The yeast telomerase RNA, TLC1, participates in two distinct modes of TLC1-TLC1 association processes in vivo

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Introduction

Materials and Methods

Plasmids

Yeast strains and growth media

Immunoprecipitation of MS2 hairpin-tagged TLC1

Immunoprecipitation of tagged proteins

Quantitative reverse transcription and PCR (qRT-PCR)

Calculation of fraction TLC1 in dimer form

Telomere length analysis

Results

Co-immunoprecipitation assays demonstrate TLC1-TLC1 association in vivo

The 3′ region of TLC1 is important for TLC1-TLC1 association

TLC1-TLC1 association is dependent on nuclear export and is cell cycle-regulated

Telomerase holoenzyme formation is not required for TLC1-TLC1 association

Ku and Sir4, but not telomere silencing or tethering to the nuclear periphery, promote the same mode of TLC1-TLC1 association

Ku/Sir4 and the 3′-cleaved TLC1 precursor sequence promote TLC1-TLC1 association by different modes

Lack of evidence for Est2-Est2 physical association

Discussion

Supplemental Information

Additional Information and Declarations

Competing Interests

The authors declare there are no competing interests.

Author Contributions

Tet Matsuguchi conceived and designed the experiments, performed the experiments, analyzed the data, wrote the paper, prepared figures and/or tables, reviewed drafts of the paper.

Elizabeth Blackburn conceived and designed the experiments, contributed reagents/materials/analysis tools, wrote the paper, reviewed drafts of the paper.

Data Availability

The following information was supplied regarding data availability:

The raw data can be found in the Supplemental Information.

Funding

This work was supported by an NIH grant (R01GM026259). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.

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