1. TOR signaling regulates microtubule structure and function
Jae H. Choi, Neil R. Adames, Ting-Fung Chan, Chenbo Zeng, John A. Cooper, X.F.Steven Zheng 
Current Biology 
Volume 10, Issue 14, Pages (July 2000)
DOI: /S (00)

2. Figure 1
The rapamycin sensitivity of yeast mutants defective in kinesin-related proteins (KRPs) and interaction between Tor1p and Bik1p. (a,b) The rapamycin sensitivity of cin8Δ, kip2Δ and bik1Δ/pac14Δ mutant strains. The isogenic haploid strains indicated were grown in YPD medium with (right panels) and without (left panels) 10 nM rapamycin. Each spot represents a tenfold dilution of the culture. (c) Tor1p interacts with Bik1p in vivo. The left panel shows binding of Tor1p–Myc9 to Bik1p–ProA5. Lysates of yeast cells expressing Tor1p–Myc9 and Bik1p–ProA5 or Bik1p were purified on Ig–Sepharose beads, and analyzed by western blotting. The right panel shows binding of Bik1p–Myc9 to Tor1p–ProA5. T, total lysate; IP, the IG–Sepharose-bound materials. The sample in each T lane represents 10% of that in the corresponding IP lane.
Current Biology  , DOI: ( /S (00) )

3. Figure 2
Rapamycin causes defects in microtubule stability, spindle elongation and spindle position. (a) Morphological abnormalities after rapamycin treatment. Log-phase cells carrying a wild-type (pTOR1WT) or dominant rapamycin-resistant TOR1 (pTOR1S1972I) were treated without any drug (control), or with 100 nM rapamycin or FK506 at 30°C for 20 min. In (a,c) the panels labeled DAPI have been stained to show the nucleus; the panels labeled TUB have been stained with anti-tubulin antibody to show microtubules. (b) Reduction of pre-anaphase spindle length and anaphase spindle elongation kinetics by rapamycin. (c) Rapamycin destabilizes the existing spindles. The cdc2, cdc13 and cdc15 ts mutants were arrested at S, G2 and M, respectively, at 36°C for 3 h. The arrested cells were then treated with 200 nM rapamycin, 200 nM FK506, or 50 mg/ml cycloheximide (+ CHX). (d) Western analysis of in vitro polymerized microtubules. Rapamycin interferes with microtubule polymerization. High-speed extracts from yeast cells treated without (lane 1) or with 200 nM rapamycin for 20 min (lane 2) were incubated without or with GTP and taxol (lanes 3–6). The results were analyzed by western blotting with an anti-tubulin antibody.
Current Biology  , DOI: ( /S (00) )

4. Figure 3
Rapamycin treatment causes defects in nuclear migration, spindle orientation and spindle position. (a,b) Rapamycin causes defects in nuclear segregation and position. Rapamycin (2 nM) was added to the early exponential wild-type yeast culture and incubated for two generations. In (a) bright-field images were overlain with DAPI images to show the positions of nuclear DNA. Binucleate cells, indicating abnormal segregation, are indicated by arrows. The percentages of cells with aberrant nuclear position are given in (b). For each strain, the table shows the total cells counted (n), the percentages of cells at the different bud sizes, and the percentages of cells with the specified abnormalities relative to total cells counted. (c) Rapamycin affects nuclear migration to the neck. Cells were arrested with a factor, released into fresh media with or without 2 nM rapamycin for 30 min, fixed, and stained with DAPI. The distance between the neck and the nearest edge of DAPI staining was measured and divided by the distance between the neck and the distal edge of the mother cortex to derive the nucleus migration index. The scale along the bottom of the histograms is in increments of 0.1. (d) Analysis of spindle orientations in wild-type cells expressing GFP–tubulin, with and without rapamycin. All measurements were done on pre-anaphase spindles at 1 min before onset of anaphase B. (e) Analysis of spindle positions in wild-type cells expressing GFP–tubulin, with or without rapamycin.
Current Biology  , DOI: ( /S (00) )