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Cytoskeleton (Hoboken, N.J.) 2023Accurate placement of the cleavage furrow is crucial for successful cell division. Recent advancements have revealed that diverse mechanisms have evolved across...
Accurate placement of the cleavage furrow is crucial for successful cell division. Recent advancements have revealed that diverse mechanisms have evolved across different branches of the phylogenetic tree. Here, we employed Dictyostelium cells to validate previous models. We observed that during metaphase and early anaphase, mitotic spindles exhibited random rotary movements which ceased when the spindle elongated by approximately 7 μm. At this point, astral microtubules reached the polar cell cortex and fixed the spindle axis, causing cells to elongate by extending polar pseudopods and divide along the spindle axis. Therefore, the position of the furrow is determined when the spindle orientation is fixed. The distal ends of astral microtubules stimulate the extension of pseudopods at the polar cortex. One signal for pseudopod extension may be phosphatidylinositol trisphosphate in the cell membrane, but there appears to be another unknown signal. At the onset of polar pseudopod extension, cortical flow began from both poles toward the equator. We suggest that polar stimulation by astral microtubules determines the furrow position, induces polar pseudopod extension and cortical flow, and accumulates the elements necessary for the construction of the contractile ring.
Topics: Dictyostelium; Phylogeny; Cytokinesis; Microtubules; Spindle Apparatus; Anaphase
PubMed: 37650534
DOI: 10.1002/cm.21784 -
Frontiers in Immunology 2022Chemotaxis plays an essential role in recruitment of leukocytes to sites of inflammation. Eukaryotic cells sense chemoattractant with G protein-coupled receptors (GPCRs)... (Review)
Review
Chemotaxis plays an essential role in recruitment of leukocytes to sites of inflammation. Eukaryotic cells sense chemoattractant with G protein-coupled receptors (GPCRs) and chemotax toward gradients with an enormous concentration range through adaptation. Cells in adaptation no longer respond to the present stimulus but remain sensitive to stronger stimuli. Thus, adaptation provides a fundamental strategy for eukaryotic cells to chemotax through a gradient. Ras activation is the first step in the chemosensing GPCR signaling pathways that displays a transient activation behavior in both model organism and mammalian neutrophils. Recently, it has been revealed that C2GAP1 and CAPRI control the GPCR-mediated adaptation in and human neutrophils, respectively. More importantly, both Ras inhibitors regulate the sensitivity of the cells. These findings suggest an evolutionarily conserved molecular mechanism by which eukaryotic cells gate concentration range of chemoattractants for chemotaxis.
Topics: Animals; Humans; Chemotaxis; Dictyostelium; Chemotactic Factors; Receptors, G-Protein-Coupled; Signal Transduction; Mammals
PubMed: 36341344
DOI: 10.3389/fimmu.2022.1020117 -
Cells Dec 2021The lamellipodia and pseudopodia of migrating cells are produced and maintained by the Scar/WAVE complex. Thus, actin-based cell migration is largely controlled through...
The lamellipodia and pseudopodia of migrating cells are produced and maintained by the Scar/WAVE complex. Thus, actin-based cell migration is largely controlled through regulation of Scar/WAVE. Here, we report that the Abi subunit-but not Scar-is phosphorylated in response to extracellular signalling in cells. Like Scar, Abi is phosphorylated after the complex has been activated, implying that Abi phosphorylation modulates pseudopodia, rather than causing new ones to be made. Consistent with this, Scar complex mutants that cannot bind Rac are also not phosphorylated. Several environmental cues also affect Abi phosphorylation-cell-substrate adhesion promotes it and increased extracellular osmolarity diminishes it. Both unphosphorylatable and phosphomimetic Abi efficiently rescue the chemotaxis of Abi KO cells and pseudopodia formation, confirming that Abi phosphorylation is not required for activation or inactivation of the Scar/WAVE complex. However, pseudopodia and Scar patches in the cells with unphosphorylatable Abi protrude for longer, altering pseudopod dynamics and cell speed. , in which Scar and Abi are both unphosphorylatable, can still form pseudopods, but migrate substantially faster. We conclude that extracellular signals and environmental responses modulate cell migration by tuning the behaviour of the Scar/WAVE complex after it has been activated.
Topics: Cell Adhesion; Cell Movement; Chemotactic Factors; Dictyostelium; Extracellular Space; Mutation; Osmotic Pressure; Phosphorylation; Protozoan Proteins; Pseudopodia; Signal Transduction; Wiskott-Aldrich Syndrome Protein Family
PubMed: 34943993
DOI: 10.3390/cells10123485 -
The Journal of Biological Chemistry Dec 2020In animals, the response to chronic hypoxia is mediated by prolyl hydroxylases (PHDs) that regulate the levels of hypoxia-inducible transcription factor α (HIFα). PHD...
