Abstract
8-Oxo-2'-deoxyguanosine (8-oxodG) is one of the most important oxidative DNA lesions, and G-rich telomeric DNA is especially susceptible to oxidative DNA damage. RecQ helicases WRN and BLM and telomere-binding protein POT1 are thought to play roles in telomere maintenance. This study examines the ability of WRN, BLM, and RecQ5 to unwind and POT1 to bind telomeric D-loops containing 8-oxodG. The results demonstrate that WRN and BLM preferentially unwind telomeric D-loops containing 8-oxodG and that POT1 binds with higher affinity to telomeric D-loops with 8-oxodG but shows no preference for telomeric single-stranded DNA with 8-oxodG. We speculate that telomeric D-loops with 8-oxodG may have a greater tendency to form G-quadruplex DNA structures than telomeric DNA lacking 8-oxodG.
Highlights
Telomeres are structures at the ends of the eukaryotic linear chromosomes that enhance chromosome stability by preventing DNA end-initiated recombination, exonucleolytic attack, and replication-associated terminal recession
8-Oxo-2-deoxyguanosine (8-oxodG) is one of the most important oxidative DNA lesions, and G-rich telomeric DNA is especially susceptible to oxidative DNA damage
The results demonstrate that WRN and BLM preferentially unwind telomeric D-loops containing 8-oxodG and that protection of telomeres 1 (POT1) binds with higher affinity to telomeric D-loops with 8-oxodG but shows no preference for telomeric single-stranded DNA with 8-oxodG
Summary
8-oxodG, 8-oxo-2Ј-deoxyguanosine; ssDNA, single-stranded DNA; MBN, mung bean nuclease; EMSA, electrophoretic mobility shift assay; DMS, dimethyl sulfate; TMS, telomestatin; GST, glutathione S-transferase; TBE, Tris borate-EDTA; DTT, dithiothreitol; BSA, bovine serum albumin; WRN, Werner syndrome protein; BLM, Bloom syndrome protein. Several lesions can be generated by oxidative damage, including 8-oxodG, thymine glycol, formamidopyrimidine-2Ј-deoxyguanosine, formamidopyrimidine-2Ј-deoxyadenosine, and many more. Many of these lesions may have important biological roles, most of them are not well studied. It is improbable to assess the effects of all the lesions due to the complications involved in the synthesis of DNA strands containing the damages. Among these lesions, 8-oxodG is an abundant and well characterized lesion and is of great biological significance. These findings indicate that these proteins have important roles in damaged telomeric DNA processing and repair
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