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Cytoplasmic streaming with extremely high velocity (∼70 μm s−1) occurs in cells of the characean algae (Chara). Because cytoplasmic streaming is caused by myosin XI, it has been suggested that a myosin XI with a velocity of 70 μm s−1, the fastest myosin measured so far, exists in Chara cells. However, the velocity of the previously cloned Chara corallina myosin XI (CcXI) was about 20 μm s−1, one-third of the cytoplasmic streaming velocity in Chara. Recently, the genome sequence of Chara braunii has been published, revealing that this alga has four myosin XI genes. We cloned these four myosin XI (CbXI-1, 2, 3, and 4) and measured their velocities. While the velocities of CbXI-3 and CbXI-4 motor domains (MDs) were similar to that of CcXI MD, the velocities of CbXI-1 and CbXI-2 MDs were 3.2 times and 2.8 times faster than that of CcXI MD, respectively. The velocity of chimeric CbXI-1, a functional, full-length CbXI-1 construct, was 60 μm s−1. These results suggest that CbXI-1 and CbXI-2 would be the main contributors to cytoplasmic streaming in Chara cells and show that these myosins are ultrafast myosins with a velocity 10 times faster than fast skeletal muscle myosins in animals. We also report an atomic structure (2.8-Å resolution) of myosin XI using X-ray crystallography. Based on this crystal structure and the recently published cryo-electron microscopy structure of acto-myosin XI at low resolution (4.3-Å), it appears that the actin-binding region contributes to the fast movement of Chara myosin XI. Mutation experiments of actin-binding surface loops support this hypothesis.

Recent advances in cell biology enable precise molecular perturbations. The spatiotemporal organization of cells and organisms, however, also depends on physical processes such as diffusion or cytoplasmic flows, and strategies to perturb physical transport inside cells are not yet available. Here, we demonstrate focused-light-induced cytoplasmic streaming (FLUCS). FLUCS is local, directional, dynamic, probe-free, physiological, and is even applicable through rigid egg shells or cell walls. We explain FLUCS via time-dependent modelling of thermoviscous flows. Using FLUCS, we demonstrate that cytoplasmic flows drive partitioning-defective protein (PAR) polarization in Caenorhabditis elegans zygotes, and that cortical flows are sufficient to transport PAR domains and invert PAR polarity. In addition, we find that asymmetric cell division is a binary decision based on gradually varying PAR polarization states. Furthermore, the use of FLUCS for active microrheology revealed a metabolically induced fluid-to-solid transition of the yeast cytoplasm. Our findings establish how a wide range of transport-dependent models of cellular organization become testable by FLUCS. Mittasch et al. show that controlling cytoplasmic flow via focused-light-induced cytoplasmic streaming (FLUCS), a non-invasive technique, can be used to invert asymmetric cell division in Caenorhabditis elegans zygotes.

In somatic cells, the position of the cell centroid is dictated by the centrosome. The centrosome is instrumental in nucleus positioning, the two structures being physically connected. Mouse oocytes have no centrosomes, yet harbour centrally located nuclei. We demonstrate how oocytes define their geometric centre in the absence of centrosomes. Using live imaging of oocytes, knockout for the formin 2 actin nucleator, with off-centred nuclei, together with optical trapping and modelling, we discover an unprecedented mode of nucleus positioning. We document how active diffusion of actin-coated vesicles, driven by myosin Vb, generates a pressure gradient and a propulsion force sufficient to move the oocyte nucleus. It promotes fluidization of the cytoplasm, contributing to nucleus directional movement towards the centre. Our results highlight the potential of active diffusion, a prominent source of intracellular transport, able to move large organelles such as nuclei, providing in vivo evidence of its biological function. By live imaging of mouse oocytes, Verlhac and colleagues demonstrate that actin-coated vesicles together with myosin Vb participate in centring of the nucleus by creating a gradient of cytoplasmic forces.

