c-Myc quadruplex-forming sequence Pu-27 induces extensive damage in both telomeric and nontelomeric regions of DNA.
Quadruplex-forming DNA sequences are present throughout the eukaryotic genome, including in telomeric DNA. We have shown that the c-Myc promoter quadruplex-forming sequence Pu-27 selectively kills transformed cells (Sedoris, K. C., Thomas, S. D., Clarkson, C. R., Muench, D., Islam, A., Singh, R., and Miller, D. M. (2012) Genomic c-Myc quadruplex DNA selectively kills leukemia. Mol. Cancer Ther. 11, 66-76). In this study, we show that Pu-27 induces profound DNA damage, resulting in striking chromosomal abnormalities in the form of chromatid or chromosomal breaks, radial formation, and telomeric DNA loss, which induces γ-H2AX in U937 cells. Pu-27 down-regulates telomeric shelterin proteins, DNA damage response mediators (RAD17 and RAD50), double-stranded break repair molecule 53BP1, G2 checkpoint regulators (CHK1 and CHK2), and anti-apoptosis gene survivin. Interestingly, there are no changes of DNA repair molecules H2AX, BRCA1, and the telomere maintenance gene, hTERT. ΔB-U937, where U937 cells stably transfected with deleted basic domain of TRF2 is partially sensitive to Pu-27 but exhibits no changes in expression of shelterin proteins. However, there is an up-regulation of CHK1, CHK2, H2AX, BRCA1, and survivin. Telomere dysfunction-induced foci assay revealed co-association of TRF1with γ-H2AX in ATM deficient cells, which are differentially sensitive to Pu-27 than ATM proficient cells. Alt (alternating lengthening of telomere) cells are relatively resistant to Pu-27, but there are no significant changes of telomerase activity in both Alt and non-Alt cells. Lastly, we show that this Pu-27-mediated sensitivity is p53-independent. The data therefore support two conclusions. First, Pu-27 induces DNA damage within both telomeric and nontelomeric regions of the genome. Second, Pu-27-mediated telomeric damage is due, at least in part, to compromise of the telomeric shelterin protein complex.
From the Department of Medicine and Molecular Targets Program, James Graham Brown Cancer Center, University of Louisville, Louisville, Kentucky 40202 and.
Md Ashraful
MA
Islam
Shelia D
SD
Thomas
Vundavalli V
VV
Murty
Kara J
KJ
Sedoris
Donald M
DM
Miller
D064007:Ataxia Telangiectasia Mutated Proteins / Q000235:genetics / Q000378:metabolism; D001483:Base Sequence; D016923:Cell Death; D045744:Cell Line, Tumor; D004247:DNA / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D004249:DNA Damage; D054856:G-Quadruplexes; D016259:Genes, myc; D006657:Histones / Q000235:genetics / Q000378:metabolism; D006801:Humans; D009369:Neoplasms / Q000235:genetics* / Q000378:metabolism / Q000473:pathology; D010766:Phosphorylation; D011401:Promoter Regions, Genetic; D016271:Proto-Oncogene Proteins c-myc / Q000235:genetics / Q000378:metabolism; D000089804:Shelterin Complex; D016615:Telomere / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D034501:Telomere-Binding Proteins / Q000235:genetics / Q000378:metabolism; D016159:Tumor Suppressor Protein p53 / Q000235:genetics / Q000378:metabolism
C468783:H2AX protein, human; D006657:Histones; D016271:Proto-Oncogene Proteins c-myc; D000089804:Shelterin Complex; D034501:Telomere-Binding Proteins; D016159:Tumor Suppressor Protein p53; D004247:DNA; D064007:Ataxia Telangiectasia Mutated Proteins
ATM; Alt Cell; Cell Death; DNA Damage; G-quadruplex; Myc; Phospho-H2AX; Shelterin; Survivin; Telomeres