The telomeric GGGTTA repeats of Trypanosoma brucei contain the hypermodified base J in both strands

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Abstract
We have previously shown that nuclear DNA of bloodstream form Trypanosoma brucei contains a novel base β-glucosyl-hydroxymethyluracil, called J. Base J is enriched in minichromosome fractions but not in the minichromosome internal repeats, suggesting the association of J with telomeric DNA. To test whether J is present in the long telomeric (GGGTTA)n repeat arrays, which are 2–26 kb in T.brucei, we have purified these arrays both by hybrid selection and by isolating 2–26 kb fragments from DNA digested with multiple restriction enzymes. We find that in purified telomeric repeats ∼13% of T is replaced by J, compared to 0.8% in total DNA, and we estimate that ∼50% of the total J is in these repeats. Highly purified complementary strands of the repeats were obtained by alkaline CsCl equilibrium centrifugation. In the (TAACCC)n strand 14% of T was replaced by J. In the (GGGTTA)n strand ∼36% of the second T was replaced by J; the first T was not detectably replaced. Modified bases have not been found in telomeric repeats before. How the bulky base J affects telomere function and structure in bloodstream form trypanosomes remains to be determined.