Clamped-Filament Elongation Model for Actin-Based Motors

Dickinson, Richard B.; Purich, Daniel L.

doi:10.1016/s0006-3495(02)75425-8

Cited by 134 publications

(120 citation statements)

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“…Because VASP is a key component of the actoclampin molecular motors responsible for actin-based cell motility (14,20), and because VASP was originally identified as a major substrate for cGMP-stimulated protein kinase in platelets and endothelial cells (21), we investigated whether NO and CO elicited similar effects on VASP phosphorylation and its mobilization to the leading edge of motile cells. VASP function appears to be chiefly regulated by phosphorylation at Ser-157 and Ser-239, and the availability of phosphorylation site-specific antibodies provides an unambiguous way to define both the sites and extents of signal molecule-activated VASP phosphorylation.…”

Section: Resultsmentioning

confidence: 99%

“…We previously demonstrated the central role of the actin cytoskeleton in EPC migration (13), and our findings suggest that NO has a critical function within EPCs, where it regulates the distribution of vasodilator-stimulated phosphoprotein (VASP). The latter plays a pivotal role in promoting actin filament elongation at the leading edge by forming an active molecular motor complex that propels motility (14). VASP contains three distinct phosphorylation sites (Ser-157, Ser-239, and Thr-278), the first of which is preferentially phosphorylated by cAMP-dependent protein kinase (PKA) and the second by cGMP-dependent protein kinase (PKG).…”

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Carbon Monoxide and Nitric Oxide Mediate Cytoskeletal Reorganization in Microvascular Cells via Vasodilator-Stimulated Phosphoprotein Phosphorylation

et al. 2008

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OBJECTIVE-We examined the effect of the vasoactive agents carbon monoxide (CO) and nitric oxide (NO) on the phosphorylation and intracellular redistribution of vasodilator-stimulated phosphoprotein (VASP), a critical actin motor protein required for cell migration that also controls vasodilation and platelet aggregation. RESEARCH DESIGN AND METHODS-We examined the effect of donor-released CO and NO in endothelial progenitor cells (EPCs) and platelets from nondiabetic and diabetic subjects and in human microvascular endothelial cells (HMECs) cultured under low (5.5 mmol/l) or high (25 mmol/l) glucose conditions. VASP phosphorylation was evaluated using phosphorylation site-specific antibodies.RESULTS-In control platelets, CO selectively promotes phosphorylation at VASP Ser-157, whereas NO promotes phosphorylation primarily at Ser-157 and also at Ser-239, with maximal responses at 1 min with both agents on Ser-157 and at 15 min on Ser-239 with NO treatment. In diabetic platelets, neither agent resulted in VASP phosphorylation. In nondiabetic EPCs, NO and CO increased phosphorylation at Ser-239 and Ser-157, respectively, but this response was markedly reduced in diabetic EPCs. In endothelial cells cultured under low glucose conditions, both CO and NO induced phosphorylation at Ser-157 and Ser-239; however, this response was completely lost when cells were cultured under high glucose conditions. In control EPCs and in HMECs exposed to low glucose, VASP was redistributed to filopodia-like structures following CO or NO exposure; however, redistribution was dramatically attenuated under high glucose conditions. CONCLUSIONS-Vasoactive gases CO and NO promote cytoskeletal changes through site-and cell type-specific VASP phosphorylation, and in diabetes, blunted responses to these agents may lead to reduced vascular repair and tissue perfusion. T he gaseous signal molecules nitric oxide (NO) and carbon monoxide (CO) exert multiple modulatory actions in regulating vascular function. While NO effects have been recognized for over a decade, similar vasoregulatory action of CO was established only recently. CO is generated by heme oxygenase (HO)-1 under a wide variety of conditions (e.g., cell exposure to such stressors as hypoxia, growth factors, and cytokine stimulation) that activate the enzyme (1,2). Unlike its highly reactive cognate NO, which participates in multiple redox reactions, CO is a relatively stable gas that exhibits extraordinary affinity for heme centers (3-5). Like NO, the signaling effects of CO rely in part on its ability to form a complex with the heme moiety of soluble guanylate cyclase (sGC), stimulating the synthesis of the diffusible second messenger guanosine 3Ј5Ј-cyclic monophosphate (cGMP) (6). The sGC/cGMP pathway plays a critical role in mediating the effects of CO on vascular relaxation and inhibition of platelet aggregation and coagulation (7,8).A recently recognized property of NO is its cell typespecific facilitation or inhibition of cell migration (9), a complex process involving molecular-...

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Section: Resultsmentioning

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Carbon Monoxide and Nitric Oxide Mediate Cytoskeletal Reorganization in Microvascular Cells via Vasodilator-Stimulated Phosphoprotein Phosphorylation

et al. 2008

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“…It is derived from the "actoclampin" model of actin filament end-tracking proteins (22)(23)(24), in which processive and tethered elongation is achieved Significance Ena/VASP proteins are tetramers that drive the processive elongation of actin filaments. Because Ena/VASP proteins are implicated in motility, embryogenesis, and cancer, it is mandatory to understand their molecular mechanism.…”

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Distinct VASP tetramers synergize in the processive elongation of individual actin filaments from clustered arrays

Brühmann

Ushakov

Winterhoff

et al. 2017

Proc. Natl. Acad. Sci. U.S.A.

