Fig. 4

Robust and prolonged endogenous FOXP3 upregulation in Jurkat cells. a, b Jurkat cells were electroporated with plasmids encoding control sgRNA (rnd), sgRNA for FOXP3 upregulation (Fox 1, 14, 15, 17 and 18; sequences in Table 1), sgRNA for EOS upregulation, sgRNA for FOXP3 upregulation in combination with an additional candidate gene (CTLA4, GATA3, GATA1, EOS, RELA, RELC, LEF1, IRF4; sequences listed in Table 2) and fold activation of exogenic FOXP3, Cas9, CD25, TNFR2, ICOS and IKZF2 were measured. Experiments were repeated 3 times. Cells were lysed and RNA isolated 48 h post electroporation c, d Jurkat cells were electroporated with either a plasmid encoding FOXP3 under the control of a CMV reporter (CMV-FOXP3) or plasmids for upregulation of endogenous FOXP3 via CRISPR–dCas9 (sgRNA FOXP3) or a pcDNA3 control vector. Fold activation and protein expression were measured on days 1, 4, 6 and 8 post electroporation