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tra12155-sup-0001-TableS1.pdfPDF document101KTable S1: Yeast strains used in this study.
tra12155-sup-0002-FigureS1.pdfPDF document4150KFigure S1: Halo assays to demonstrate effect of pheromone on growth arrest in wild type and apm4 null strains. KAY120 and KAY1690 were grown to lawns in the presence of discs supplemented with 1, 2 and 5 µg α-factor. Cells were grown on YPD as described.
tra12155-sup-0003-FigureS2.pdfPDF document4150KFigure S2: Mid2-GFP levels at the plasma membrane are higher in apm4 null cells. After 30 min of exposure to pheromone the fluorescence intensity of plasma membrane staining was analysed in wild type and apm4Δ cells as a measure of the level of Mid2-GFP. Intensities were measured at two points per cell where a line drawn perpendicular to the axis of polarized growth assessed morphologically, and through the centre of the cell, bisected the plasma membrane. Data from three independent experiments and analysis of >100 cells total. In a Student t-test significance p value <0.0001. Mean wt 380.1 ± 14.24 SEM, n = 103; apm4Δ mean 637.9 ± 25.73, n = 103.
tra12155-sup-0004-FigureS3.pdfPDF document9288KFigure S3: The effect on Apm4 binding and Mid2 localization of a Y278 mutation in the Mid2 cytoplasmic tail. A) GST alone; GST-tagged Apm4; His-tagged Mid2 or Mid2 Y278 were prepared as described and their binding interaction tested using pull down assays on beads with 20 µm Mid2. Apm4 is clearly seen on a Generon NUView gel, along with a faint band for Mid2 (arrow). Mid2 was detected using anti-His tag antibodies after western blotting. This band was verified as Mid2 using mass spectrometry. B) Mid2-GFP and Mid2-Y278A-GFP were transformed into cells otherwise lacking mid2 (KAY53) or both mid2 and apm4 (KAY1700). Localization after 1 h incubation with alpha factor was observed microscopically.
tra12155-sup-0005-FigureS4.pdfPDF document9288KFigure S4: Effect of pheromone on organization of actin patches in wild type and apm4Δ cells. Abp1-GFP was transformed into KAY120 and KAY1690 cells and visualized in live cells 90 min after pheromone addition.
tra12155-sup-0006-AppendixS1.pdfPDF document13377KAppendix S1. Supplementary materials and methods.

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