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Ena/VASP proteins act as actin polymerases that drive the processive elongation of filament barbed ends in membrane protrusions or at the surface of bacterial pathogens. Based on previous analyses of fast and slow elongating VASP proteins by in vitro total internal reflection fluorescence microscopy (TIRFM) and kinetic and thermodynamic measurements, we established a kinetic model of Ena/VASP-mediated actin filament elongation. At steady state, it entails that tetrameric VASP uses one of its arms to processively track growing filament barbed ends while three G-actin-binding sites (GABs) on other arms are available to recruit and deliver monomers to the filament tip, suggesting that VASP operates as a single tetramer in solution or when clustered on a surface, albeit processivity and resistance toward capping protein (CP) differ dramatically between both conditions. Here, we tested the model by variation of the oligomerization state and by increase of the number of GABs on individual polypeptide chains. In excellent agreement with model predictions, we show that in solution the rates of filament elongation directly correlate with the number of free GABs. Strikingly, however, irrespective of the oligomerization state or presence of additional GABs, filament elongation on a surface invariably proceeded with the same rate as with the VASP tetramer, demonstrating that adjacent VASP molecules synergize in the elongation of a single filament. Additionally, we reveal that actin ATP hydrolysis is not required for VASP-mediated filament assembly. Finally, we show evidence for the requirement of VASP to form tetramers and provide an amended model of processive VASPmediated actin assembly in clustered arrays.actin | Ena/VASP | TIRF | cluster | formin

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“…The ''actoclampin'' model (4,5) proposes that the adhesive bonds are never broken, and instead that the bacterium advances through the discrete, ATP-hydrolysis-driven advancement of ''end-clamping'' proteins that bind the bacterium to the elongating ends of actin filaments. In this case, the biochemical rates of hydrolysis and filament growth reactions determine the rate of movement.…”

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“…Under the simplest assumptions, the tethered elastic Brownian ratchet and elastic gel models predict that bacteria will move faster at higher temperatures, and also that the steepness of the temperature dependence will be related in a complex way to the characteristic average speed of that bacterium. In the actoclampin model, speed is strongly coupled to the rate of ATP hydrolysis (4), so the temperature dependence of all bacteria should follow that of ATP hydrolysis. The simple kinetic form of the cooperative thermal breakage model predicts that (i) the speed of individual bacteria will increase approximately as an Arrhenius-type function of temperature and (ii) among bacteria, the temperature dependence will be ''rate-compensated'' (7,8), i.e., the prefactor A and the apparent activation energy, E a , of the Arrhenius dependence among individual bacteria will be correlated over a wide range.…”

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Adhesion controls bacterial actin polymerization-based movement

Soo

Theriot²

2005

Proc. Natl. Acad. Sci. U.S.A.

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As part of its infectious life cycle, the bacterial pathogen Listeria monocytogenes propels itself through the host-cell cytoplasm by triggering the polymerization of host-cell actin near the bacterial surface, harnessing the activity of several cytoskeletal proteins used during actin-based cell crawling. To distinguish among several classes of biophysical models of actin-based bacterial movement, we used a high-throughput tracking technique to record the movement of many individual bacteria during temperature shifts. The speed of each bacterium varied strongly with temperature, closely following the Arrhenius rate law. Among bacteria, the prefactor A of the Arrhenius dependence unexpectedly varied exponentially with apparent activation energy, E a, over a wide range (8 -21 kcal͞mol), reminiscent of the ''rate compensation effect'' of classical catalytic reactions. Average E a were increased for mutant bacteria deficient in binding Ena͞VASP proteins and bacteria moving in diluted extract. These two effects were additive. The observed temperature and rate compensation effects are consistent with a class of simple kinetic models in which the bacterium advances through the thermally driven, cooperative breakage of groups of adhesive bonds on its surface. The estimated number of coupled adhesive bonds N on the bacterial surface varies between 10 and 40 bonds. In contrast to other models, this model correctly predicts an experimentally observed negative correlation between bacterial speed and actin gel density. The idea that speed depends on adhesion, rather than polymerization, suggests several alternative mechanisms by which known cytoskeletal regulatory proteins could control cellular movement.ActA ͉ Listeria ͉ motility M any types of eukaryotic cells, including immune system lymphocytes, neurons, and cancer cells, use the forces generated by polymerizing actin filament networks to change shape and to move. In the actin polymerization-based movement of intracellular bacterial pathogens such as Listeria monocytogenes, individual bacteria harness the same cytoskeletal machinery to move within the host-cell cytoplasm (1).Several classes of biophysical models have been proposed to explain the mechanism of actin polymerization-based movement. The models differ in which steps in the motility cycle are proposed to control the rate of bacterial movement.The ''tethered elastic Brownian ratchet'' model (2) proposes that a large force generated by the network of polymerizing actin filaments in the ''comet tail'' behind the bacterium continuously induces the breakage of individual adhesive bonds between the bacterium and the surrounding actin gel, pushing the bacterium forward. In this model, bacteria move at the speed at which propulsive and adhesive forces are balanced; this equilibrium velocity depends strongly on the specific properties of actin polymerization and adhesion. The related ''elastic gel '' model (3) proposes that the elastic expansion of the actin gel behind the bacterium also contributes to propulsion, ''s...

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Clamped-Filament Elongation Model for Actin-Based Motors

Cited by 134 publications

References 50 publications

Carbon Monoxide and Nitric Oxide Mediate Cytoskeletal Reorganization in Microvascular Cells via Vasodilator-Stimulated Phosphoprotein Phosphorylation

Carbon Monoxide and Nitric Oxide Mediate Cytoskeletal Reorganization in Microvascular Cells via Vasodilator-Stimulated Phosphoprotein Phosphorylation

Distinct VASP tetramers synergize in the processive elongation of individual actin filaments from clustered arrays

Adhesion controls bacterial actin polymerization-based movement

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