AID-dependent histone acetylation is detected in immunoglobulin S regions

Abstract

Class switch recombination (CSR) is regulated by the expression of activation-induced deaminase (AID) and germline transcripts (GLTs). AID-dependent double-strand breaks (DSBs) are introduced into switch (S) regions and stimulate CSR. Although histone acetylation (Ac) has been well documented in transcription regulation, its role in DNA damage repair remains largely unexplored. The 1B4.B6 B cell line and normal splenic B cells were activated to undergo CSR and analyzed for histone Ac by chromatin immunoprecipitation (ChIP). A detailed study of the Iγ3-Sγ3-Cγ3 locus demonstrated that acetylated histones are focused to the Iγ3 exon and the Sγ3 region but not to the intergenic areas. Histone H3 Ac is strongly correlated with GLT expression at four S regions, whereas H4 Ac was better associated with B cell activation and AID expression. To more directly examine the relationship between H4 Ac and AID, LPS-activated AID KO and WT B cells were analyzed and express comparable levels of GLTs. In AID-deficient B cells, both histones H3 and H4 are reduced where H4 is more severely affected as compared with WT cells. Our findings raise the intriguing possibility that histone H4 Ac at S regions is a marker for chromatin modifications associated with DSB repair during CSR.

NF‐κB binds to the immunoglobulin Sγ3 region in vivo during class switch recombination

“…2A). The μ and γ3 GLT are constitutively expressed in unstimulated splenic B cells, as previously observed [21]. These GLT continued their expression following LPS activation and were not affected by IL-4 treatment, whereas γ1GLT expression was IL-4 inducible.…”

“…These GLT continued their expression following LPS activation and were not affected by IL-4 treatment, whereas γ1GLT expression was IL-4 inducible. The levels of GLT and AID transcripts remain essentially constant from the 24 to 48-h time points [21]. The post-switch transcript (PST) expression profile, taken after 5 days of activation, confirms successful CSR in our B cell cultures (Fig.…”

“…The earliest time that completed CSR has been detected is 72 h following stimulation [21,22]. Consequently, time points prior to 72 h of B cell activation are likely to represent predisposing events for CSR, whereas at later times these alterations may be associated with post-recombination modifications.…”

“…The NF-κB-binding sites of SNIP motifs are replete throughout the Sγ3 and Sγ1 TR but the repetitive nature of S regions necessitates using primers for ChIP located immediately flanking the TR ( [21] and Supplementary Fig. 2).…”

“…Histone hyper Ac has been taken as a measure of S region accessibility [21,30,31]. We found that histone hyper Ac is stimulated by LPS alone and augmented by IL-4, indicating that B cell activation is sufficient to stimulate chromatin remodeling [21]. To explore the relationship between GLT expression, histone hyper Ac, and NF-κB binding at the Sγ1 region we used ChIP analysis to examine p50 and p65 binding.…”