UnAIDed Class Switching in Activated B-Cells Reveals Intrinsic Features of a Self-Cleaving IgH Locus

Abstract

Activation-induced deaminase (AID) is the major actor of immunoglobulin (Ig) gene diversification in germinal center B-cells. From its first description, it was considered as mandatory for class switch recombination (CSR), and this discovery initiated a long quest for all of the AID-interacting factors controlling its activity. The mechanisms focusing AID-mediated DNA lesions to given target sequences remain incompletely understood with regards the detailed characterization of optimal substrates in which cytidine deamination will lead to double strand breaks (DSBs) and chromosomal cleavage. In an effort to reconsider whether such CSR breaks absolutely require AID, we herein provide evidence, based on deep-sequencing approaches, showing that this dogma is not absolute in both human and mouse B lymphocytes. In activated B-cells from either AID-deficient mice or human AID-deficient patients, we report an intrinsic ability of the IgH locus to undergo “on-target” cleavage and subsequent synapsis of broken regions in conditions able to yield low-level CSR. DNA breaks occur in such conditions within the same repetitive S regions usually targeted by AID, but their repair follows a specific pathway with increased usage of microhomology-mediated repair. These data further demonstrate the role of AID machinery as not initiating de novo chromosomal cleavage but rather catalyzing a process which spontaneously initiates at low levels in an appropriately conformed IgH locus.

Increased frequencies of IgH locus suicide recombination points on Chronic Lymphocytic Leukemia with low rate of AID related somatic mutations, cMYC overexpression and short telomeres

“…LSR could be induced in CLL proliferation centers in secondary lymphoid organs 1 . It has been shown that some residual CSR can occur in absence of AID 2 . This seems to result from random DNA breaks and inaccurate DSB repair as also implicated in chromosome translocations.…”

Roles of G4-DNA and G4-RNA in Class Switch Recombination and Additional Regulations in B-Lymphocytes

“…However, AID by itself is not sufficient for CSR to occur, and a large part of CSR regulation involves regulated accessibility of specific targeted regions within the IgH locus. A minimal level of CSR is even detectable in an accessible IgH locus in the absence of AID, highlighting the importance of the structure and the accessibility of S regions to recombination [ 69 ]. These data show that transcribed S regions undergo rare DNA breaks even in the absence of AID, likely due to the presence of G4s and R-loops at these regions.…”

“…AID G133V has genome-wide chromatin localization defects especially in the Sµ region [ 150 ]. Of note, while CSR is principally AID-dependent (and not only G4-dependent), a recent study showed that the structure of S regions and their programmed accessibility are sufficient for preserving a basal level of CSR in AID-independent conditions [ 69 ]. Indeed, in both AID-deficient mice and AID-mutant patients, CSR junctions remain detectable at low levels upon B cell activation.…”

G4-DNA, Class Switch Recombination and Potential Additional Regulations in B-Lymphocytes

“…However, AID by itself is not sufficient for CSR to occur, and a large part of CSR regulation involves regulated accessibility of specific targeted regions within the IgH locus. A minimal level of CSR is even detectable in an accessible IgH locus in the absence of AID, highlighting the importance of the structure and the accessibility of S regions to recombination [72]. Regulated accessibility to CSR is notably (but not only) related to transcriptional regulation of the S regions targeted for CSR [73][74][75][76].…”

“…AID G133V has genome-wide chromatin localization defects, and especially to Sµ region [159]. Of note, while CSR is principally AID-dependent (and not only G4-dependent), a recent study showed that the structure of S regions and their programmed accessibility are sufficient for preserving a basal level of CSR in AID-independent conditions [72]. Indeed, in both AID-deficient mice and AID-mutant patients, CSR junctions remain detectable at low level upon B cell activation.…”