List of yeast strains.

<p>List of yeast strains.</p

<i>BUD2</i> is required for the spindle position checkpoint.

<p><i>arp1Δ GFP-TUB1</i> cells with the additional indicated mutations were assayed for checkpoint integrity by video analysis. Cells with long (late-anaphase) spindles in the mother of a budded cell were followed over time. Checkpoint integrity is the percent of cells in which the spindle that remained intact, i.e. did not break down, for a time greater than the mean plus two standard deviations of the time for normal mitotic exit. A. <i>bud2Δ</i> mutants have a defect in the spindle position checkpoint, with failure to maintain arrest in about half of cells. <i>bub2Δ</i> is a positive control known to have a complete defect. The <i>bud2Δ</i> phenotype does not depend on <i>LTE1</i>, based on the <i>bud2Δ lte1Δ</i> double mutant. <i>BUD6</i> is in a pathway upstream of <i>LTE1</i>, as described previously <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0036127#pone.0036127-Nelson1" target="_blank">[2]</a> and confirmed here. The <i>bud2 bud6</i> double mutant has an exacerbated phenotype, confirming that <i>BUD2</i> is in a genetic pathway independent of <i>BUD6</i> and <i>LTE1</i>. The <i>bud2Δ bud6Δ</i> double mutant does not have a complete loss of phenotype, as <i>bub2</i> does, suggesting a possible third input into the checkpoint control of mitotic exit. B. The bud-site-selection pathway has no role in the spindle position checkpoint. Mutants lacking either Rsr1/Bud1, the only known substrate of Bud2, or Bud5, the GEF for Rsr1/Bud1, have no checkpoint defect. Deleting <i>RSR1/BUD1</i> does not suppress the checkpoint defect of a <i>bud2Δ</i> mutant. <i>bud3</i> and <i>bud5</i> null mutants, defective in axial and all budding patterns, respectively, are also normal.</p

Musculature of the mouse intestinal wall.

<p>(A) A frozen section of fluorescently labelled small intestine visualised by widefield fluorescence microscopy (red shows F-actin, green shows laminin, blue shows nuclei). (B–E) Fluorescently stained wholemounts of small and large intestine, visualised using confocal fluorescence microscopy (red shows F-actin, blue shows nuclei). (A) A section through the wall of the small intestine. Nuclei (blue) are stained with DAPI, F-actin (red) highlights the smooth muscle cells and apical surface of the gut epithelium. The gut epithelium is continuous from the crypts (C) and over the villi (V). Basement membranes (green) are stained with an anti-laminin antibody, and the basal surfaces of gut epithelial cells are directly attached to the basement membrane (arrowhead). The muscularis externa is composed of outer longitudinal and inner circularly oriented smooth muscle fibres. The muscularis mucosae (just above the dashed line) closely follows the crypt bases, with some fibres extending up into the villi (dashed arrow). Pericryptal fibroblasts surround the crypt epithelium (arrows). (B) A longitudinal section through the small intestine shows the base of crypts (outlined by dashed circles). Smooth muscle fibres of the muscularis mucosae are oriented parallel (arrows) to the longitudinal muscle. (C) A transverse section of small intestine shows the muscularis mucosae (arrow). It is comprised of a single cell layer that forms an incomplete layer or meshwork beneath crypts, just above the circular muscle (CM). (D) A longitudinal section through the colon shows the outer (solid arrow) and inner (dashed arrow) more disorganised layers of the muscularis mucosae. The outer layer is oriented parallel to the longitudinal muscle and the inner layer parallel with the circular muscle, as indicated by double headed arrows. (E) A transverse section through the colon shows the muscularis mucosae (arrow) at the base of the crypts, which is much thicker than that found in the small intestine. Scale bars = .</p

A comparison of model results with cell area data for three healthy murine colonic epithelium samples.

<p>(a) Experimental data obtained from three wildtype murine tissue samples, taken from midway down the length of the colon. (b) A histogram of cross-sectional area data collected from the simulation of 1000 hours. These data have been dimensionalised, scaling 1 cell width to , and the frequency has been averaged over total cell number to show the proportion of cells in each bin.</p

Cell division and death statistics for the cross-sectional model.

<p>(a) The spatial distribution of anoikis events along the -axis, (c) the spatial distribution of division events along the -axis.</p

Defining the location of the basement membrane.

<p>The red dashed line indicates the location of the basement membrane, which is defined to pass through the midpoints of the springs connecting neighbouring epithelial and stromal cell centres. The local discrete curvature is calculated for each epithelial and stromal node pair, and the midpoints of the neighbouring springs are taken to form a piecewise linear curve defined by three points. An example is indicated by the three points marked by black circles.</p

The spatial distribution of anoikis and division events that occur in the epithelial layer when density-dependent inhibition of mitosis and random cell death are implemented.

<p>(a) Anoikis events, (b) division events, (c) epithelial cell locations at the final timestep, (d) six typical migratory tracks for epithelial cells in the monolayer (showing the -coordinates only).</p

Model parameters.

<p>Unless stated otherwise, the parameters in the model assume the values given in this table. The distances in the model have been scaled with cell width (<a href="http://www.ploscompbiol.org/article/info:doi/10.1371/journal.pcbi.1002515#pcbi.1002515-Alberts1" target="_blank">[35]</a>, <a href="http://www.ploscompbiol.org/article/info:doi/10.1371/journal.pcbi.1002515#pcbi.1002515-Smallwood1" target="_blank">[39]</a>), time is measured in hours, and all other variables have been scaled so that (see Equation (2)) <a href="http://www.ploscompbiol.org/article/info:doi/10.1371/journal.pcbi.1002515#pcbi.1002515-VanLeeuwen2" target="_blank">[15]</a>, <a href="http://www.ploscompbiol.org/article/info:doi/10.1371/journal.pcbi.1002515#pcbi.1002515-Meineke1" target="_blank">[16]</a>.</p

A comparison of the final state of the epithelial layer for the three cases considered.

<p>The direction of the arrows indicate the extension of the epithelial layer when random cell death is included at the edges (blue curve), compared to the two cases where this second form of apoptosis is not present.</p

The spatial distribution of anoikis and division events that occur in the epithelial layer when density-dependent inhibition of mitosis is implemented.

<p>(a) Anoikis events, (b) division events, (c) epithelial cell locations at the final timestep, (d) six typical migratory tracks for epithelial cells in the monolayer (showing the -coordinates only).</p