Abstract
In Saccharomyces cerevisiae, the sequence-specific binding of the negative regulator Rap1p provides a mechanism to measure telomere length: as the telomere length increases, the binding of additional Rap1p inhibits telomerase activity in cis. We provide evidence that the association of Rap1p with telomeric DNA in vivo occurs in part by sequence-independent mechanisms. Specific mutations in EST2 (est2-LT) reduce the association of Rap1p with telomeric DNA in vivo. As a result, telomeres are abnormally long yet bind an amount of Rap1p equivalent to that observed at wild-type telomeres. This behavior contrasts with that of a second mutation in EST2 (est2-up34) that increases bound Rap1p as expected for a strain with long telomeres. Telomere sequences are subtly altered in est2-LT strains, but similar changes in est2-up34 telomeres suggest that sequence abnormalities are a consequence, not a cause, of overelongation. Indeed, est2-LT telomeres bind Rap1p indistinguishably from the wild type in vitro. Taken together, these results suggest that Est2p can directly or indirectly influence the binding of Rap1p to telomeric DNA, implicating telomerase in roles both upstream and downstream of Rap1p in telomere length homeostasis.
Publication types
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Comparative Study
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Research Support, Non-U.S. Gov't
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Research Support, U.S. Gov't, Non-P.H.S.
MeSH terms
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Carrier Proteins / physiology
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Chromatin / metabolism*
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Chromosomes, Fungal / metabolism*
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Chromosomes, Fungal / ultrastructure
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DNA Helicases / deficiency
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DNA Helicases / physiology
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DNA, Fungal / metabolism
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DNA-Binding Proteins / deficiency
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DNA-Binding Proteins / physiology
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Gene Silencing
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Mutation, Missense
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Protein Binding
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RNA / physiology
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Recombinant Fusion Proteins / physiology
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Repressor Proteins / physiology
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Saccharomyces cerevisiae / cytology
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Saccharomyces cerevisiae / genetics*
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Saccharomyces cerevisiae Proteins / physiology*
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Shelterin Complex
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Telomerase / physiology*
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Telomere / metabolism
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Telomere / ultrastructure*
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Telomere-Binding Proteins / deficiency
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Telomere-Binding Proteins / physiology*
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Transcription Factors / physiology*
Substances
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Carrier Proteins
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Chromatin
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DNA, Fungal
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DNA-Binding Proteins
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RAP1 protein, S cerevisiae
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RIF2 protein, S cerevisiae
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RNA, recombinant
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Recombinant Fusion Proteins
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Repressor Proteins
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Saccharomyces cerevisiae Proteins
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Shelterin Complex
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Telomere-Binding Proteins
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Transcription Factors
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telomerase RNA
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RIF1 protein, S cerevisiae
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RNA
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EST2 protein, S cerevisiae
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Telomerase
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PIF1 protein, S cerevisiae
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DNA Helicases