The Arabidopsis FRA1 kinesin contributes to the organization of cellulose microfibrils through an unknown mechanism. The cortical localization of this kinesin during interphase raises the possibility that it transport...The Arabidopsis FRA1 kinesin contributes to the organization of cellulose microfibrils through an unknown mechanism. The cortical localization of this kinesin during interphase raises the possibility that it transports cell wallrelated cargoes along cortical microtubules that either directly or indirectly influence cellulose microfibril patterning. To determine whether FRA1 is an authentic motor protein, we combined bulk biochemical assays and single molecule fluorescence imaging to analyze the motor properties of recombinant, GFP-tagged FRA1 containing the motor and coiled-coil domains （designated as FRAI（707）-GFP）. We found that FRAI（707）-GFP binds to microtubules in an ATP-dependent manner and that its ATPase activity is dramatically stimulated by the presence of microtubules. Using single molecule studies, we found that FRAI（707）-GFP moves processively along microtubule tracks at a velocity of about 0.4 μm s-1. In addition, we found that FRAI（707）-GFP is a microtubule plus-end-directed motor and that it moves along microtubules as a dimer. Interestingly, our single molecule analysis shows that the processivity of FRAI（707）-GFP is at least twice the processivity of conventional kinesin, making FRA1 the most processive kinesin to date. Together, our data show that FRA1 is a bona fide motor protein that has the potential to drive Iong-distance transport of cargo along cortical microtubules.展开更多
The effect of sintering dispersed dispersion and nano-emulsion particles of high molecular weightpolytetrafluoroethylene(PTFE)on a substrate as a function of“melt”time and temperature is described.Folded chain singl...The effect of sintering dispersed dispersion and nano-emulsion particles of high molecular weightpolytetrafluoroethylene(PTFE)on a substrate as a function of“melt”time and temperature is described.Folded chain singlecrystals parallel to the substrate and as ribbons on-edge(with double striations),as well as bands,are produced for longersintering times;particle merger and diffusion of individual molecules,crystallizing as folded chain,single(or few)molecule,single crystals when“trapped”on the substrate by cooling occur for shorter sintering times.It is suggested the observedstructures develop with sintering time,in a mesomorphic melt.The structure of the nascent particles is also discussed.展开更多
Formins are conserved regulators of actin cytoskeletal organization and dynamics that have been impli- cated to be important for cell division and cell polarity. The mechanism by which diverse formins regulate actin d...Formins are conserved regulators of actin cytoskeletal organization and dynamics that have been impli- cated to be important for cell division and cell polarity. The mechanism by which diverse formins regulate actin dynamics in plants is still not well understood. Using in vitro single-molecule imaging technology, we directly observed that the FH1-FH2 domain of an Arabidopsis thaliana formin, AtFH14, processively at- taches to the barbed end of actin filaments as a dimer and slows their elongation rate by 90%. The attach- ment persistence of FH1-FH2 is concentration dependent. Furthermore, by use of the triple-color total internal reflection fluorescence microscopy, we found that ABP29, a barbed-end capping protein, com- petes with FH1-FH2 at the filament barbed end, where its binding is mutually exclusive with AtFH14. In the presence of different plant profilin isoforms, FH1-FH2 enhances filament elongation rates from about 10 to 42 times. Filaments buckle when FH1-FH2 is anchored specifically to cover slides, further indicating that AtFH 14 moves processively on the elongating barbed end. At high concentration, AtFH 14 bundles actin filaments randomly into antiparallel or parallel spindle-like structures; however, the FH1-FH2-mediated bundles become thinner and longer in the presence of plant profilins. This is the direct demonstration of a processive formin from plants. Our results also illuminate the molecular mechanism of AtFH14 in regulating actin dynamics via association with profilin.展开更多
文摘The Arabidopsis FRA1 kinesin contributes to the organization of cellulose microfibrils through an unknown mechanism. The cortical localization of this kinesin during interphase raises the possibility that it transports cell wallrelated cargoes along cortical microtubules that either directly or indirectly influence cellulose microfibril patterning. To determine whether FRA1 is an authentic motor protein, we combined bulk biochemical assays and single molecule fluorescence imaging to analyze the motor properties of recombinant, GFP-tagged FRA1 containing the motor and coiled-coil domains （designated as FRAI（707）-GFP）. We found that FRAI（707）-GFP binds to microtubules in an ATP-dependent manner and that its ATPase activity is dramatically stimulated by the presence of microtubules. Using single molecule studies, we found that FRAI（707）-GFP moves processively along microtubule tracks at a velocity of about 0.4 μm s-1. In addition, we found that FRAI（707）-GFP is a microtubule plus-end-directed motor and that it moves along microtubules as a dimer. Interestingly, our single molecule analysis shows that the processivity of FRAI（707）-GFP is at least twice the processivity of conventional kinesin, making FRA1 the most processive kinesin to date. Together, our data show that FRA1 is a bona fide motor protein that has the potential to drive Iong-distance transport of cargo along cortical microtubules.
文摘The effect of sintering dispersed dispersion and nano-emulsion particles of high molecular weightpolytetrafluoroethylene(PTFE)on a substrate as a function of“melt”time and temperature is described.Folded chain singlecrystals parallel to the substrate and as ribbons on-edge(with double striations),as well as bands,are produced for longersintering times;particle merger and diffusion of individual molecules,crystallizing as folded chain,single(or few)molecule,single crystals when“trapped”on the substrate by cooling occur for shorter sintering times.It is suggested the observedstructures develop with sintering time,in a mesomorphic melt.The structure of the nascent particles is also discussed.
文摘Formins are conserved regulators of actin cytoskeletal organization and dynamics that have been impli- cated to be important for cell division and cell polarity. The mechanism by which diverse formins regulate actin dynamics in plants is still not well understood. Using in vitro single-molecule imaging technology, we directly observed that the FH1-FH2 domain of an Arabidopsis thaliana formin, AtFH14, processively at- taches to the barbed end of actin filaments as a dimer and slows their elongation rate by 90%. The attach- ment persistence of FH1-FH2 is concentration dependent. Furthermore, by use of the triple-color total internal reflection fluorescence microscopy, we found that ABP29, a barbed-end capping protein, com- petes with FH1-FH2 at the filament barbed end, where its binding is mutually exclusive with AtFH14. In the presence of different plant profilin isoforms, FH1-FH2 enhances filament elongation rates from about 10 to 42 times. Filaments buckle when FH1-FH2 is anchored specifically to cover slides, further indicating that AtFH 14 moves processively on the elongating barbed end. At high concentration, AtFH 14 bundles actin filaments randomly into antiparallel or parallel spindle-like structures; however, the FH1-FH2-mediated bundles become thinner and longer in the presence of plant profilins. This is the direct demonstration of a processive formin from plants. Our results also illuminate the molecular mechanism of AtFH14 in regulating actin dynamics via association with profilin.
