Transcription pause and elongation regulators mediate somatic hypermutation

Abstract

Somatic hypermutation (SHM) creates point mutations in immunoglobulin (Ig) gene variable (V) regions in activated B cells to support antibody affinity maturation1. SHM is initiated by activation induced deaminase (AID), a cytosine deaminase whose single-stranded DNA (ssDNA) substrate is thought to arise during RNA Polymerase II (Pol II) transcription, perhaps in the context of Pol II stalling2,3. The factors and mechanisms that mediate AID chromatin recruitment and generate its putative stalled Pol II substrate during SHM are unknown. Negative Elongation Factor (NELF) stabilizes Pol II in a paused state near the transcription start site. Release of NELF from Pol II and the Pol II elongation-versus-termination decision are regulated by opposing super elongation complex (SEC) kinase and Integrator-protein phosphatase 2A (INT-PP2A) activities4. Here we demonstrate that NELF, SEC components MLLT1 and MLLT3, and the INT-PP2A phosphatase module are required for SHM. Unexpectedly, we find that NELF can associate with Pol II in the AID “target window” in the gene body and that in the absence of NELF, AID fails to deaminate its targets despite efficient recruitment to chromatin and transcription by Pol II. MLLT1 and MLLT3 interact with AID through their histone binding YEATS domain and are required for enrichment of AID at its target sites. We propose a physiological function for NELF-Pol II association in gene bodies in creating the stalled Pol II substrate for AID action during SHM.