title: Fluorescent analogues of myosin II for tracking the behavior of different myosin isoforms in living cells.
abstract: Fluorescently labeled smooth muscle myosin II is often used to study myosin II dynamics in non-muscle cells. In order to provide more specific tools for tracking non-muscle myosin II in living cytoplasm, fluorescent analogues of non-muscle myosin IIA and IIB were prepared and characterized. In addition, smooth and non-muscle myosin II were labeled with both cy5 and rhodamine so that comparative, dynamic studies may be performed. Non-muscle myosin IIA was purified from bovine platelets, non-muscle myosin IIB from bovine brain, and smooth muscle myosin II from turkey gizzards. After being fluorescently labeled with tetramethylrhodamine-5-iodoacetamide or with a succinimidyl ester of cy5, they retained the following properties: (1) reversible assembly into thick filaments, (2) actin-activatable MgATPase, (3) phosphorylation by myosin light chain kinase, (4) increased MgATPase upon light-chain phosphorylation, (5) interconversion between 6S and 10S conformations, and (6) distribution into endogenous myosin II-containing structures when microinjected into cultured cells. These fluorescent analogues can be used to visualize isoform-specific dynamics of myosin II in living cells.
authors.affiliation: Department of Anatomy and Cell Biology, State University of New York at Buffalo School of Medicine and Biomedical Sciences, 14214, USA. jkolega@ubmedb.buffalo.edu
authors.forename: J
authors.initials: J
authors.lastname: Kolega
mesh_terms: D000818:Animals; D001792:Blood Platelets / Q000737:chemistry; D001921:Brain / Q000378:metabolism; D002232:Carbocyanines; D002478:Cells, Cultured; D005456:Fluorescent Dyes; D009130:Muscle, Smooth / Q000737:chemistry; D009218:Myosins / Q000737:chemistry / Q000145:classification / Q000378:metabolism*; D012235:Rhodamines
chemical_list: D002232:Carbocyanines; D005456:Fluorescent Dyes; D012235:Rhodamines; C085321:cyanine dye 5; D009218:Myosins