Supplementary Figure 2: Expression of miR-31 after treatment of CD8⁺ T cells with cytokines or TLR agonists.

(a) Naïve OT-I CD8⁺ T cells were treated for 24 hours with 1 μg/ml SIINFEKL peptide and 20 ng/ml IL-2, or 20 ng/ml of the indicated cytokines for 48 hours. MiR-31 expression was assayed by qPCR relative to input naïve OT-I. (b) Naïve OT-I T cells were treated for 48 hours with CD3+CD28 antibodies or 20 ng/ml TGF-b for 48 hours. MiR-31 expression was assayed by qPCR relative to input naïve OT-I. (c) CD8⁺ T cells were treated for 24 hours with PMA/ionomycin, 10 μg/ml LPS, or 3 μg/ml CpG 2006. MiR-31 expression was assayed by qPCR relative to input CD8⁺ T cells. (d) miR-31 expression relative to the small nucleolar RNA sno234 was measured by qPCR 24 hours following activation of CD4⁺ or CD8⁺ T cells (CD3+CD28 antibodies), NK cells (PMA/ionomycin) and B cells (PMA/ionomycin), compared to cells without activation stimulus. (e) miR-31 expression relative to sno-234 in OT-I CD8⁺ T cells activated by SIINFEKL peptide-pulsed splenocytes for 72 hours, and then cultured in the presence of IL-2 or IL-15 for 8 or 15 days. (f) miR-31 expression relative to sno234 in ex vivo sorted CD4⁺ and CD8⁺ T cell subsets: naïve (CD44^low CD62L⁺), effector memory (EM, CD44^hi CD62L^-), central memory (CM, CD44^hi CD62L⁺), Treg (CD4⁺ CD25⁺ GITR⁺) subsets. Data are representative of two (a-e) experiments. Data show mean ± s.d.