Actins, Caco-2 Cells, Cell Polarity, Cell Shape, Cofilin 1, Down-Regulation, Epithelial Cells, Heterocyclic Compounds with 4 or More Rings, Humans, Intestinal Mucosa, Microfilaments, Myosin Type II, Nocodazole, Organ Culture Techniques, Polymers, RNA Stability, Type C Phospholipases
The development of luminal organs begins with the formation of spherical cysts composed of a single layer of epithelial cells. Using a model three-dimensional cell culture, this study examines the role of a cytoskeletal motor, myosin II, in cyst formation. Caco-2 and SK-CO15 intestinal epithelial cells were embedded into Matrigel, and myosin II was inhibited by blebbistatin or siRNA-mediated knockdown. Whereas control cells formed spherical cysts with a smooth surface, inhibition of myosin II induced the outgrowth of F-actin-rich surface protrusions. The development of these protrusions was abrogated after inhibition of F-actin polymerization or of phospholipase C (PLC) activity, as well as after overexpression of a dominant-negative ADF/cofilin. Surface protrusions were enriched in microtubules and their formation was prevented by microtubule depolymerization. Myosin II inhibition caused a loss of peripheral F-actin bundles and a submembranous extension of cortical microtubules. Our findings suggest that inhibition of myosin II eliminates the cortical F-actin barrier, allowing microtubules to reach and activate PLC at the plasma membrane. PLC-dependent stimulation of ADF/cofilin creates actin-filament barbed ends and promotes the outgrowth of F-actin-rich protrusions. We conclude that myosin II regulates the spherical shape of epithelial cysts by controlling actin polymerization at the cyst surface.
Ivanov AI, Hopkins AM, Brown GT, Gerner-Smidt K, Babbin BA, Parkos CA, Nusrat A. Myosin II regulates the shape of three-dimensional intestinal epithelial cysts. Journal of Cell Science. 2008;121(Pt 11):1803-14.