Evaporation-Induced Pattern Formation and Wetting in Active Microtubule-Kinesin Droplets

Abstract

Active networks composed of biopolymers and motor proteins provide versatile biomimetic systems that have advanced active matter physics and deepened our understanding of cytoskeletal dynamics and self-organization under diverse stimuli. In these systems, activity arises in aqueous solutions where motor proteins cross-link biopolymers and generate active stress driving the emergent network behavior. Here, we establish the active network in the form of a sessile, multi-component droplet on a substrate and investigate how evaporation influences its dynamics. We focus on how mass loss and compositional changes in the droplet reshape the behavior of the active suspension. We show that capillary and Marangoni flows drive the self-organization of microtubules into a distinctive radial arrangement within the droplet. The cross-linking ability of motor proteins gives rise to a striking non-monotonic wetting behavior, where the extensile stresses generated by the motor proteins strongly affect the characteristic timescale of the contact-line retracting and subsequent expansion. Using a combined experimental and theoretical approach, we demonstrate the crucial role of crosslinking in evaporating microtubule networks, and explain how active stresses together with evaporation-induced flows govern the dynamics of reconstituted microtubule systems and their wetting behavior. Evaporating droplets have recently attracted significant attention in the scientific community, and the findings of the setup presented in this study can have broad implications, ranging from self-organization and mechanical pattern formation in biological systems to questions about the origin of life.

Figures (12)

• Figure 1: Dynamics of active microtubule network inside an evaporating droplet. a) Schematic representation of the experimental setup. Microtubule active network embedded into a droplet of a mixture of salty buffer and PEG on a PLL-g-PEG functionalised substrate. The inset shows the biological building blocks constituting the network, namely microtubules arranged into bundles due to PEG depletion force and kinesin-streptavidin motor clusters. b) Micrographs showing the emergent behaviour of the self-organizing active network during the droplet evaporation over a time of 30 s. The pattern can be described by two zones marked as zone I, and zone II, which we refer to as the radial corona and the dense network, respectively. c) Dynamics of the contact line of the droplet and isotropic network during evaporation.
• Figure 2: Comparison of the dynamics of the active microtubule network in the presence (a–d) and absence (e–f) of PEG. (a)-(d): Micrographs and schematics of the droplet containing PEG for time interval of t = 0 -- 20 s, showing an inward contraction of the microtubule network and the contact line at short time, and an outward extension at long times (t = 30 -- 50 s). In the schematics the yellow arrows represent the evaporation of the droplet, the orange arrows the Marangoni flow, the blue arrows the capillary flow and the red dashed arrows the network contraction force. (e) and (f): Micrographs and schematics of the active microtubule network in a droplet without PEG, showing a slight inward motion of the contact line and the contraction of the network (zone II, which is highlighted by white arrows). Scale bars: 200 $\mu$m.
• Figure 3: a) Schematic representation of an evaporating droplet of radius R and PEG-induced Marangoni flow on a PLL-g-PEG functionalised surface. b) Measurements of the internal flow velocity of the droplet obtained with high resolution micro-PIV. Cross-sectional view of the droplet composed of M2B and PEG with molecular weight of 6 kDa: free interface (blue circles and fitting line), radial velocity (black arrows) and velocity profiles (black lines) at a distance from the contact line r$^\prime$ = R - r, where $R$ is the radius of the droplet and $r$ is the radial position in a cylindrical coordinate system with the origin at the center of the droplet. Inset: close-up of the Marangoni flow near the contact line at t = 30 s. See SI for more details.
• Figure 4: a) Temporal evolution of a microtubule network cross-linked by kinesin motors in a solution without ATP t = 0 -- 120 s. At t = 22 s the annular region close to the contact line and rich of thin radially organized bundles of microtubules is visible (see the zoom-in view of the annular zone on the right panel). In the time frame t = 45 -- 120 s non-monotonic behaviour can be observed. Scale bar: 200 $\mu$m. b) Micrographs of the evaporating droplet containing microtubule network with PEG in the absence of motor proteins. Scale bar: 500 $\mu$m.
• Figure 5: Dynamics of the contact line (a) and the network radius (b) under different experimental setups. Dark colors represent the averages, and lighter colors individual experiments. The radius is normalized by the initial radius. In all cases with motors, the network radius first decreases and then increases. This behavior is less evident in the average for the No PEG case while is clear in individual experiments. The reduced evidence in the average arises because the non-monotonic behavior occurs at different times in different experiments (see Figure S6 in SI). In (a) and (b), $R_0$ ($R_0^{n}$) denotes the initial radius of the droplet (network), and $R(t)$ ($R_n(t)$) denotes the radius of the droplet (network) over time. (c) The average turnover timescale, $\tau_s$ (black), and the initial retraction timescale, $\tau_{Init}$ (red), are shown for different conditions. The initial network retraction timescale, $\tau_{Init}$, defined as the timescale over which the network radius contracts to $90\%$ of its initial value, scales linearly with the turnover timescale, $\tau_s$. Specifically, the timescale over which the network starts expanding, $\tau_s$, scales linearly with $\tau_{Init}$, with a proportionality factor of approximately $3.4$. Error bars represent the standard deviation across different experiments.