t(8;14)易位是Burkitt淋巴瘤的标志并导致c-MYC失调。在易位过程中,8号染色体上的c-MYC基因与14号染色体上的Ig开关区并列。尽管已经研究了c-MYC启动子的脆弱性机制,对其他c-MYC断点区域知之甚少。我们已经分析了伯基特淋巴瘤患者c-MYC基因座外显子1/内含子1的易位断裂点。结果表明,断点区域,当存在于质粒上时,可以以转录依赖的方式折叠成R环确认。亚硫酸氢钠修饰试验显示Burkitt淋巴瘤细胞系染色体DNA上存在显著的单丝,Raji,和正常的淋巴细胞,揭示覆盖高达100bp区域的不同R环。此外,ChIP-DRIP分析揭示了R环抗体可以与断点区结合。Further,我们显示了在基因组的非模板链上形成稳定的平行分子内G-四链体。最后,体外培养纯化的AID或细胞内AID的过表达导致c-MYC断点区突变频率增强。有趣的是,抗-γH2AX可以与在细胞内的c-MYC断裂点区域产生的DSB结合。发现R-环和G-四链体的形成是相互排斥的。因此,我们的结果表明,AID可以与R环和G4DNA的单链区域结合,导致胞嘧啶脱氨基为尿嘧啶,并在其中一条DNA链中诱导DNA断裂,导致双股断裂,这可能最终导致t(8;14)染色体易位。
t(8;14) translocation is the hallmark of Burkitt\'s lymphoma and results in c-MYC deregulation. During the translocation, c-MYC gene on chromosome 8 gets juxtaposed to the Ig switch regions on chromosome 14. Although the promoter of c-MYC has been investigated for its mechanism of fragility, little is known about other c-MYC breakpoint regions. We have analyzed the translocation break points at the exon 1/intron 1 of c-MYC locus from patients with Burkitt\'s lymphoma. Results showed that the breakpoint region, when present on a plasmid, could fold into an R-loop confirmation in a transcription-dependent manner. Sodium bisulfite modification assay revealed significant single-strandedness on chromosomal DNA of Burkitt\'s lymphoma cell line, Raji, and normal lymphocytes, revealing distinct R-loops covering up to 100 bp region. Besides, ChIP-DRIP analysis reveals that the R-loop antibody can bind to the breakpoint region. Further, we show the formation of stable parallel intramolecular G-quadruplex on non-template strand of the genome. Finally, incubation of purified AID in vitro or overexpression of AID within the cells led to enhanced mutation frequency at the c-MYC breakpoint region. Interestingly, anti-γH2AX can bind to DSBs generated at the c-MYC breakpoint region within the cells. The formation of R-loop and G-quadruplex was found to be mutually exclusive. Therefore, our results suggest that AID can bind to the single-stranded region of the R-loop and G4 DNA, leading to the deamination of cytosines to uracil and induction of DNA breaks in one of the DNA strands, leading to double-strand break, which could culminate in t(8;14) chromosomal translocation.