-
ELife May 2022The cytoskeleton - a collection of polymeric filaments, molecular motors, and crosslinkers - is a foundational example of active matter, and in the cell assembles into...
The cytoskeleton - a collection of polymeric filaments, molecular motors, and crosslinkers - is a foundational example of active matter, and in the cell assembles into organelles that guide basic biological functions. Simulation of cytoskeletal assemblies is an important tool for modeling cellular processes and understanding their surprising material properties. Here, we present (a Living Ensemble Simulator), a novel computational framework designed to surmount the limits of conventional simulation methods. We model molecular motors with crosslinking kinetics that adhere to a thermodynamic energy landscape, and integrate the system dynamics while efficiently and stably enforcing hard-body repulsion between filaments. Molecular potentials are entirely avoided in imposing steric constraints. Utilizing parallel computing, we simulate tens to hundreds of thousands of cytoskeletal filaments and crosslinking motors, recapitulating emergent phenomena such as bundle formation and buckling. This simulation framework can help elucidate how motor type, thermal fluctuations, internal stresses, and confinement determine the evolution of cytoskeletal active matter.
Topics: Computer Simulation; Cytoskeleton; Microtubules; Models, Biological; Molecular Motor Proteins
PubMed: 35617115
DOI: 10.7554/eLife.74160 -
Journal of Integrative Plant Biology Jan 2023Microtubule dynamics and organization are important for plant cell morphogenesis and development. The microtubule-based motor protein kinesins are mainly responsible for...
Microtubule dynamics and organization are important for plant cell morphogenesis and development. The microtubule-based motor protein kinesins are mainly responsible for the transport of some organelles and vesicles, although several have also been shown to regulate microtubule organization. The ARMADILLO REPEAT KINESIN (ARK) family is a plant-specific motor protein subfamily that consists of three members (ARK1, ARK2, and ARK3) in Arabidopsis thaliana. ARK2 has been shown to participate in root epidermal cell morphogenesis. However, whether and how ARK2 associates with microtubules needs further elucidation. Here, we demonstrated that ARK2 co-localizes with microtubules and facilitates microtubule bundling in vitro and in vivo. Pharmacological assays and microtubule dynamics analyses indicated that ARK2 stabilizes cortical microtubules. Live-cell imaging revealed that ARK2 moves along cortical microtubules in a processive mode and localizes both at the plus-end and the sidewall of microtubules. ARK2 therefore tracks and stabilizes the growing plus-ends of microtubules, which facilitates the formation of parallel microtubule bundles.
Topics: Arabidopsis; Arabidopsis Proteins; Microtubules; Kinesins; Microtubule-Associated Proteins
PubMed: 36169006
DOI: 10.1111/jipb.13373 -
Annals of the New York Academy of... Sep 2022During migration, cells invade, repair, and create barriers leading to the formation of new cellular contacts in target tissues. Cell migration requires many proteins... (Review)
Review
During migration, cells invade, repair, and create barriers leading to the formation of new cellular contacts in target tissues. Cell migration requires many proteins that collectively form the cytoskeleton. The main cytoskeletal elements are actin filaments, microtubules (MTs), and intermediate filaments. These structures work in concert with a large number of accessory proteins that contribute in a variety of ways to regulate filament assembly and turnover, to alter the configuration or arrangement of filaments by bundling or crosslinking, to link the cytoskeleton to other structures in the cell, such as membranes and junctions, and to transport cargo along the filaments. Sperm flagella protein-1 (Spef1), also designated calponin homology and microtubules-associated protein (CLAMP), is a multifunctional protein that interacts with cytoskeletal structures, including MTs, actin filaments, and focal adhesions in epithelia. In this review, we outline Spef1/CLAMP structure and expression in several cellular models. The function of Spef1/CLAMP in flagellar and ciliary motility, MT-binding and stability, regulation of planar cell polarity, and potential contribution to the maintenance of actin-based structures, such as lamellipodia and filopodia during cell migration, are also discussed.
Topics: Actins; Calcium-Binding Proteins; Cell Movement; Cell Polarity; Humans; Male; Microfilament Proteins; Microtubules; Semen; Calponins
PubMed: 35710871
DOI: 10.1111/nyas.14845 -
Frontiers in Cell and Developmental... 2022Pancreatic islet β cells regulate glucose homeostasis glucose-stimulated insulin secretion (GSIS). Cytoskeletal polymers microtubules (MTs) serve as tracks for the... (Review)
Review
Pancreatic islet β cells regulate glucose homeostasis glucose-stimulated insulin secretion (GSIS). Cytoskeletal polymers microtubules (MTs) serve as tracks for the transport and positioning of secretory insulin granules. MT network in β cells has unique morphology with several distinct features, which support granule biogenesis ( Golgi-derived MT array), net non-directional transport ( interlocked MT mesh), and control availability of granules at secretion sites ( submembrane MT bundle). The submembrane MT array, which is parallel to the plasma membrane and serves to withdraw excessive granules from the secretion hot spots, is destabilized and fragmented downstream of high glucose stimulation, allowing for regulated secretion. The origin of such an unusual MT network, the features that define its functionality, and metabolic pathways that regulate it are still to a large extent elusive and are a matter of active investigation and debate. Besides the MT network itself, it is important to consider the interplay of molecular motors that drive and fine-tune insulin granule transport. Importantly, activity of kinesin-1, which is the major MT-dependent motor in β cells, transports insulin granules, and has a capacity to remodel MT network, is also regulated by glucose. We discuss yet unknown potential avenues toward understanding how MT network and motor proteins provide control for secretion in coordination with other GSIS-regulating mechanisms.
