Figure 5.
Figure 5. A nonphosphorylatable IRSp53 mutant rescues the Par1b phenotype. (A) Parental MDCK cells and five clones expressing RNAi-resistant IRSp53-A4 and four clones of IRSp53-WT cells were transiently transfected with mock or IRSp53shmir cDNA and analyzed for cell spreading 2 h after plating. (B) 2-h spreading in parental cells, eight IRSp53AA, and five IRSp53WT clones that were transduced with control or Par1b adenovirus. (C) Paxillin-mCherry in WT clone #1 and A4 clone #5 expressing Par1b-GFP 30 min after plating on collagen I. Arrowheads point to FAs (Video 9, bottom). (D) Par1b-MDCK cells transfected with mock or IRSp53 cDNAs (WT, A4, DD, and ΔSH3) were quantitatively analyzed for lumen polarity as described in Materials and methods. *, P < 0.0001 with difference to parental or mock-transfected cells. Error bars indicate SEM. Bars: (A and B) 30 µm; (C and D) 10 µm.

A nonphosphorylatable IRSp53 mutant rescues the Par1b phenotype. (A) Parental MDCK cells and five clones expressing RNAi-resistant IRSp53-A4 and four clones of IRSp53-WT cells were transiently transfected with mock or IRSp53shmir cDNA and analyzed for cell spreading 2 h after plating. (B) 2-h spreading in parental cells, eight IRSp53AA, and five IRSp53WT clones that were transduced with control or Par1b adenovirus. (C) Paxillin-mCherry in WT clone #1 and A4 clone #5 expressing Par1b-GFP 30 min after plating on collagen I. Arrowheads point to FAs (Video 9, bottom). (D) Par1b-MDCK cells transfected with mock or IRSp53 cDNAs (WT, A4, DD, and ΔSH3) were quantitatively analyzed for lumen polarity as described in Materials and methods. *, P < 0.0001 with difference to parental or mock-transfected cells. Error bars indicate SEM. Bars: (A and B) 30 µm; (C and D) 10 µm.

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