GSE167873
Role of Dot1L and H3K79 methylation in regulating somatic hypermutation of immunoglobulin genes
Somatic hypermutation (SHM) and class switch recombination (CSR) of the immunoglobulin (Ig) genes allow B cells to make antibodies that protect us against a wide variety of pathogens. SHM is mediated by activation induced deaminase (AID), occurs at a million times higher frequency than other mutations in the mammalian genome and is largely restricted to the variable (V) and switch (S) regions of Ig genes. Using the Ramos human Burkitt's lymphoma cell line, we find that H3K79me2/3 and its methyltransferase Dot1L are more abundant on the V region than the constant (C) region which does not undergo mutation. In primary naïve mouse B cells examined ex vivo, the H3K79me2/3 modification appears constitutively in the donor Sµ and is inducible in the recipient S?1 upon CSR stimulation. Knockout (KO) and inhibition of Dot1L in Ramos cells significantly reduces V region mutation and the abundance of H3K79me2/3 on the V region and is associated with a decrease of Pol II and its S2 phosphorylated form at the IgH locus. KO of Dot1L also decreases the abundance of BRD4 and CDK9 (a subunit of P-TEFb complex) on the V region and this is accompanied by decreased nascent transcripts throughout the IgH gene. Treatment with JQ1 (inhibitor of BRD4) or DRB (inhibitor of CDK9) decreased SHM and the abundance of Pol II S2P at the IgH locus. With all these factors playing a role in transcription elongation, our studies extend our insights into the chromatin context and dynamics of transcription required for SHM. Overall design: Examination of Dot1L in chromatin accessibility by ATAC-seq in human B cell line Ramos.