Active mechanical stabilization of the viscoplastic intracellular space of Dictyostelia cells by microtubule–actin crosstalk

Abstract

The micro-viscoelasticity of the intracellular space of Dictyostelium discoideum cells is studied by evaluating the intracellular transport of magnetic force probes and their viscoelastic responses to force pulses of 20-700 pN. The role of the actin cortex, the microtubule (MT) aster and their crosstalk is explored by comparing the behaviour of wild-type cells, myosin II null mutants, latrunculin A and benomyl treated cells. The MT coupled beads perform irregular local and long range directed motions which are characterized by measuring their velocity distributions (P(v)). The correlated motion of the MT and the centrosome are evaluated by microfluorescence of GFP-labelled MTs. P(v) can be represented by log-normal distributions with long tails and it is determined by random sweeping motions (v $ 0.5 lm/s) of the MTs (caused by tangential forces on the filament ends coupled to the actin cortex) and by intermittent bead transports parallel to the MTs (v max $ 1.5 lm/s). The tails are due to spontaneous filament deflections (with speeds up to 10 lm/s) attributed to pre-stressing of the MT by local cortical tensions, generated by dynactin motors generating plus-end directed forces in the MTs. The viscoelastic responses are strongly non-linear and are mostly directed opposite or perpendicular to the force, showing that the cytoplasm behaves as an active viscoplastic body with time and force dependent drag coefficients. Nano-Newton loads exerted on the soft MT are balanced by traction forces arising at the MT ends coupled to the actin cortex and the centrosome, respectively. The mechanical coupling between the soft microtubules and the viscoelastic actin cortex provides cells with high mechanical stability despite the softness of the cytoplasm.

Mechanical tension contributes to clustering of neurotransmitter vesicles at presynaptic terminals

“…In axons of Drosophila embryos, the cytoskeleton consists of actin and microtubules. Thus, Factin may bridge and transfer forces between focal complexes at the terminal and microtubules (24). Under an acute imposed stretch, the axonal cytoskeleton deforms elastically when the tension is high (18).…”

Temporal Analysis of Active and Passive Transport in Living Cells

“…In addition, the cytoplasm is highly crowded, consisting of organelles, fixed compartments and freely moving vesicles. Microbeads are readily phagocytosed by DD cells and remain trapped in endosomes [16], thus resembling endogenous vesicles. They can bind to and be dragged along microtubules by dynein and kinesin motors.…”

“…Experiments were performed using 1:4 m ferromagnetic beads as tracer particles, internalized by DD amoebas in the vegetative state, when they exhibit a quasispherical shape and no aggregation behavior [16]. Bright-field imaging was performed using an Axiovert 200 microscope (Zeiss, Germany) and a CCD camera (C 4880-80 Hamamatsu, Germany).…”

Axonal Guidance by Surface Microstructuring for Intracellular Transport Investigations

“…[11] Vesicles undergo two types of motion in cells: They are either actively transported by the molecular motors dynein and kinesin along microtubules, or they undergo thermal diffusion in a crowed active intracellular medium. [12] To retrieve these complex properties observed in active vesicle transport, several theoretical models have emerged. Lipowsky et al [13][14][15][16] proposed two specific models, namely, a "tug-of-war" scenario for the two microtubule-associated motor types, in contrast to coordinated motor transport.…”