We describe the unique structural features of a large telomere repeat
DNA complex (TRDC) of >20 kb generated by a simple PCR using (TTAGGG)(4) and (CCCTAA)(4) as both primers and templates. Although large, as determined by conventional
agarose gel electrophoresis, the TRDC was found to consist of short
single-stranded DNA telomere repeat units of between several hundred and 3000 bases, indicating that it is a non-covalent complex comprising short cohesive telomere repeat units. S1 nuclease digestion showed that the TRDC contains both single- and double-stranded portions stable enough to survive
glycerol density gradient centrifugation, precipitation with
ethanol and gel electrophoresis. Sedimentation analysis suggests that a part of the TRDC is non-linear and consists of a three-dimensional network structure.
After treatment with Werner
DNA helicase the TRDC dissociated into smaller fragments, provided that human
replication protein A was present, indicating that: (i) the TRDC is a new substrate for the
Werner syndrome helicase; (ii) the telomere repeat sequence re-anneals rapidly unless unwound single-stranded regions are protected by
replication protein A; (iii) the TRDC may provide a new clue to understanding deleterious telomere-totelomere interactions that can lead to
genomic instability. Some properties of the TRDC account for the extra-chromosomal telomere repeat (ECTR)
DNA that exists in
telomerase-negative immortalized cell lines and may be involved in maintaining telomeres.