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. Author manuscript; available in PMC: 2020 Dec 29.
Published in final edited form as: Cell Rep. 2020 Nov 24;33(8):108433. doi: 10.1016/j.celrep.2020.108433

Figure 2. Experimental Validation of IL-27 Predicative Network Identifies TFs that Regulate Il10 In Vitro and In Vivo.

Figure 2.

(A) Log2 fold change of Il10 mRNA levels in WT versus KO Tr1 cells differentiated in vitro with IL-27 for 72 h, quantified by qPCR. Blue, positive regulator; red, negative regulator; gray, not statistically significant. Data are displayed as mean of 2–3 replicates.

(B) Statistically significant ChIP-seq binding sites of ATF-3 ATF-3, Fosl2, T-bet, and IRF8 in the Il10 locus.

(C) Luciferase activity in 293T cells transfected with luciferase reporters for the indicated cis-regulatory elements of Il10 and plasmids encoding the depicted TFs. Firefly luciferase activity is normalized to constitutive Renilla luciferase activity.

(D) WT and Hlx+/− mice were injected intraperitoneally (i.p.) with anti-CD3. Il10 mRNA in CD4+ T cells MACS purified from mesenteric lymph nodes was measured by qPCR.

(E) 5 x 105 in vitro differentiated WT (diamonds) and Hlx+/−(squares) Tr1 cells were transferred i.p. into Rag1−/− recipients. Rag1−/− (circles) did not receive any cells. Changes in body weight were monitored weekly. n = 5.