Mutational landscape of the G-quadruplex DNA in cMyc proto-oncogene promoter: Insights into the structural polymorphism and transcriptional regulation

Noncanonical four-stranded DNA structures known as G-quadruplexes (G4s) play crucial roles in gene regulation and have recently been identified as hotspots for single-nucleotide variations (SNVs). In the cMyc proto-oncogene promoter, the Pu27 G4 element serves as a key transcriptional switch, yet its susceptibility to SNVs and the resulting molecular consequences remain unclear. Here, we systematically analyze SNVs in the cMyc promoter G4 using the dbSNP database and identify 17 mutation sites distributed across its 27-nt sequence, indicating a high mutation frequency. Biophysical studies reveal that SNVs in loop regions primarily enhance G4 stability by introducing additional hydrogen bonds and π-π interactions while maintaining its parallel topology. In contrast, SNVs within G-runs destabilize the G4, leading to structural polymorphism, mixed topologies, and increased conformational dynamics; certain SNVs induce noncanonical G4 architectures such as G-vacancies and bulges. Functional assays show that these structural changes differentially modulate transcription from the cMyc promoter, with stabilizing SNVs generally suppressing expression and destabilizing SNVs producing divergent effects. Overall, our findings establish G4s as dynamic sensors of genetic variation and provide a mechanistic framework for understanding how non-B DNA motifs contribute to transcriptional dysregulation and genome instability in cancer.