DNA-PROTEIN INTERACTIONS AT THE TELOMERIC REPEATS OF SCHIZOSACCHAROMYCES-POMBE

Gel retardation assays using a probe containing the repeat region of a Schizosaccharomyces pombe chromosomal telomere identified four specif ic DNA-protein complexes in S. pombe total protein extracts (I, I', II a and IIb). The proteins responsible for these complexes bound to the telomeric repeat region irrespective of whether or not the repeats wer e in close proximity to the end of a DNA molecule, and none of them bo und strongly to single-stranded DNA. The protein responsible for compl ex I (TeRF I) was separated from the activity responsible for complexe s IIa and IIb (TeRF II) using heparin-Sepharose chromatography. Both f actors were efficiently cross-competed by an oligonucleotide containin g the 18 bp sequence 5'-GGTT-ACAGGTTACAGGTT-3', which corresponds to t wo complete telomeric repeat units, Mutation of the T residues at posi tions 4 and 11 in the oligonucleotide dramatically reduced binding by TeRF II, but had no affect on binding by TeRF I. The protein responsib le for complex I' did not bind strongly to either the wild-type or mut ant oligonucleotide.