Figure 4: Ectopic expression of GBPs in Ifnar1–/– cells and efficiency of Gbp2 and Gbp5 knockdown.

From: Guanylate-binding proteins promote activation of the AIM2 inflammasome during infection with Francisella novicida

Supplementary Figure 4

(a–c) Ectopic expression of GBP2 or GBP5 do not complement type-I-IFN receptor deficiency. Ifnar1–/– macrophages were transduced with lentivirus encoding either GFP only (EV = empty vector) and GFP-GBP2 and GFP-GBP5. 48 h post transduction, macrophages were infected with F. novicida at the indicated MOI. IL-1β concentration in the supernatant was determined at 7 h and 10.5 h post infection (a). Specific ectopic expression was verified by quantifying the Gbp3 (control), Gbp2 and Gbp5 transcript levels. Results are expressed as fold induction relative to the transcript level in Ifnar1–/– macrophages transduced with empty vector control (b). Graphs show mean and s.d. of triplicate assays. The percentage of transduced cells was determined by flow cytometry based on GFP expression (c). (d) RT-PCR for Gbp2 and Gbp5 expression from unprimed wild-type, Gbp2–/– and Gbp5–/– BMDMs treated with Non-Targeting or the indicated gene-specific siRNA for 22 h and infected for 8 h with wild-type F. novicida U112. Graphs show mean and s.d. of quadruplicate assays and data are representative of independent two experiments. *, p<0.05; **, p<0.01; NS, not significant (two-tailed unpaired t-test). ND, not detected.

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