Myosin II regulates the shape of three-dimensional intestinal epithelial cysts.
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
Andrei I
AI
Epithelial Pathobiology Research Unit, Department of Pathology and Laboratory Medicine, Emory University, Atlanta, GA 30322, USA. Andrei_Ivanov@urmc.rochester.edu
Hopkins
Ann M
AM
Brown
G Thomas
GT
Gerner-Smidt
Kirsten
K
Babbin
Brian A
BA
Parkos
Charles A
CA
Nusrat
Asma
A
D008841:Actin Cytoskeleton / Q000378:metabolism*; D000199:Actins / Q000378:metabolism; D018938:Caco-2 Cells; D016764:Cell Polarity / Q000502:physiology; D048430:Cell Shape / Q000187:drug effects / Q000502:physiology*; D051338:Cofilin 1 / Q000378:metabolism; D015536:Down-Regulation / Q000187:drug effects / Q000502:physiology; D004847:Epithelial Cells / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D006576:Heterocyclic Compounds, 4 or More Rings / Q000494:pharmacology; D006801:Humans; D007413:Intestinal Mucosa / Q000166:cytology / Q000196:embryology* / Q000378:metabolism*; D024462:Myosin Type II / Q000235:genetics / Q000378:metabolism*; D015739:Nocodazole / Q000494:pharmacology; D009924:Organ Culture Techniques; D011108:Polymers / Q000378:metabolism; D020871:RNA Stability; D010738:Type C Phospholipases / Q000378:metabolism
D000199:Actins; D051338:Cofilin 1; D006576:Heterocyclic Compounds, 4 or More Rings; D011108:Polymers; C472645:blebbistatin; D010738:Type C Phospholipases; D024462:Myosin Type II; D015739:Nocodazole