We investigate the myosin XI-driven transport network in Arabidopsis using protein–protein interaction, subcellular localization, gene knockout, and bioinformatics analyses. The two major groups of nodes in this network are myosins XI and their membrane-anchored receptors (MyoB) that, together, drive endomembrane trafficking and cytoplasmic streaming in the plant cells. The network shows high node connectivity and is dominated by generalists, with a smaller fraction of more specialized myosins and receptors. We show that interaction with myosins and association with motile vesicles are common properties of the MyoB family receptors. We identify previously uncharacterized myosin-binding proteins, putative myosin adaptors that belong to two unrelated families, with four members each (MadA and MadB). Surprisingly, MadA1 localizes to the nucleus and is rapidly transported to the cytoplasm, suggesting the existence of myosin XI-driven nucleocytoplasmic trafficking. In contrast, MadA2 and MadA3, as well as MadB1, partition between the cytosolic pools of motile endomembrane vesicles that colocalize with myosin XI-K and diffuse material that does not. Gene knockout analysis shows that MadB1–4 contribute to polarized root hair growth, phenocopying myosins, whereas MadA1–4 are redundant for this process. Phylogenetic analysis reveals congruent evolutionary histories of the myosin XI, MyoB, MadA, and MadB families. All these gene families emerged in green algae and show concurrent expansions via serial duplication in flowering plants. Thus, the myosin XI transport network increased in complexity and robustness concomitantly with the land colonization by flowering plants and, by inference, could have been a major contributor to this process.

Cytoplasmic streaming in Drosophila oocytes is a microtubule-based bulk cytoplasmic movement. Streaming efficiently circulates and localizes mRNAs and proteins deposited by the nurse cells across the oocyte. This movement is driven by kinesin-1, amajor microtubule motor. Recently, we have shown that kinesin-1 heavy chain (KHC) can transport one microtubule on another microtubule, thus driving microtubule–microtubule sliding in multiple cell types. To study the role of microtubule sliding in oocyte cytoplasmic streaming, we used a Khc mutant that is deficient in microtubule sliding but able to transport a majority of cargoes. We demonstrated that streaming is reduced by genomic replacement of wild-type Khc with this sliding-deficient mutant. Streaming can be fully rescued by wild-type KHC and partially rescued by a chimeric motor that cannot move organelles but is active in microtubule sliding. Consistent with these data, we identified two populations of microtubules in fast-streaming oocytes: a network of stable microtubules anchored to the actin cortex and free cytoplasmic microtubules that moved in the ooplasm. We further demonstrated that the reduced streaming in sliding-deficient oocytes resulted in posterior determination defects. Together, we propose that kinesin-1 slides free cytoplasmic microtubules against cortically immobilized microtubules, generating forces that contribute to cytoplasmic streaming and are essential for the refinement of posterior determinants.

Found in many large eukaryotic cells, particularly in plants, cytoplasmic streaming is the circulation of their contents driven by fluid entrainment from particles carried by molecular motors at the cell periphery. In the more than two centuries since its discovery, streaming has frequently been conjectured to aid in transport and mixing of molecular species in the cytoplasm and, by implication, in cellular homeostasis, yet no theoretical analysis has been presented to quantify these processes. We show by a solution to the coupled dynamics of fluid flow and diffusion appropriate to the archetypal \"rotational streaming\" of algal species such as Chara and Nitella that internal mixing and the transient dynamical response to changing external conditions can indeed be enhanced by streaming, but to an extent that depends strongly on the pitch of the helical flow. The possibility that this may have a developmental consequence is illustrated by the coincidence of the exponential growth phase of Nitella and the point of maximum enhancement of those processes.

