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. 2013 Jun 19;41(16):7713–7724. doi: 10.1093/nar/gkt514

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© The Author(s) 2013. Published by Oxford University Press.

This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/3.0/), which permits unrestricted reuse, distribution, and reproduction in any medium, provided the original work is properly cited.

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Figure 3. — Characterization of Est1 function in conjunction with TLC1 alleles. (A) Top: FISH assay detecting the Tlc1 RNA in WT, tlc1Δ, est1Δ strains or strains that harbor the various TLC1 alleles. The Tlc1 RNA in all cells with different TLC1 alleles display a similar nuclear distribution. Each image is a representation of one cell from the corresponding strain and the signals from both DAPI (blue, DNA) and Cy3 (green, Tlc1) channels were merged. Scale bar = 500 nm. Bottom: graphical representation of cellular localization from 50 cells of each WT, est1Δ, ΔSL, ΔSL + IL SA5, SA3 and OPM strains. (B) Overexpression of Est1 does not recapitulate telomere elongation in the ΔSL, SA5 and SA3 mutants. Genomic DNA from NLYH80 (+ indicated TLC1 alleles) was extracted after 50 generations of growth in the following conditions: lanes 3–7: cells contain an empty pRS423, the complementing TLC1 plasmid (pAZ1) and the indicated variant alleles of TLC1. Lanes 8–12: cells contain pRS423-EST1 (2 µm plasmid for overexpression), the complementing TLC1 plasmid (pAZ1) and TLC1 variants. Lanes 13–17: cells contain pRS423-EST1 and only the TLC1 alleles as indicated. Lanes 2 and 18: yku70Δ (short telomere length control). Lanes 1 and 19 (M): radiolabeled 1 kb DNA ladder.