Deletion of Iα promoter/exon inhibits CSR to IgA. (A) Schematic structure of the IgH locus in CH12F3-2 line. The non-expressed allele is a partially rearranged DJH allele that underwent Sµ/Sα recombination, thus deleting all upstream inducible I promoters. The mitogen and cytokines inducing the different I promoters are indicated on top. The sites flanking Iα promoter/exon targeted by the gRNAs are indicated with arrows. The Eµ/Iµ enhancer/promoter between the variable and the constant regions, and the 3′RR super-enhancer downstream of the locus are shown. (B) Flow cytometry analysis of Iα-deleted clones. The 8 clones obtained by CRIPR/Cas9 were analyzed by FACS for IgA surface expression. The parental CH12 line was used as a control prior to (UNS) and following LIT (LPS + IL4 + TGFβ) stimulation. LIT-activated splenic B cells were also included as a control (n = 3). (C) Representative FACS plot obtained with CH12 cells and an Iα-deleted clone (n° 1), before and following LIT stimulation.