Programmed cell death (PCD) functions in the developmental remodeling of leaf shape in higher plants, a process analogous to digit formation in the vertebrate limb. In this study, we provide a cytological characterization of the time course of events as PCD remodels young expanding leaves of the lace plant. Tonoplast rupture is the first PCD event in this system, indicated by alterations in cytoplasmic streaming, loss of anthocyanin color, and ultrastructural appearance. Nuclei become terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling positive soon afterward but do not become morphologically altered until late stages of PCD. Genomic DNA is fragmented, but not into internucleosomal units. Other cytoplasmic changes, such as shrinkage and degradation of organelles, occur later. This form of PCD resembles tracheary element differentiation in cytological execution but requires unique developmental regulation so that discrete panels of tissue located equidistantly between veins undergo PCD while surrounding cells do not.

Recent studies of aquatic and land plants show that similar phenomena determine intracellular transport of organelles and vesicles. This suggests that aspects of cell signaling involved in development and response to external stimuli are conserved across species. The movement of molecular motors along cytoskeletal filaments directly or indirectly entrains the fluid cytosol, driving cyclosis (i.e., cytoplasmic streaming) and affecting gradients of molecular species within the cell, with potentially important metabolic implications as a driving force for cell expansion. Research has shown that myosin XI functions in organelle movement driving cytoplasmic streaming in aquatic and land plants. Despite the conserved cytoskeletal machinery propelling organelle movement among aquatic and land plants, the velocities of cyclosis in plant cells varies according to cell types, developmental stage of the cell, and plant species. Here, we synthesize recent insights into cytoplasmic streaming, molecular gradients, cytoskeletal and membrane dynamics, and expand current cellular models to identify important gaps in current research.

We investigate how an amoeba mechanically moves its own center of gravity using the model organism Physarum plasmodium. Time-dependent velocity fields of protoplasmic streaming over the whole plasmodia were measured with a particle image velocimetry program developed for this work. Combining these data with measurements of the simultaneous movements of the plasmodia revealed a simple physical mechanism of locomotion. The shuttle streaming of the protoplasm was not truly symmetric due to the peristalsis-like movements of the plasmodium. This asymmetry meant that the transport capacity of the stream was not equal in both directions, and a net forward displacement of the center of gravity resulted. The generality of this as a mechanism for amoeboid locomotion is discussed.

The toxicity of heavy metals (Cd, Zn, and Pb) was assessed by in vivo observations of their effect on cytoplasmic streaming in Allium cepa L. bulb scale epidermal cells. On the basis of our results, the order of toxicity of the studied cations is Zn < Pb [double less-than sign] Cd. The difference in toxicity between cadmium and lead was found to be very large. When cytoplasmic streaming was assessed, this difference was threefold. When the total content of cadmium and lead (determined by inductively coupled plasma mass spectrometry) was the criterion, the difference in toxicity was 15-fold. Fractionation of the tissue and enzymatic digestion of the cells revealed that the largest proportion of cadmium was located in the cell walls (56%), whereas almost all of the lead (97.6%) was accumulated in an insoluble form. The speciation of water-soluble Pb and Cd fractions is discussed on the basis of analysis by capillary zone electrophoresis interfaced with inductively coupled plasma mass spectrometry of water extracts from epidermal cells. Lead and cadmium appeared to be bound mainly to salts, which explains their toxicity. Cadmium was complexed (detoxified) by organic acids, while thiols were the metal-complexing species for lead. Histidine formed complexes with both cadmium and lead. Ultrastructural analyses showed that lead was encapsulated in small vesicles in the cytoplasm. Fluorescence studies of the endoplasmic reticulum (ER) revealed that it underwent extensive fragmentation under the influence of lead, with numerous ER vesicles appearing in the cells. In other words, the lead deposits in the cytoplasm were contained in vesicles arising from fragmentation of the ER. These observations indicate that epidermal cells have a rapid and effective mechanism for detoxifying lead involving the ER, and this may be one of the mechanisms accounting for the lower toxicity of lead in comparison with cadmium. The suitability of Allium cepa bulb scale epidermal cells for use in ecotoxicological studies is also discussed. Step-by-step directions for this test are given.