Summary
Local three-stranded DNA-RNA hybrid regions of the genomes (R-loops) have been detected either by binding of a monoclonal antibody (DRIP assay) or by enzymatic recognition by RNaseH. Such a structure has been postulated for the mouse and human telomeres, clearly suggested by the identification of the complementary RNA TERRA. It was, however, evidenced by DRIP exclusively in human cancer and not in normal human nor in any mouse cell. Based on the observation that in several fractionation procedures, DNA-RNA hybrids copurify with double-stranded DNA, we developed a preparative approach that allows detection of stable DNA-RNA triplexes in a complex genome, their physical isolation and their RNA nucleotide sequence. We then define in the normal mouse and human genomes the notion of terminal R-loop complexes including TERRA molecules synthesized from local promoters of every chromosome. In addition to the telomeric structures since, the finding of the RNA peak, applied in addition to the whole murine sperm genomes, has highlighted a reproducible profile of the R-loop complexes from entire genome suggesting a defined profile for a future generation.
