title: Actin dynamics and competition for myosin monomer govern the sequential amplification of myosin filaments.
abstract: The cellular mechanisms governing non-muscle myosin II (NM2) filament assembly are largely unknown. Using EGFP-NM2A knock-in fibroblasts and multiple super-resolution imaging modalities, we characterized and quantified the sequential amplification of NM2 filaments within lamellae, wherein filaments emanating from single nucleation events continuously partition, forming filament clusters that populate large-scale actomyosin structures deeper in the cell. Individual partitioning events coincide spatially and temporally with the movements of diverging actin fibres, suppression of which inhibits partitioning. These and other data indicate that NM2A filaments are partitioned by the dynamic movements of actin fibres to which they are bound. Finally, we showed that partition frequency and filament growth rate in the lamella depend on MLCK, and that MLCK is competing with centrally active ROCK for a limiting pool of monomer with which to drive lamellar filament assembly. Together, our results provide new insights into the mechanism and spatio-temporal regulation of NM2 filament assembly in cells.
authors.affiliation: Cell Biology and Physiology Center, National Heart, Lung and Blood Institute, National Institutes of Health Bethesda, Maryland 20892, USA.
authors.forename: Jordan R
authors.initials: JR
authors.lastname: Beach
authors.affiliation: Cell Biology and Physiology Center, National Heart, Lung and Blood Institute, National Institutes of Health Bethesda, Maryland 20892, USA.
authors.forename: Kyle S
authors.initials: KS
authors.lastname: Bruun
authors.affiliation: Howard Hughes Medical Institute, Janelia Research Campus, Ashburn, Virginia 20147, USA.
authors.forename: Lin
authors.initials: L
authors.lastname: Shao
authors.affiliation: Howard Hughes Medical Institute, Janelia Research Campus, Ashburn, Virginia 20147, USA.
authors.forename: Dong
authors.initials: D
authors.lastname: Li
authors.affiliation: Cell Biology and Physiology Center, National Heart, Lung and Blood Institute, National Institutes of Health Bethesda, Maryland 20892, USA.
authors.forename: Zac
authors.initials: Z
authors.lastname: Swider
authors.affiliation: Cell Biology and Physiology Center, National Heart, Lung and Blood Institute, National Institutes of Health Bethesda, Maryland 20892, USA.
authors.forename: Kirsten
authors.initials: K
authors.lastname: Remmert
authors.affiliation: Laboratory of Molecular Cardiology, National Heart, Lung and Blood Institute, National Institutes of Health Bethesda, Maryland 20892, USA.
authors.forename: Yingfan
authors.initials: Y
authors.lastname: Zhang
authors.affiliation: Laboratory of Molecular Cardiology, National Heart, Lung and Blood Institute, National Institutes of Health Bethesda, Maryland 20892, USA.
authors.forename: Mary A
authors.initials: MA
authors.lastname: Conti
authors.affiliation: Laboratory of Molecular Cardiology, National Heart, Lung and Blood Institute, National Institutes of Health Bethesda, Maryland 20892, USA.
authors.forename: Robert S
authors.initials: RS
authors.lastname: Adelstein
authors.affiliation: Cell Biology and Physiology Center, National Heart, Lung and Blood Institute, National Institutes of Health Bethesda, Maryland 20892, USA.
authors.forename: Nasser M
authors.initials: NM
authors.lastname: Rusan
authors.affiliation: Howard Hughes Medical Institute, Janelia Research Campus, Ashburn, Virginia 20147, USA.
authors.forename: Eric
authors.initials: E
authors.lastname: Betzig
authors.affiliation: Cell Biology and Physiology Center, National Heart, Lung and Blood Institute, National Institutes of Health Bethesda, Maryland 20892, USA.
authors.forename: John A
authors.initials: JA
authors.lastname: Hammer
mesh_terms: D000199:Actins / Q000378:metabolism*; D000205:Actomyosin / Q000378:metabolism; D000818:Animals; D003599:Cytoskeleton / Q000378:metabolism*; D005347:Fibroblasts / Q000378:metabolism*; D055879:Gene Knock-In Techniques; D051379:Mice; D018994:Myosin Light Chains / Q000235:genetics*; D009219:Myosin-Light-Chain Kinase / Q000378:metabolism*; D009218:Myosins / Q000378:metabolism*; D010446:Peptide Fragments / Q000378:metabolism*
chemical_list: D000199:Actins; C524495:Mlc2a protein, mouse; D018994:Myosin Light Chains; D010446:Peptide Fragments; C090954:SM 1 peptide; D000205:Actomyosin; D009219:Myosin-Light-Chain Kinase; D009218:Myosins