JCS015842 Supplementary Material

Files in this Data Supplement:

• Supplemental Figure S1 -

  Fig. S1. Analogous morphology of Caco-2 cysts formed after 72 hours in the 3D Matrigel and Caco-2 colonies growing for the same time on Matrigel-coated coverslips.

• Supplemental Figure S2 -

  Fig. S2. Phospholipase C and microtubules are involved in the formation of surface protrusions in NMMIIA-depleted cysts. SK-CO15 cells were transfected with NMMIIA-specific siRNA, embedded into 3D Matrigel and 72 hours later were treated for 12 hours with either vehicle or PLC inhibitor U-73122 (1 µM) or microtubule-depolymerizing drug vinblastin (20 µM). Inhibition of PLC and microtubule depolymerization prevented the formation of peripheral F-actin-rich protrusions in NMMIIA-depleted cysts (*P<0.01).

• Supplemental Figure S3 -

  Fig. S3. The involvement of microtubule plus ends in the formation of blebbistatin-induced surface protrusions. Preformed Caco-2 cysts were treated for 12 hours with either blebbistatin alone or in combination with a low concentration of nocodazole (100 nM). Inhibition of microtubule plus-end dynamics with nocodazole prevented formation of peripheral F-actin-rich protrusions in blebbistatin-treated cysts (*P<0.01).

• Supplemental Figure S4 -

  Fig. S4. Stabilization of microtubules in blebbistatin-treated epithelial cells. Caco-2 cells were treated with either blebbistatin or vehicle, and the amount of stable and unstable microtubules determined by Triton X-100 fractionation and western blotting. A significant increase in the amount of Triton-insoluble (stable) microtubules was observed after inhibition of myosin II (*P<0.05).