PubMed: 35874834
DOI: 10.3389/fcell.2022.915206 -
The Journal of Cell Biology Mar 2022Contractile ring constriction during cytokinesis is thought to compact central spindle microtubules to form the midbody, an antiparallel microtubule bundle at the...
Contractile ring constriction during cytokinesis is thought to compact central spindle microtubules to form the midbody, an antiparallel microtubule bundle at the intercellular bridge. In Caenorhabditis elegans, central spindle microtubule assembly requires targeting of the CLASP family protein CLS-2 to the kinetochores in metaphase and spindle midzone in anaphase. CLS-2 targeting is mediated by the CENP-F-like HCP-1/2, but their roles in cytokinesis and midbody assembly are not known. We found that although HCP-1 and HCP-2 mostly function cooperatively, HCP-1 plays a more primary role in promoting CLS-2-dependent central spindle microtubule assembly. HCP-1/2 codisrupted embryos did not form central spindles but completed cytokinesis and formed functional midbodies capable of supporting abscission. These central spindle-independent midbodies appeared to form via contractile ring constriction-driven bundling of astral microtubules at the furrow tip. This work suggests that, in the absence of a central spindle, astral microtubules can support midbody assembly and that midbody assembly is more predictive of successful cytokinesis than central spindle assembly.
Topics: Animals; Caenorhabditis elegans; Caenorhabditis elegans Proteins; Embryo, Nonmammalian; Green Fluorescent Proteins; Microtubules; Spindle Apparatus
PubMed: 34994802
DOI: 10.1083/jcb.202011085 -
Biology Nov 2020Fascin, an actin-binding protein, regulates many developmental migrations and contributes to cancer metastasis. Specifically, Fascin promotes cell motility, invasion,... (Review)
Review
Fascin, an actin-binding protein, regulates many developmental migrations and contributes to cancer metastasis. Specifically, Fascin promotes cell motility, invasion, and adhesion by forming filopodia and invadopodia through its canonical actin bundling function. In addition to bundling actin, Fascin has non-canonical roles in the cell that are thought to promote cell migration. These non-canonical functions include regulating the activity of other actin-binding proteins, binding to and regulating microtubules, mediating mechanotransduction to the nucleus via interaction with the Linker of the Nucleoskeleton and Cytoskeleton (LINC) Complex, and localizing to the nucleus to regulate nuclear actin, the nucleolus, and chromatin modifications. The many functions of Fascin must be coordinately regulated to control cell migration. While much remains to be learned about such mechanisms, Fascin is regulated by post-translational modifications, prostaglandin signaling, protein-protein interactions, and transcriptional means. Here, we review the structure of Fascin, the various functions of Fascin and how they contribute to cell migration, the mechanisms regulating Fascin, and how Fascin contributes to diseases, specifically cancer metastasis.
PubMed: 33212856
DOI: 10.3390/biology9110403 -
Cells Sep 2022To cope with continuous physiological and environmental stresses, cells of all sizes require an effective wound repair process to seal breaches to their cortex. Once a... (Review)
Review
To cope with continuous physiological and environmental stresses, cells of all sizes require an effective wound repair process to seal breaches to their cortex. Once a wound is recognized, the cell must rapidly plug the injury site, reorganize the cytoskeleton and the membrane to pull the wound closed, and finally remodel the cortex to return to homeostasis. Complementary studies using various model organisms have demonstrated the importance and complexity behind the formation and translocation of an actin ring at the wound periphery during the repair process. Proteins such as actin nucleators, actin bundling factors, actin-plasma membrane anchors, and disassembly factors are needed to regulate actin ring dynamics spatially and temporally. Notably, Rho family GTPases have been implicated throughout the repair process, whereas other proteins are required during specific phases. Interestingly, although different models share a similar set of recruited proteins, the way in which they use them to pull the wound closed can differ. Here, we describe what is currently known about the formation, translocation, and remodeling of the actin ring during the cell wound repair process in model organisms, as well as the overall impact of cell wound repair on daily events and its importance to our understanding of certain diseases and the development of therapeutic delivery modalities.
Topics: Actins; Cell Membrane; Cytoskeleton; Microtubules; rho GTP-Binding Proteins
PubMed: 36139352
DOI: 10.3390/cells11182777 -
Journal of Integrative Plant Biology Mar 2023In plants, cortical microtubules anchor to the plasma membrane in arrays and play important roles in cell shape. However, the molecular mechanism of microtubule binding...
