DNA and RNA can fold into a variety of alternative conformations. In recent years, a particular nucleic acid structure was discussed to play a role in malignant transformation and cancer development. This structure is called a G-quadruplex (G4). G4 structure formation can drive genome instability by creating mutations, deletions and stimulating recombination events. The importance of G4 structures in the characterization of malignant cells was currently demonstrated in breast cancer samples. In this analysis a correlation between G4 structure formation and an increased intratumor heterogeneity was identified. This suggests that G4 structures might allow breast cancer stratification and supports the identification of new personalized treatment options. Because of the stability of G4 structures and their presence within most human oncogenic promoters and at telomeres, G4 structures are currently tested as a therapeutic target to downregulate transcription or to block telomere elongation in cancer cells. To date, different chemical molecules (G4 ligands) have been developed that aim to target G4 structures. In this review we discuss and compare G4 function and relevance for therapeutic approaches and their impact on cancer development for three cancer entities, which differ significantly in their amount and type of mutations: pancreatic cancer, leukemia and malignant melanoma. G4 structures might present a promising new strategy to individually target tumor cells and could support personalized treatment approaches in the future.

Translation efficiency can be affected by mRNA stability and secondary structures, including G-quadruplex structures (G4s). The highly conserved DEAH-box helicase DHX36/RHAU resolves G4s on DNA and RNA in vitro, however a systems-wide analysis of DHX36 targets and function is lacking. We map globally DHX36 binding to RNA in human cell lines and find it preferentially interacting with G-rich and G4-forming sequences on more than 4500 mRNAs. While DHX36 knockout (KO) results in a significant increase in target mRNA abundance, ribosome occupancy and protein output from these targets decrease, suggesting that they were rendered translationally incompetent. Considering that DHX36 targets, harboring G4s, preferentially localize in stress granules, and that DHX36 KO results in increased SG formation and protein kinase R (PKR/EIF2AK2) phosphorylation, we speculate that DHX36 is involved in resolution of rG4 induced cellular stress.

The concept that G-quadruplex (G4) structures can form within DNA or RNA in vitro has been long known and extensively discussed. In recent years, accumulating evidences imply that G-quadruplex structures form in vivo. Initially, inefficient regulation of G-quadruplex structures was mainly associated with genome instability. However, due to the location of G-quadruplex motifs and their evolutionary conservation, different cellular functions of these structures have been postulated (e.g. in telomere maintenance, DNA replication, transcription, and translation). Regardless of their function, efficient and controlled formation and unwinding are very important, because ‘mis’-regulated G-quadruplex structures are detrimental for a given process, causing genome instability and diseases. Several helicases have been shown to target and regulate specific G-quadruplex structures. This mini-review focuses on the biological consequences of G4 disruption by different helicases in vivo.

DNA molecules can adopt a variety of alternative structures. Among these structures are G-quadruplex DNA structures (G4s), which support cellular function by affecting transcription, translation, and telomere maintenance. These structures can also induce genome instability by stalling replication, increasing DNA damage, and recombination events. G-quadruplex-driven genome instability is connected to tumorigenesis and other genetic disorders. In recent years, the connection between genome stability, DNA repair and G4 formation was further underlined by the identification of multiple DNA repair proteins and ligands which bind and stabilize said G4 structures to block specific DNA repair pathways. The relevance of G4s for different DNA repair pathways is complex and depends on the repair pathway itself. G4 structures can induce DNA damage and block efficient DNA repair, but they can also support the activity and function of certain repair pathways. In this review, we highlight the roles and consequences of G4 DNA structures for DNA repair initiation, processing, and the efficiency of various DNA repair pathways.

Previously we reported the production and characterization of monoclonal antibody 1H6 raised against (T₄G₄)₂ intermolecular guanine quadruplex (G4) DNA structures (Henderson A. et al. (2014) Nucleic Acids Res., 42, 860–869; Hoffmann R.F. et al. (2016) Nucleic Acids Res., 44, 152–163). It was shown that 1H6 strongly stains nuclei and has exquisite specificity for heterochromatin by immuno-electron microscopy. Here we extend our studies of 1H6 reactivity using enzyme-linked immunosorbent assay (ELISA) and microscale thermophoresis (MST). As previously reported, 1H6 was found to strongly bind intermolecular G4 structures with a (T₄G₄)₂ sequence motif. However, using both methods we did not detect significant binding to G4 structures without thymidines in their sequence motif or to G4 structures made with (T₂G₄)₂ oligonucleotides. In addition, we observed strong, sequence-specific binding of 1H6 by ELISA to immobilized single stranded poly(T) DNA but not to immobilized poly(C) or poly(A) homo-polymers. Cross-reactivity of 1H6 to poly(T) was not measured in solution using MST. 1H6 was furthermore found to bind to selected areas on DNA fibers but only after DNA denaturation. Based on these observations we propose that 1H6 binds with high affinity to adjacent T's that are restricted in their movement in selected G4 structures and denatured DNA. Cross-reactivity of 1H6 to immobilized single stranded T-rich DNA next to its previously reported specificity for bona fide G4 structures needs to be taken into account in the interpretation of 1H6 binding to (sub-) cellular structures.

Accurate DNA replication is essential for genomic stability and cancer prevention. Homologous recombination is important for high-fidelity DNA damage tolerance during replication. How the homologous recombination machinery is recruited to replication intermediates is unknown. Here, we provide evidence that a Rad51 paralog-containing complex, the budding yeast Shu complex, directly recognizes and enables tolerance of predominantly lagging strand abasic sites. We show that the Shu complex becomes chromatin associated when cells accumulate abasic sites during S phase. We also demonstrate that purified recombinant Shu complex recognizes an abasic analog on a double-flap substrate, which prevents AP endonuclease activity and endonuclease-induced double-strand break formation. Shu complex DNA binding mutants are sensitive to methyl methanesulfonate, are not chromatin enriched, and exhibit increased mutation rates. We propose a role for the Shu complex in recognizing abasic sites at replication intermediates, where it recruits the homologous recombination machinery to mediate strand specific damage tolerance.