In animals, the response to chronic hypoxia is mediated by prolyl hydroxylases (PHDs) that regulate the levels of hypoxia-inducible transcription factor α (HIFα). PHD homologues exist in other types of eukaryotes and prokaryotes where they act on non HIF substrates. To gain insight into the factors underlying different PHD substrates and properties, we carried out biochemical and biophysical studies on PHD homologues from the cellular slime mold, and the protozoan parasite, , both lacking HIF. The respective prolyl-hydroxylases (DdPhyA and TgPhyA) catalyze prolyl-hydroxylation of S-phase kinase-associated protein 1 (Skp1), a reaction enabling adaptation to different dioxygen availability. Assays with full-length Skp1 substrates reveal substantial differences in the kinetic properties of DdPhyA and TgPhyA, both with respect to each other and compared with human PHD2; consistent with cellular studies, TgPhyA is more active at low dioxygen concentrations than DdPhyA. TgSkp1 is a DdPhyA substrate and DdSkp1 is a TgPhyA substrate. No cross-reactivity was detected between DdPhyA/TgPhyA substrates and human PHD2. The human Skp1 E147P variant is a DdPhyA and TgPhyA substrate, suggesting some retention of ancestral interactions. Crystallographic analysis of DdPhyA enables comparisons with homologues from humans, , and prokaryotes, informing on differences in mobile elements involved in substrate binding and catalysis. In DdPhyA, two mobile loops that enclose substrates in the PHDs are conserved, but the C-terminal helix of the PHDs is strikingly absent. The combined results support the proposal that PHD homologues have evolved kinetic and structural features suited to their specific sensing roles.
Topics: Amino Acid Sequence; Animals; Binding Sites; Biocatalysis; Crystallography, X-Ray; Dictyostelium; Humans; Hydroxylation; Hypoxia-Inducible Factor 1, alpha Subunit; Kinetics; Molecular Dynamics Simulation; Oxygen; Prolyl Hydroxylases; Protein Structure, Tertiary; Protozoan Proteins; Recombinant Proteins; S-Phase Kinase-Associated Proteins; Sequence Alignment; Substrate Specificity; Toxoplasma
PubMed: 32934009
DOI: 10.1074/jbc.RA120.013998 -
Current Biology : CB May 2023Eukaryotic cells can undergo chemorepulsion, but the molecular mechanisms behind this phenomenon have remained unclear. Using Dictyostelium cells, a new study shows that...
Eukaryotic cells can undergo chemorepulsion, but the molecular mechanisms behind this phenomenon have remained unclear. Using Dictyostelium cells, a new study shows that competition of two ligands for the same receptors results in chemorepulsion, thus revealing a simple rule for eukaryotic cells to achieve negative chemotaxis.
Topics: Chemotaxis; Dictyostelium; Eukaryotic Cells
PubMed: 37160099
DOI: 10.1016/j.cub.2023.03.076 -
ELife May 2021Filopodia are thin, actin-based structures that cells use to interact with their environments. Filopodia initiation requires a suite of conserved proteins but the...
Filopodia are thin, actin-based structures that cells use to interact with their environments. Filopodia initiation requires a suite of conserved proteins but the mechanism remains poorly understood. The actin polymerase VASP and a MyTH-FERM (MF) myosin, DdMyo7 in amoeba, are essential for filopodia initiation. DdMyo7 is localized to dynamic regions of the actin-rich cortex. Analysis of VASP mutants and treatment of cells with anti-actin drugs shows that myosin recruitment and activation in requires localized VASP-dependent actin polymerization. Targeting of DdMyo7 to the cortex alone is not sufficient for filopodia initiation; VASP activity is also required. The actin regulator locally produces a cortical actin network that activates myosin and together they shape the actin network to promote extension of parallel bundles of actin during filopodia formation. This work reveals how filopodia initiation requires close collaboration between an actin-binding protein, the state of the actin cytoskeleton and MF myosin activity.
Topics: Actins; Cell Adhesion Molecules; Dictyostelium; Microfilament Proteins; Movement; Myosins; Phosphoproteins; Protozoan Proteins; Pseudopodia; Time Factors
PubMed: 34042588
DOI: 10.7554/eLife.68082 -
Small GTPases Sep 2019Rap1 belongs to the Ras family of small GTPases, which are involved in a multitude of cellular signal transduction pathways and have extensively been linked to cancer... (Review)
Review
Rap1 belongs to the Ras family of small GTPases, which are involved in a multitude of cellular signal transduction pathways and have extensively been linked to cancer biogenesis and metastasis. The small GTPase is activated in response to various extracellular and intracellular cues. Rap1 has conserved functions in amoeba and mammalian cells, which are important for cell polarity, substrate and cell-cell adhesion and other processes that involve the regulation of cytoskeletal dynamics. Moreover, our recent study has shown that Rap1 is required for the formation of the replication-permissive vacuole of an intracellular bacterial pathogen. Here we review the function and regulation of Rap1 in these distinct processes, and we discuss the underlying signal transduction pathways.
