RNA-binding proteins control the G2-M checkpoint of the germinal center B cell [TrAEL-seq]

How germinal centre (GC) B cells undergo rapid cell division while maintaining genome stability is poorly understood. Here, we show that the RNA-binding proteins ZFP36L1 and ZFP36L2 act downstream of antigen-sensing and protect GC B cells from replication stress by controlling a cell cycle-related RNA post-transcriptional regulon. ZFP36L1 and ZFP36L2 safeguard faithful completion of mitosis by restraining the expression of CDK1 and cyclin B1, whilst controlling their activity through regulation of a p21-mediated negative feedback loop. In the absence of ZFP36L1 and ZFP36L2, GC B cells arrest in G2-M and die by apoptosis, resulting in curtailed GC responses. This is associated with stalling of the DNA replication fork at active replication initiation zones, which causes replication stress and increased activity of the ATR/CHK1 DNA damage response. Our findings reveal that gene regulation by RNA-binding proteins is essential for a functional G2-M checkpoint to operate in GC B cells. Naive B cells isolated from peripheral lymph nodes of Zfp36l1 fl/fl Zfp36l2 fl/fl and Zfp36l1 fl/fl Zfp36l2 fl/fl CD23cre mice (2 replicates each), were activated in vitro for 7 days in iGCB culture conditions. Agarose-embedded DNA from 2x10^6 iGCB cells were taken and processed into TrAEL-seq libraries, to track replication fork sites and profile replication origin usage genome-wide