Fas is the direct target of miR-146a in GC B cells. (A) Fas expression in various lymphocyte subsets of lymph nodes from WT and miR-146a TG mice, determined by FACS (gated as CD3⁺CD4⁺, CD3⁺CD8⁺, CD3⁺CD4^-CD8^-TCRαβ⁺ DN T cells, non-GC B cells [gated as B220⁺PNA^-CD38^high] and GC B cells [gated as B220⁺PNA⁺CD38^low]). (B) Fas downregulation in GC B cells (gated as B220⁺PNA⁺CD38^low) in dLNs from TG mice. The mean fluorescence intensity (MFI) of the Fas expression detected by FACS was calculated and presented. (C) Putative miR-146a-binding sites in the 3′-UTR of Fas from the rat, mouse, and human. The sequences are partially complementary to each other (left). Cotransfection of 293T cells with the pEZX-Luc vector containing the Fas 3′-UTR (from the rat, mouse, or human) and a plasmid encoding either miR-146a or miR-scramble (Ctrl), and the pEZX-Luc empty vector was also used as a control (bottom). Data are expressed in terms of mean ± standard deviation values. Comparison of means was performed with an unpaired Student 2-tailed t test (**P < .01).