Topics: Animals; Bacterial Infections; Cytoskeleton; Dictyostelium; Humans; Neoplasm Metastasis; Neoplasm Proteins; Neoplasms; Protozoan Proteins; Shelterin Complex; Signal Transduction; Telomere-Binding Proteins
PubMed: 28632994
DOI: 10.1080/21541248.2017.1331721 -
BMC Biology Dec 2019Cellular functions can be regulated by cell-cell interactions that are influenced by extra-cellular, density-dependent signaling factors. Dictyostelium grow as...
BACKGROUND
Cellular functions can be regulated by cell-cell interactions that are influenced by extra-cellular, density-dependent signaling factors. Dictyostelium grow as individual cells in nutrient-rich sources, but, as nutrients become depleted, they initiate a multi-cell developmental program that is dependent upon a cell-density threshold. We hypothesized that novel secreted proteins may serve as density-sensing factors to promote multi-cell developmental fate decisions at a specific cell-density threshold, and use Dictyostelium in the identification of such a factor.
RESULTS
We show that multi-cell developmental aggregation in Dictyostelium is lost upon minimal (2-fold) reduction in local cell density. Remarkably, developmental aggregation response at non-permissive cell densities is rescued by addition of conditioned media from high-density, developmentally competent cells. Using rescued aggregation of low-density cells as an assay, we purified a single, 150-kDa extra-cellular protein with density aggregation activity. MS/MS peptide sequence analysis identified the gene sequence, and cells that overexpress the full-length protein accumulate higher levels of a development promoting factor (DPF) activity than parental cells, allowing cells to aggregate at lower cell densities; cells deficient for this DPF gene lack density-dependent developmental aggregation activity and require higher cell density for cell aggregation compared to WT. Density aggregation activity co-purifies with tagged versions of DPF and tag-affinity-purified DPF possesses density aggregation activity. In mixed development with WT, cells that overexpress DPF preferentially localize at centers for multi-cell aggregation and define cell-fate choice during cytodifferentiation. Finally, we show that DPF is synthesized as a larger precursor, single-pass transmembrane protein, with the p150 fragment released by proteolytic cleavage and ectodomain shedding. The TM/cytoplasmic domain of DPF possesses cell-autonomous activity for cell-substratum adhesion and for cellular growth.
CONCLUSIONS
We have purified a novel secreted protein, DPF, that acts as a density-sensing factor for development and functions to define local collective thresholds for Dictyostelium development and to facilitate cell-cell communication and multi-cell formation. Regions of high DPF expression are enriched at centers for cell-cell signal-response, multi-cell formation, and cell-fate determination. Additionally, DPF has separate cell-autonomous functions for regulation of cellular adhesion and growth.
Topics: Cell Communication; Dictyostelium; Protozoan Proteins
PubMed: 31791330
DOI: 10.1186/s12915-019-0714-9 -
Sub-cellular Biochemistry 2022Macropinocytosis is a nonspecific mechanism by which cells compulsively "drink" the surrounding extracellular fluids in order to feed themselves or sample the molecules...
Macropinocytosis is a nonspecific mechanism by which cells compulsively "drink" the surrounding extracellular fluids in order to feed themselves or sample the molecules therein, hence gaining information about their environment. This process is cell-intrinsically incompatible with the migration of many cells, implying that the two functions are antagonistic. The migrating cell uses a molecular switch to stop and explore its surrounding fluid by macropinocytosis, after which it employs the same molecular machinery to start migrating again to examine another location. This cycle of migration/macropinocytosis allows cells to explore tissues, and it is key to a range of physiological processes. Evidence of this evolutionarily conserved antagonism between the two processes can be found in several cell types-immune cells, for example, being particularly adept-and ancient organisms (e.g., the social amoeba Dictyostelium discoideum). How macropinocytosis and migration are negatively coupled is the subject of this chapter.
Topics: Cell Movement; Dictyostelium; Pinocytosis
PubMed: 35378704
DOI: 10.1007/978-3-030-94004-1_5 -
Cells Nov 2021The social amoeba provides an excellent model for research across a broad range of disciplines within biology. The organism diverged from the plant, yeast, fungi and... (Review)
Review
The social amoeba provides an excellent model for research across a broad range of disciplines within biology. The organism diverged from the plant, yeast, fungi and animal kingdoms around 1 billion years ago but retains common aspects found in these kingdoms. has a low level of genetic complexity and provides a range of molecular, cellular, biochemical and developmental biology experimental techniques, enabling multidisciplinary studies to be carried out in a wide range of areas, leading to research breakthroughs. Numerous laboratories within the United Kingdom employ as their core research model. This review introduces and then highlights research from several leading British research laboratories, covering their distinct areas of research, the benefits of using the model, and the breakthroughs that have arisen due to the use of as a tractable model system.
Topics: Animals; Biology; Dictyostelium; Drug Discovery; Models, Biological; Protein Processing, Post-Translational; Research; United Kingdom
PubMed: 34831258
DOI: 10.3390/cells10113036