Overexpressing and Inhibiting miRNA-155

To overexpress miRNA-155, the pre-miRNA with flanking sequences was amplified from genomic DNA using primers listed in Table S6. Insert was cloned into the retroviral MXW-PGK-IRES-GFP vector [30 (link)] using XhoI and EcoRI. To validate miRNA overexpression, GFP+ cells were sorted ∼2 weeks after transduction using a MoFlo sorter (Dako cytomation). We inhibited miR-155 using a retrovirally expressed miR-155 sponge with 14 binding sites (Table S6) [16 (link)]. The 3′UTR of DET1, TRIM32, JARID2, PSIP1, NIAM and the ∼300nt coding sequence fragment of HOMEZ containing the potential miR-155-binding sites were amplified from genomic DNA using primers listed in Table S6 and cloned into the psiCHECK2 vector (Promega, Madison, WI). To inhibit miR-155 target genes, shRNA oligos were designed (Table S6) and cloned into the retroviral pMDH1-PGK-GFP 2.0 vector [30 (link)]. The sequence of NIAMsh1 was based on a previous study [22 (link)]. Retroviral transductions were performed as previously described [28 (link)].