Figure 3. Round- and bent-shaped mutant cells maintain wildtype-like cell polarity when constrained inside straight channels – a feedback loop.

A. Top, A straight microfluidic channel containing orb6^ts mutant cells growing at the restrictive 36°C temperature. Bottom, orb6^ts cells expressing mCherry-atb2p (tubulin) and tea1p-GFP (+TIP) (PT.893). Outside the straight channel, orb6^ts cells have lost their bipodal polarity and are round with microtubules oriented in all directions and with +TIP protein tea1p delocalized (observed in ~87% of control cells; n=30). Inside the channel, orb6^ts cells are forced to maintain a rod-shape with microtubules maintaining their alignment along the cell long-axis and +TIP protein tea1p specifically localized to the growing cell tips (observed in 60% of cells; n=15). Bar, 10 μm.

B. Top, a straight microfluidic channel containing mto1Δ mutant cells. Bottom, mto1Δ cells expressing mCherry-atb2p (tubulin) and tea1p-GFP (+TIP) (PT.910). Outside the channel, the single microtubule bundle is located at the convex surface of the bent mto1Δ cell. The +TIP protein tea1p shows asymmetry in its localization at the old cell tips and also begins to localize ectopically at a new cortical site (yellow arrow). Inside the channel, the microtubule bundle of mto1Δ cells shows no preference for a particular cell surface, but instead interacts with the bipodal cell tips. As a consequence, cell tip proteins show symmetrical localization at the cell tips and not at other ectopic sites (observed in ~88% of cells, n=17; control cells n=17). Bar, 10 μm.

C. Top, mto1Δ cells from (B). We defined the Length Ratio as the ratio of the width of the tea1-GFP (or bud6-GFP) signal divided by the width of the cell tip. Bottom, comparison plots of Length Ratios of tea1-GFP and bud6-GFP in mto1Δ cells growing outside versus inside the straight channels.