GGA center dot TCC-interrupted triplets in long GAA center dot TTC repeatsinhibit the formation of tripler and sticky DNA structures, alleviate transcription inhibition, and reduce genetic instabilities

Large expansions of GAA.TTC repeats in the first intron of the frataxin (X2 5) gene are the principal mutation responsible for Friedreich's ataxia (FRD A). Sticky DNA, based on R.R.Y triplexes, was found at the expanded GAA.TTC repeats from FRDA patients. The (GAAGGA.TCCTTC)(65) repeat occurs in the s ame frataxin locus but is nonpathogenic and does not form sticky DNA. To el ucidate the behavior of sticky DNA, we introduced various extents of GGA.TC C interruptions into the long GAA.TTC repeat, More than 20% of GGA.TCC inte rruptions abolished the formation of sticky DNA. However, the GAA.TTC repea ts with less than 11% of GGA.TCC interruptions formed triplexes and/or stic ky DNA similar to the uninterrupted repeat sequence. These triplexes showed different P1 nuclease sensitivities, and the GGA.TCC interruptions were sl ightly more sensitive than the surrounding GAA.TTC repeats. Furthermore, ge netic instability investigations in Escherichia coli revealed that a small number (4%) of interruptions substantially stabilized the long GAA.TTC trac ts. Furthermore, the greater the extent of interruptions of the GAA.TTC rep eats, the less inhibition of in vitro transcription was observed, as expect ed, based on the capacity of interruptions to inhibit the formation of stic ky DNA We propose that the interruptions introduce base mismatches into the R.R.Y triplex, which explains the observed chemical and biological propert ies.