In Vitro Reconstitution of Dynamic Co-organization of Microtubules and Actin Filaments in Emulsion Droplets

Vendel, K.J.A.; Alkemade, Celine; Andrea, Nemo; Koenderink, Gijsje H.; Dogterom, Marileen

doi:10.1007/978-1-0716-0219-5_5

Cited by 3 publications

(4 citation statements)

References 32 publications

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“…Nevertheless, our theoretical predictions on the response of the system to the change of the binding/diffusion ratio, could in principle be validated by experiments, as transport in the three-dimensional domain has recently been observed in droplets [ 14 ]. While directly manipulating the transport and binding rates may be harder to achieve experimentally, tuning the diffusion coefficient by changing the viscosity of the medium seems feasible.…”

Section: Discussionmentioning

confidence: 95%

“…Because preliminary experiments have revealed that the interaction between actin and microtubules via TipAct can change the dynamic properties of the latter [ 13 ], we first define a general model that accounts for this change in the dynamics. Nevertheless, our main interest lies in the actin transport mechanism observed in the experiments performed in the droplet, where no change in the dynamics of microtubules has been recorded [ 14 ]. Therefore, we will provide a solution of the model under the assumption that the dynamic properties of microtubules do not change as a consequence of the interaction with actin filaments.…”

Section: Methodsmentioning

confidence: 99%

“…This possibility was explored in recent in vitro experiments [ 10 , 13 , 14 ] employing the engineered protein construct TipAct [ 6 ] specifically designed to bind to both actin and microtubules. These experiments revealed a dual action of TipAct.…”

Section: Introductionmentioning

confidence: 99%

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Microtubule-based actin transport and localization in a spherical cell

Saltini

Mulder

2020

R. Soc. open sci.

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The interaction between actin filaments and microtubules is crucial for many eukaryotic cellular processes, such as, among others, cell polarization, cell motility and cellular wound healing. The importance of this interaction has long been recognized, yet very little is understood about both the underlying mechanisms and the consequences for the spatial (re)organization of the cellular cytoskeleton. At the same time, understanding the causes and the consequences of the interaction between different biomolecular components are key questions for in vitro research involving reconstituted biomolecular systems, especially in the light of current interest in creating minimal synthetic cells. In this light, recent in vitro experiments have shown that the actin-microtubule interaction mediated by the cytolinker TipAct, which binds to actin lattice and microtubule tips, causes the directed transport of actin filaments. We develop an analytical theory of dynamically unstable microtubules, nucleated from the centre of a spherical cell, in interaction with actin filaments. We show that, depending on the balance between the diffusion of unbound actin filaments and propensity to bind microtubules, actin is either concentrated in the centre of the cell, where the density of microtubules is highest, or becomes localized to the cell cortex.

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Section: Discussionmentioning

confidence: 95%

Section: Methodsmentioning

confidence: 99%

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Microtubule-based actin transport and localization in a spherical cell

Saltini

Mulder

2020

R. Soc. open sci.

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“…For example, TIRF-based assays employing yeast or Xenopus extracts have reconstituted contractile actomyosin rings 37 , mitotic spindles 26 , 38 , components of actin or microtubule assembly 39 , 40 , and even dynamics at the centrosome and kinetochores 36 , 41 , 42 , 43 . Moreover, such systems may pave the way toward artificial cell systems that have lipids or signaling factors present 44 , 45 , 46 .…”

Section: Discussionmentioning

confidence: 99%

Visualizing Actin and Microtubule Coupling Dynamics <em>In Vitro</em> by Total Internal Reflection Fluorescence (TIRF) Microscopy

Henty-Ridilla¹

2022

JoVE

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Traditionally, the actin and microtubule cytoskeletons have been studied as separate entities, restricted to specific cellular regions or processes, and regulated by different suites of binding proteins unique for each polymer. Many studies now demonstrate that the dynamics of both cytoskeletal polymers are intertwined and that this crosstalk is required for most cellular behaviors. A number of proteins involved in actinmicrotubule interactions have already been identified (i.e., Tau, MACF, GAS, formins, and more) and are well characterized with regard to either actin or microtubules alone. However, relatively few studies showed assays of actin-microtubule coordination with dynamic versions of both polymers. This may occlude emergent linking mechanisms between actin and microtubules. Here, a total internal reflection fluorescence (TIRF) microscopy-based in vitro reconstitution technique permits the visualization of both actin and microtubule dynamics from the one biochemical reaction. This technique preserves the polymerization dynamics of either actin filament or microtubules individually or in the presence of the other polymer. Commercially available Tau protein is used to demonstrate how actin-microtubule behaviors change in the presence of a classic cytoskeletal crosslinking protein. This method can provide reliable functional and mechanistic insights into how individual regulatory proteins coordinate actinmicrotubule dynamics at a resolution of single filaments or higher-order complexes.

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How cytoskeletal crosstalk makes cells move: Bridging cell-free and cell studies

Conboy,

Istúriz Petitjean,

van der Net

et al. 2024

Biophysics Reviews

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Cell migration is a fundamental process for life and is highly dependent on the dynamical and mechanical properties of the cytoskeleton. Intensive physical and biochemical crosstalk among actin, microtubules, and intermediate filaments ensures their coordination to facilitate and enable migration. In this review, we discuss the different mechanical aspects that govern cell migration and provide, for each mechanical aspect, a novel perspective by juxtaposing two complementary approaches to the biophysical study of cytoskeletal crosstalk: live-cell studies (often referred to as top-down studies) and cell-free studies (often referred to as bottom-up studies). We summarize the main findings from both experimental approaches, and we provide our perspective on bridging the two perspectives to address the open questions of how cytoskeletal crosstalk governs cell migration and makes cells move.

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In Vitro Reconstitution of Dynamic Co-organization of Microtubules and Actin Filaments in Emulsion Droplets

Cited by 3 publications

References 32 publications

Microtubule-based actin transport and localization in a spherical cell

Microtubule-based actin transport and localization in a spherical cell

Visualizing Actin and Microtubule Coupling Dynamics <em>In Vitro</em> by Total Internal Reflection Fluorescence (TIRF) Microscopy

How cytoskeletal crosstalk makes cells move: Bridging cell-free and cell studies

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