Long interspersed nuclear element-1 hypomethylation and oxidative stress: correlation and bladder cancer diagnostic potential

Although, increased oxidative stress and hypomethylation of long interspersed nuclear element-1 (LINE-1) associate with bladder cancer (BCa) development, the relationship between these alterations is unknown. We evaluated the oxidative stress and hypomethylation of the LINE-1 in 61 BCa patients and 45 normal individuals. To measure the methylation levels and to differentiate the LINE-1 loci into hypermethylated, partially methylated and hypomethylated, peripheral blood cells, urinary exfoliated cells and cancerous tissues were evaluated by combined bisulfite restriction analysis PCR. The urinary total antioxidant status (TAS) and plasma protein carbonyl content were determined. The LINE-1 methylation levels and patterns, especially hypomethylated loci, in the blood and urine cells of the BCa patients were different from the levels and patterns in the healthy controls. The urinary TAS was decreased, whereas the plasma protein carbonyl content was increased in the BCa patients relative to the controls. A positive correlation between the methylation of LINE-1 in the blood-derived DNA and urinary TAS was found in both the BCa and control groups. The urinary hypomethylated LINE-1 loci and the plasma protein carbonyl content provided the best diagnostic potential for BCa prediction. Based on post-diagnostic samples, the combination test improved the diagnostic power to a sensitivity of 96% and a specificity of 96%. In conclusion, decreased LINE-1 methylation is associated with increased oxidative stress both in healthy and BCa subjects across the various tissue types, implying a dose-response association. Increases in the LINE-1 hypomethylation levels and the number of hypomethylated loci in both the blood- and urine-derived cells and increase in the oxidative stress were found in the BCa patients. The combination test of the urinary hypomethylated LINE-1 loci and the plasma protein carbonyl content may be useful for BCa screening and monitoring of treatment.     

Characteristics of replication-independent endogenous double-strand breaks in Saccharomyces cerevisiae

Background

Replication-independent endogenous double-strand breaks (RIND-EDSBs) occur in both humans and yeast in the absence of inductive agents and DNA replication. In human cells, RIND-EDSBs are hypermethylated, preferentially retained in the heterochromatin and unbound by γ-H2AX. In single gene deletion yeast strains, the RIND-EDSB levels are altered; the number of RIND-EDSBs is higher in strains with deletions of histone deacetylase, endonucleases, topoisomerase, or DNA repair regulators, but lower in strains with deletions of the high-mobility group box proteins or Sir2. In summary, RIND-EDSBs are different from pathologic DSBs in terms of their causes and consequences. In this study, we identified the nucleotide sequences surrounding RIND-EDSBs and investigated the features of these sequences as well as their break locations.

Results

In recent work, we detected RIND-EDSBs using ligation mediated PCR. In this study, we sequenced RIND-EDSB PCR products of resting state Saccharomyces cerevisiae using next-generation sequencing to analyze RIND-EDSB sequences. We found that the break locations are scattered across a number of chromosomes. The number of breaks correlated with the size of the chromosomes. Most importantly, the break occurrences had sequence pattern specificity. Specifically, the majority of the breaks occurred immediately after the sequence “ACGT” (P = 2.2E-156). Because the “ACGT” sequence does not occur primarily in the yeast genome, this specificity of the “ACGT” sequence cannot be attributed to chance.

Conclusions

RIND-EDSBs occur non-randomly; that is, they are produced and retained by specific mechanisms. Because these particular mechanisms regulate their generation and they possess potentially specific functions, RIND-EDSBs could be epigenetic marks.

Electronic supplementary material

The online version of this article (doi:10.1186/1471-2164-15-750) contains supplementary material, which is available to authorized users.