DNA secondary structures, including G-quadruplex (G4) structures, are non-B-form DNA conformations that can form in vivo and play roles in genome stability and cellular processes. G4 structures are stacked nucleic acid structures formed by guanine-rich sequences and are stabilized by monovalent cations. They can adopt various topologies, such as parallel, antiparallel, or hybrid forms, and are found in diverse genomic regions, including telomeres, promoters, and ribosomal DNA. G4 structures are thought to influence DNA replication, transcription, and genome stability by resolving or stabilizing these structures. Telomeres, which are essential for chromosome protection, contain high concentrations of G4 motifs due to their G-rich sequences. Studies in ciliates and yeast show that G4 structures can form at telomeres and are resolved during DNA replication. Telomerase, a telomere-specific enzyme, can be influenced by G4 structures, and small molecule ligands that stabilize G4 structures are being tested for their potential to inhibit telomerase activity. G4 structures also play roles in transcription by influencing the accessibility of DNA to transcription machinery. G4 motifs are enriched in promoter regions of many genes, and their presence can repress or enhance transcription depending on their location. For example, G4 structures in the promoter region of the MYC gene can repress transcription, and compounds that stabilize G4 structures can reduce MYC expression. G4 structures are also involved in meiosis and recombination. They may assist in the formation of the telomere-dependent bouquet structure during meiosis and promote homologous recombination. Additionally, G4 structures are implicated in the regulation of antigenic variation in pathogens like Neisseria gonorrhoeae, where G4-based recombination mechanisms control the expression of surface antigens. Despite growing evidence for the in vivo existence of G4 structures, their physiological relevance remains debated. While in vitro studies show that G4 structures can form and resolve, direct in vivo evidence is limited. However, genetic experiments and the presence of G4-specific antibodies suggest that G4 structures do exist in vivo and may have functional roles in genome stability and cellular processes. Further research is needed to fully understand the mechanisms and implications of G4 structures in DNA biology.DNA secondary structures, including G-quadruplex (G4) structures, are non-B-form DNA conformations that can form in vivo and play roles in genome stability and cellular processes. G4 structures are stacked nucleic acid structures formed by guanine-rich sequences and are stabilized by monovalent cations. They can adopt various topologies, such as parallel, antiparallel, or hybrid forms, and are found in diverse genomic regions, including telomeres, promoters, and ribosomal DNA. G4 structures are thought to influence DNA replication, transcription, and genome stability by resolving or stabilizing these structures. Telomeres, which are essential for chromosome protection, contain high concentrations of G4 motifs due to their G-rich sequences. Studies in ciliates and yeast show that G4 structures can form at telomeres and are resolved during DNA replication. Telomerase, a telomere-specific enzyme, can be influenced by G4 structures, and small molecule ligands that stabilize G4 structures are being tested for their potential to inhibit telomerase activity. G4 structures also play roles in transcription by influencing the accessibility of DNA to transcription machinery. G4 motifs are enriched in promoter regions of many genes, and their presence can repress or enhance transcription depending on their location. For example, G4 structures in the promoter region of the MYC gene can repress transcription, and compounds that stabilize G4 structures can reduce MYC expression. G4 structures are also involved in meiosis and recombination. They may assist in the formation of the telomere-dependent bouquet structure during meiosis and promote homologous recombination. Additionally, G4 structures are implicated in the regulation of antigenic variation in pathogens like Neisseria gonorrhoeae, where G4-based recombination mechanisms control the expression of surface antigens. Despite growing evidence for the in vivo existence of G4 structures, their physiological relevance remains debated. While in vitro studies show that G4 structures can form and resolve, direct in vivo evidence is limited. However, genetic experiments and the presence of G4-specific antibodies suggest that G4 structures do exist in vivo and may have functional roles in genome stability and cellular processes. Further research is needed to fully understand the mechanisms and implications of G4 structures in DNA biology.