An in vitro assay for saccharomyces telomerase requires EST1

Jing Jer Lin; Virginia A. Zakian

doi:10.1016/S0092-8674(05)80017-0

An in vitro assay for saccharomyces telomerase requires EST1

Jing Jer Lin, Virginia A. Zakian

Molecular Biology

Research output: Contribution to journal › Article › peer-review

102 Scopus citations

Abstract

Telomerase activity was demonstrated in cell-free extracts from S. cerevisiae through the use of a PCR-based assay. As expected, this activity was eliminated by RNase or phenol treatment of the extract and was dependent on dGTP and dTTP. Telomerase was not detected in extracts prepared from cells grown for ∼ 30 or more cell divisions in the absence of the EST1 product, Est1 p. TLC1 RNA, which determines the sequence of telomeric DNA in vivo, was present in normal amounts in est1Δ cells. Moreover, TLC1 RNA specifically precipitated with epitope-tagged Est1p. These data indicate that Estlp is either a subunit of yeast telomerase or an accessory protein associated with telomerase that is essential in vitro for its activity.

Original language	English (US)
Pages (from-to)	1127-1135
Number of pages	9
Journal	Cell
Volume	81
Issue number	7
DOIs	https://doi.org/10.1016/S0092-8674(05)80017-0
State	Published - Jun 30 1995

All Science Journal Classification (ASJC) codes

General Biochemistry, Genetics and Molecular Biology

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10.1016/S0092-8674(05)80017-0

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@article{74e0c58262f34f1cbce621c79d9bf211,

title = "An in vitro assay for saccharomyces telomerase requires EST1",

abstract = "Telomerase activity was demonstrated in cell-free extracts from S. cerevisiae through the use of a PCR-based assay. As expected, this activity was eliminated by RNase or phenol treatment of the extract and was dependent on dGTP and dTTP. Telomerase was not detected in extracts prepared from cells grown for ∼ 30 or more cell divisions in the absence of the EST1 product, Est1 p. TLC1 RNA, which determines the sequence of telomeric DNA in vivo, was present in normal amounts in est1Δ cells. Moreover, TLC1 RNA specifically precipitated with epitope-tagged Est1p. These data indicate that Estlp is either a subunit of yeast telomerase or an accessory protein associated with telomerase that is essential in vitro for its activity.",

author = "Lin, {Jing Jer} and Zakian, {Virginia A.}",

note = "Funding Information: Correspondence should be addressed to V. A. Z. at Princeton University. We thank J. W. Shay for communicating results prior to publication; M. A. Piatyszek, J. Cooper, and S. Hahn for useful technical discussions; S. EIledge and S. Balakumaran for supplying plasmids prior to publication; and V. Schulz and J. Stavenhagen for their comments on the manuscript. This work was supported by National Institutes of Health grant GM26938 to V. A. Z. and by a Damon Runyon-Walter Winchell Cancer Research Fund fellowship (DRG-1205) to J.-J. L.",

year = "1995",

month = jun,

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doi = "10.1016/S0092-8674(05)80017-0",

language = "English (US)",

volume = "81",

pages = "1127--1135",

journal = "Cell",

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T1 - An in vitro assay for saccharomyces telomerase requires EST1

AU - Lin, Jing Jer

AU - Zakian, Virginia A.

N1 - Funding Information: Correspondence should be addressed to V. A. Z. at Princeton University. We thank J. W. Shay for communicating results prior to publication; M. A. Piatyszek, J. Cooper, and S. Hahn for useful technical discussions; S. EIledge and S. Balakumaran for supplying plasmids prior to publication; and V. Schulz and J. Stavenhagen for their comments on the manuscript. This work was supported by National Institutes of Health grant GM26938 to V. A. Z. and by a Damon Runyon-Walter Winchell Cancer Research Fund fellowship (DRG-1205) to J.-J. L.

PY - 1995/6/30

Y1 - 1995/6/30

N2 - Telomerase activity was demonstrated in cell-free extracts from S. cerevisiae through the use of a PCR-based assay. As expected, this activity was eliminated by RNase or phenol treatment of the extract and was dependent on dGTP and dTTP. Telomerase was not detected in extracts prepared from cells grown for ∼ 30 or more cell divisions in the absence of the EST1 product, Est1 p. TLC1 RNA, which determines the sequence of telomeric DNA in vivo, was present in normal amounts in est1Δ cells. Moreover, TLC1 RNA specifically precipitated with epitope-tagged Est1p. These data indicate that Estlp is either a subunit of yeast telomerase or an accessory protein associated with telomerase that is essential in vitro for its activity.

AB - Telomerase activity was demonstrated in cell-free extracts from S. cerevisiae through the use of a PCR-based assay. As expected, this activity was eliminated by RNase or phenol treatment of the extract and was dependent on dGTP and dTTP. Telomerase was not detected in extracts prepared from cells grown for ∼ 30 or more cell divisions in the absence of the EST1 product, Est1 p. TLC1 RNA, which determines the sequence of telomeric DNA in vivo, was present in normal amounts in est1Δ cells. Moreover, TLC1 RNA specifically precipitated with epitope-tagged Est1p. These data indicate that Estlp is either a subunit of yeast telomerase or an accessory protein associated with telomerase that is essential in vitro for its activity.

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SN - 0092-8674

VL - 81

SP - 1127

EP - 1135

JO - Cell

JF - Cell

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ER -

An in vitro assay for saccharomyces telomerase requires EST1

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