Yeast telomeric sequences function as chromosomal anchorage points in vivo.
Site-specific recombination in Saccharomyces cerevisiae was used to generate non-replicative DNA rings containing yeast telomeric sequences. In topoisomerase mutants expressing Escherichia coli topoisomerase I, the rings adopted a novel DNA topology consistent with the ability of yeast telomeric DNA to block or retard the axial rotation of DNA. DNA fragments bearing portions of the terminal repeat sequence C1-3 A/TG1-3 were both necessary and sufficient to create a barrier to DNA rotation. Synthetic oligonucleotide sequences containing Rap1p binding sites, a well represented motif in naturally occurring C1-3A arrays, also conferred immobilization; mutant Rap1p binding sites and telomeric sequences from other organisms were not sufficient. DNA anchoring was diminished by addition of competing telomeric sequences, implicating a role for an as yet unidentified limiting trans-acting factor. Though Rap1p is a likely protein constituent of the DNA anchor, deletion of the non-essential C-terminal domain did not affect the topology of telomeric DNA rings. Similarly, disruption of SIR2, SIR3 and SIR4, genes which influence a variety of telomere functions in yeast, also had no effect. We propose that telomeric DNA supports the formation of a SIR-independent macromolecular protein-DNA assembly that hinders the motion of DNA because of its linkage to an insoluble nuclear structure. Potential roles for DNA anchoring in telomere biology are discussed.
Mirabella
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A
Department of Pharmacology, UMDNJ-Robert Wood Johnson Medical School, Piscataway, NJ 08854, USA.
Gartenberg
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MR
D001665:Binding Sites / Q000235:genetics; D002875:Chromosomes / Q000378:metabolism*; D004247:DNA / Q000378:metabolism*; D004264:DNA Topoisomerases, Type I / Q000378:metabolism*; D004278:DNA, Superhelical / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000378:metabolism / Q000494:pharmacology; D004587:Electrophoresis, Agar Gel; D004926:Escherichia coli / Q000201:enzymology; D009690:Nucleic Acid Conformation; D012091:Repetitive Sequences, Nucleic Acid; D012441:Saccharomyces cerevisiae / Q000378:metabolism*; D016615:Telomere / Q000378:metabolism*
D004278:DNA, Superhelical; D004268:DNA-Binding Proteins; D004247:DNA; D004264:DNA Topoisomerases, Type I