Extended Data Fig. 8: Identification of Runx2, Runx3 and Gata3 as repressors of TFH genes, acting downstream of Bcl-6.

Related to Fig. 7 a, Schematic of the RV⁺ SMARTA cell transfer system used for LCMV_Arm infection. WT SMARTA CD4 T cells transduced with pMIG (GFP-RV⁺), pMIG-Runx3myc (Runx3-RV⁺), or pMIG-Runx2myc (Runx2-RV⁺) were transferred to C57BL/6 host mice, followed by infection of the host mice with LCMV_Arm, and analyzed 7 days after infection. Related to Fig. 7i-j and Extended Data Fig. 7o-u. b, Total GFP⁺ RV⁺ SMARTA cells [RV⁺ high] were FACS sorted from in vitro culture. Whole cell lysates were analyzed by immunoblotting using anti-myc tag and anti-GAPDH antibodies. Relative Runx expressions are indicated. c, SMARTA CD4 T cells transduced with pMIG (GFP-RV⁺), pMIG-Runx3myc (Runx3-RV⁺ [High]), or pMIG-Runx2myc (Runx2-RV⁺ [High]) were transferred to C57BL/6 host mice, followed by infection of the host mice with LCMV_Arm, and analyzed 7 days later. Frequency of CD45.1⁺ SMARTA cells among total CD4 T cells from spleen of LCMV_Arm infected mice. Two independent experiments were performed; each dot represents one mouse (n = 4). Data are mean ± s.d., unpaired two-tailed Student’s t-test. d, Representative flow cytometry of T_FH RV⁺ [High] SMARTA cell subsets from spleen of LCMV_Arm infected mice. Quantification of results from left panels. e, Bottom 10% [Low], bottom 10-30% [Med] and remaining 30-100% [High] GFP⁺ Runx3-RV⁺ SMARTA cells, and total GFP⁺ RV⁺ SMARTA cells were FACS sorted from in vitro culture. Whole cell lysates were analyzed by immunoblotting using anti-Runx3 and anti-GAPDH antibodies. Relative Runx expressions are indicated. f, Bottom 10% GFP⁺ Runx3-RV⁺ (Runx3-RV⁺ [Low]), bottom 30% GFP⁺ Runx2-RV⁺ (Runx2-RV⁺ [Med]), and total GFP⁺ RV⁺ (GFP-RV⁺) SMARTA cells were FACS sorted from in vitro culture. Whole cell lysates were analyzed by immunoblotting using anti-myc tag and anti-GAPDH antibodies. Relative Runx expressions are indicated. g, SMARTA CD4 T cells transduced with pMIG (GFP-RV⁺), pMIG-Runx3myc (Runx3-RV⁺ [Low]), or pMIG-Runx2myc (Runx2-RV⁺ [Med]) were transferred to C57BL/6 host mice, followed by infection of the host mice with LCMV_Arm, and analyzed 7 days later. Frequency of CD45.1⁺ SMARTA cells among total CD4 T cells from spleen of LCMV_Arm infected mice. Two independent experiments were performed; each dot represents one mouse (n = 5). Data are mean ± s.d., unpaired two-tailed Student’s t-test. h, Representative flow cytometry of T_FH Runx3-RV⁺ [Low] and Runx2-RV⁺ [Med] SMARTA cell subsets from spleen of LCMV_Arm infected mice. Quantification of results from left panels. h, Representative flow cytometry of T_FH Runx3-RV⁺ [Low] and Runx2-RV⁺ [Med] SMARTA cell subsets from spleen of LCMV_Arm infected mice. Quantification of results from left panels.