title: A new role for myosin II in vesicle fission.
abstract: An endocytic vesicle is formed from a flat plasma membrane patch by a sequential process of invagination, bud formation and fission. The scission step requires the formation of a tubular membrane neck (the fission pore) that connects the endocytic vesicle with the plasma membrane. Progress in vesicle fission can be measured by the formation and closure of the fission pore. Live-cell imaging and sensitive biophysical measurements have provided various glimpses into the structure and behaviour of the fission pore. In the present study, the role of non-muscle myosin II (NM-2) in vesicle fission was tested by analyzing the kinetics of the fission pore with perforated-patch clamp capacitance measurements to detect single vesicle endocytosis with millisecond time resolution in peritoneal mast cells. Blebbistatin, a specific inhibitor of NM-2, dramatically increased the duration of the fission pore and also prevented closure during large endocytic events. Using the fluorescent markers FM1-43 and pHrodo Green dextran, we found that NM-2 inhibition greatly arrested vesicle fission in a late phase of the scission event when the pore reached a final diameter of ∼ 5 nm. Our results indicate that loss of the ATPase activity of myosin II drastically reduces the efficiency of membrane scission by making vesicle closure incomplete and suggest that NM-2 might be especially relevant in vesicle fission during compound endocytosis.
authors: Dpto. Fisiología Médica y Biofísica, Facultad de Medicina, Universidad de Sevilla, Seville, Spain.
authors: Juan A
authors: JA
authors: Flores
authors: Dpto. Fisiología Médica y Biofísica, Facultad de Medicina, Universidad de Sevilla, Seville, Spain.
authors: Santiago
authors: S
authors: Balseiro-Gomez
authors: Dpto. Fisiología Médica y Biofísica, Facultad de Medicina, Universidad de Sevilla, Seville, Spain.
authors: Jose M
authors: JM
authors: Cabeza
authors: Dpto. Fisiología Médica y Biofísica, Facultad de Medicina, Universidad de Sevilla, Seville, Spain.
authors: Jorge
authors: J
authors: Acosta
authors: Dpto. Fisiología Médica y Biofísica, Facultad de Medicina, Universidad de Sevilla, Seville, Spain.
authors: Pilar
authors: P
authors: Ramirez-Ponce
authors: Dpto. Fisiología Médica y Biofísica, Facultad de Medicina, Universidad de Sevilla, Seville, Spain.
authors: Eva
authors: E
authors: Ales
mesh_terms: D000818:Animals; D002462:Cell Membrane / Q000187:drug effects / Q000378:metabolism; D004705:Endocytosis / Q000187:drug effects / Q000502:physiology; D005089:Exocytosis / Q000187:drug effects / Q000502:physiology; D006576:Heterocyclic Compounds, 4 or More Rings / Q000494:pharmacology; D008407:Mast Cells / Q000187:drug effects / Q000378:metabolism; D051379:Mice; D024462:Myosin Type II / Q000037:antagonists & inhibitors / Q000378:metabolism*; D011485:Protein Binding; D011726:Pyridinium Compounds / Q000378:metabolism; D000644:Quaternary Ammonium Compounds / Q000378:metabolism; D022161:Transport Vesicles / Q000187:drug effects / Q000378:metabolism*
chemical_list: C073804:FM1 43; D006576:Heterocyclic Compounds, 4 or More Rings; D011726:Pyridinium Compounds; D000644:Quaternary Ammonium Compounds; C472645:blebbistatin; D024462:Myosin Type II