Formation of multiple G-quadruplexes contributes toward BCR fragility associated with chronic myelogenous leukemia.

Abstract

The Philadelphia chromosome, the translocation between BCR and ABL genes, is seen in 95% of chronic myeloid leukemia (CML) patients. Although discovered >60 years ago, the molecular mechanism of BCR fragility is unclear. Here, we have identified several G4 DNA motifs at the BCR fragile region of CML patients. Various lines of experimentation revealed that the breakpoint regions could fold into multiple intramolecular G-quadruplex structures. The sodium bisulfite modification assay revealed single strandedness in the fragile region when present on a plasmid and human genome. Circular dichroism spectroscopy revealed the parallel G4 DNA formation, leading to polymerase arrest at the BCR breakpoints. Intracellular recombination assay revealed that DNA breakage at the BCR fragile region could join with the break generated by ISceI endonuclease. Finally, purified AID could bind and deaminate cytosines when present on single-stranded DNA generated due to G4 DNA, both in vitro and inside the cells. Therefore, our results suggest that AID binds to G4 DNA present at the BCR fragile region, resulting in the deamination of cytosines to uracil and induction of DNA breaks in one of the DNA strands, which can later get converted into a double-strand break, leading to t(9;22) chromosomal translocation.