| Description |
Interest in oxidative modifications of the canonical bases, particularly the oxidation of guanine to 8-oxo-7,8-dihydroguanine (8-oxo-G), has led our laboratory to examine G-rich regions of the human genome for oxidative hot spots. This search led our laboratory to investigate the G-quadruplex (G4) secondary structure, which is often found within G-rich regions of the genome. The G4 field is burgeoning and has greatly expanded in the last 30 years. Recent work in our laboratory has connected the formation of 8-oxo-G with G4 folding via the base excision repair pathway (BER). Additionally, 8-oxo-G, traditionally seen as oxidative damage, may be acting as an epigenetic marker for gene regulation during oxidative stress. In our laboratory, research has been performed in mammalian systems, but little work has been done in plant or photosynthetic organisms. My goal was to develop a biological system that would allow the study of the biological role of G4s in a photosynthetic organism. Chlamydomonas reinhardtii, a soil green algae, was found to be an ideal system due to its high potential G-quadruplex-forming sequence (PQS) density, the previously developed expression system for foreign genes, and the existence of a transformation procedure requiring limited amounts of DNA. Furthermore, the physiological conditions within C. reinhardtii offered opportunities for probing the effects of the physiologically high polyamine concentration found in C. reinhardtii. iv Both the effects of magnesium concentration and polyamine concentration on the stability and structural equilibrium of two G4s selected from a cohort of characterized G4 sequences selected from the C. reinhardtii genome were examined. The results demonstrated that the high polyamine concentration drastically stabilized the G4s in vitro. The magnesium studies also showed some stabilization effects, but a more novel observation was the shift in structural equilibrium induced by the addition of magnesium. C. reinhardtii offers a novel physiological environment for studying G4s both in vitro and in vivo. |