In plants, cortical microtubules anchor to the plasma membrane in arrays and play important roles in cell shape. However, the molecular mechanism of microtubule binding proteins, which connect the plasma membrane and cortical microtubules in cell morphology remains largely unknown. Here, we report that a plasma membrane and microtubule dual-localized IQ67 domain protein, IQD21, is critical for cotyledon pavement cell (PC) morphogenesis in Arabidopsis. iqd21 mutation caused increased indentation width, decreased lobe length, and similar lobe number of PCs, whereas IQD21 overexpression had a different effect on cotyledon PC shape. Weak overexpression led to increased lobe number, decreased indentation width, and similar lobe length, while moderate or great overexpression resulted in decreased lobe number, indentation width, and lobe length of PCs. Live-cell observations revealed that IQD21 accumulation at indentation regions correlates with lobe initiation and outgrowth during PC development. Cell biological and genetic approaches revealed that IQD21 promotes transfacial microtubules anchoring to the plasma membrane via its polybasic sites and bundling at the indentation regions in both periclinal and anticlinal walls. IQD21 controls cortical microtubule organization mainly through promoting Katanin 1-mediated microtubule severing during PC interdigitation. These findings provide the genetic evidence that transfacial microtubule arrays play a determinant role in lobe formation, and the insight into the molecular mechanism of IQD21 in transfacial microtubule organization at indentations and puzzle-shaped PC development.
Topics: Arabidopsis Proteins; Microtubules; Arabidopsis; Katanin; Morphogenesis
PubMed: 36263896
DOI: 10.1111/jipb.13393 -
FASEB Journal : Official Publication of... Dec 2020Dynamin 1 is a neuronal endocytic protein that participates in vesicle formation by scission of invaginated membranes. Dynamin 1 is also expressed in the kidney;...
Dynamin 1 is a neuronal endocytic protein that participates in vesicle formation by scission of invaginated membranes. Dynamin 1 is also expressed in the kidney; however, its physiological significance to this organ remains unknown. Here, we show that dynamin 1 is crucial for microtubule organization and stabilization in glomerular podocytes. By immunofluorescence and immunoelectron microscopy, dynamin 1 was concentrated at microtubules at primary processes in rat podocytes. By immunofluorescence of differentiated mouse podocytes (MPCs), dynamin 1 was often colocalized with microtubule bundles, which radially arranged toward periphery of expanded podocyte. In dynamin 1-depleted MPCs by RNAi, α-tubulin showed a dispersed linear filament-like localization, and microtubule bundles were rarely observed. Furthermore, dynamin 1 depletion resulted in the formation of discontinuous, short acetylated α-tubulin fragments, and the decrease of microtubule-rich protrusions. Dynamins 1 and 2 double-knockout podocytes showed dispersed acetylated α-tubulin and rare protrusions. In vitro, dynamin 1 polymerized around microtubules and cross-linked them into bundles, and increased their resistance to the disassembly-inducing reagents Ca and podophyllotoxin. In addition, overexpression and depletion of dynamin 1 in MPCs increased and decreased the nocodazole resistance of microtubules, respectively. These results suggest that dynamin 1 supports the microtubule bundle formation and participates in the stabilization of microtubules.
Topics: Animals; Cells, Cultured; Dynamin I; Endocytosis; Epithelial Cells; Kidney; Male; Mice; Mice, Inbred C57BL; Mice, Knockout; Microtubule-Associated Proteins; Microtubules; Podocytes; Rats; Tubulin
PubMed: 33070431
DOI: 10.1096/fj.202001240RR -
Cytoskeleton (Hoboken, N.J.) Feb 2024The microtubule cytoskeleton is a major structural element inside cells that directs self-organization using microtubule-associated proteins and motors. It has been...
The microtubule cytoskeleton is a major structural element inside cells that directs self-organization using microtubule-associated proteins and motors. It has been shown that finite-sized, spindle-like microtubule organizations, called "tactoids," can form in vitro spontaneously from mixtures of tubulin and the antiparallel crosslinker, MAP65, from the MAP65/PRC1/Ase family. Here, we probe the ability of MAP65 to form tactoids as a function of the ionic strength of the buffer to attempt to break the electrostatic interactions binding MAP65 to microtubules and inter-MAP65 binding. We observe that, with increasing monovalent salts, the organizations change from finite tactoids to unbounded length bundles, yet the MAP65 binding and crosslinking appear to stay intact. We further explore the effects of ionic strength on the dissociation constant of MAP65 using both microtubule pelleting and single-molecule binding assays. We find that salt can reduce the binding, yet salt never negates it. Instead, we believe that the salt is affecting the ability of the MAP65 to form phase-separated droplets, which cause the nucleation and growth of tactoids, as recently demonstrated.
PubMed: 38385864
DOI: 10.1002/cm.21839