NF-kB subunits direct kinetically distinct transcriptional cascades in antigen receptor-activated B cells [scRNA-seq]

The NF-kB family of transcription factors orchestrates signal-induced gene expression in a diversity of cell types. Cellular responses to NF-kB activation are regulated at the level of cell- and signal-specificity, as well as differential use of family members (subunit specificity). Here we used time-dependent multi-omics to investigate selective functions of Rel and RelA, two closely related NF-kB proteins, in primary B lymphocytes activated via the B cell receptor. Despite large numbers of shared binding sites genome wide, Rel and RelA directed kinetically distinct cascades of gene expression in activated B cells. Single-cell RNA-Seq revealed marked heterogeneity of Rel- and RelA-specific responses and sequential binding of these factors was not a major mechanism of protracted transcription. Moreover, nuclear co-expression of Rel and RelA led to functional antagonism between the two factors. By rigorously identifying target genes of each NF-kB subunit, these studies provide insights into exclusive functions of Rel and RelA in immunity and cancer. Overall design: Twelve 10,000 targeted B cell samples from floxed RelA mice (RelA-flfl (gift from Harmel-Laws Steinbrecher), N=1 for 0, 1, and 4 hour time points), RelA conditional knock-out (RelA-cKO (RelA-flfl x Cd19-cre) (006785, The Jackson Laboratory) mice, N=1 for 0, 1, and 4 hour time points), Rel knock-out mice (Rel-KO (gift from Hsia Cheng), N=1 for 0, 1, and 18 hour time points), and wildtype C57BL/6 mice (WT (006785 - The Jackson Laboratory), N=1 for each 0, 1, and 18 hour time points) were processed though the 10X Genomics 3' Gene Expression V.3.0 pipeline.