title: A Role for the Fifth G-Track in G-Quadruplex Forming Oncogene Promoter Sequences during Oxidative Stress: Do These "Spare Tires" Have an Evolved Function?
abstract: Uncontrolled inflammation or oxidative stress generates electron-deficient species that oxidize the genome increasing its instability in cancer. The G-quadruplex (G4) sequences regulating thec-MYC,KRAS,VEGF,BCL-2,HIF-1α, andREToncogenes, as examples, are targets for oxidation at loop and 5'-core guanines (G) as showcased in this study by CO3•-oxidation of theVEGFG4. Products observed include 8-oxo-7,8-dihydroguanine (OG), spiroiminodihydantoin (Sp), and 5-guanidinohydantoin (Gh). Our previous studies found that OG and Gh, when present in the four G-tracks of the solved structure forVEGFandc-MYC, were not substrates for the base excision repair (BER) DNA glycosylases in biologically relevant KCl solutions. We now hypothesize that a fifth G-track found a few nucleotides distant from the G4 tracks involved in folding can act as a "spare tire," facilitating extrusion of a damaged G-run into a large loop that then becomes a substrate for BER. Thermodynamic, spectroscopic, and DMS footprinting studies verified the fifth domain replacing a damaged G-track with OG or Gh at a loop or core position in theVEGFG4. These new "spare tire"-containing strands with Gh in loops are now found to be substrates for initiation of BER with the NEIL1, NEIL2, and NEIL3 DNA glycosylases. The results support a hypothesis in which regulatory G4s carry a "spare-tire" fifth G-track for aiding in the repair process when these sequences are damaged by radical oxygen species, a feature observed in a large number of these sequences. Furthermore, formation and repair of oxidized bases in promoter regions may constitute an additional example of epigenetic modification, in this case of guanine bases, to regulate gene expression in which the G4 sequences act as sensors of oxidative stress.
authors.affiliation: Department of Chemistry, University of Utah , Salt Lake City, Utah 84112, United States.
authors.forename: Aaron M
authors.initials: AM
authors.lastname: Fleming
authors.affiliation: Department of Microbiology and Molecular Genetics, University of Vermont , Burlington, Vermont 05045, United States.
authors.forename: Jia
authors.initials: J
authors.lastname: Zhou
authors.affiliation: Department of Microbiology and Molecular Genetics, University of Vermont , Burlington, Vermont 05045, United States.
authors.forename: Susan S
authors.initials: SS
authors.lastname: Wallace
authors.affiliation: Department of Chemistry, University of Utah , Salt Lake City, Utah 84112, United States.
authors.forename: Cynthia J
authors.initials: CJ
authors.lastname: Burrows