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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 -
BMC Bioinformatics Jun 2017Dictyostelium discoideum, a soil-dwelling social amoeba, is a model for the study of numerous biological processes. Research in the field has benefited mightily from the...
BACKGROUND
Dictyostelium discoideum, a soil-dwelling social amoeba, is a model for the study of numerous biological processes. Research in the field has benefited mightily from the adoption of next-generation sequencing for genomics and transcriptomics. Dictyostelium biologists now face the widespread challenges of analyzing and exploring high dimensional data sets to generate hypotheses and discovering novel insights.
RESULTS
We present dictyExpress (2.0), a web application designed for exploratory analysis of gene expression data, as well as data from related experiments such as Chromatin Immunoprecipitation sequencing (ChIP-Seq). The application features visualization modules that include time course expression profiles, clustering, gene ontology enrichment analysis, differential expression analysis and comparison of experiments. All visualizations are interactive and interconnected, such that the selection of genes in one module propagates instantly to visualizations in other modules. dictyExpress currently stores the data from over 800 Dictyostelium experiments and is embedded within a general-purpose software framework for management of next-generation sequencing data. dictyExpress allows users to explore their data in a broader context by reciprocal linking with dictyBase-a repository of Dictyostelium genomic data. In addition, we introduce a companion application called GenBoard, an intuitive graphic user interface for data management and bioinformatics analysis.
CONCLUSIONS
dictyExpress and GenBoard enable broad adoption of next generation sequencing based inquiries by the Dictyostelium research community. Labs without the means to undertake deep sequencing projects can mine the data available to the public. The entire information flow, from raw sequence data to hypothesis testing, can be accomplished in an efficient workspace. The software framework is generalizable and represents a useful approach for any research community. To encourage more wide usage, the backend is open-source, available for extension and further development by bioinformaticians and data scientists.
Topics: Chromatin Immunoprecipitation; Cluster Analysis; Dictyostelium; High-Throughput Nucleotide Sequencing; Internet; Sequence Analysis, RNA; Transcriptome; User-Computer Interface
PubMed: 28578698
DOI: 10.1186/s12859-017-1706-9 -
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 -
Molecular Microbiology Jan 2023Mammalian professional phagocytic cells ingest and kill invading microorganisms and prevent the development of bacterial infections. Our understanding of the sequence of...
Mammalian professional phagocytic cells ingest and kill invading microorganisms and prevent the development of bacterial infections. Our understanding of the sequence of events that results in bacterial killing and permeabilization in phagosomes is still largely incomplete. In this study, we used the Dictyostelium discoideum amoeba as a model phagocyte to study the fate of the bacteria Klebsiella pneumoniae inside phagosomes. Our analysis distinguishes three consecutive phases: bacteria first lose their ability to divide (killing), then their cytosolic content is altered (permeabilization), and finally their DNA is degraded (digestion). Phagosomal acidification and production of free radicals are necessary for rapid killing, membrane-permeabilizing proteins BpiC and AlyL are required for efficient permeabilization. These results illustrate how a combination of genetic and microscopical tools can be used to finely dissect the molecular events leading to bacterial killing and permeabilization in a maturing phagosome.
Topics: Animals; Dictyostelium; Phagosomes; Klebsiella pneumoniae; Membrane Proteins; Bacteria; Mammals
PubMed: 36416195
DOI: 10.1111/mmi.15004 -
Cells Feb 2019The nucleoli of have a comparatively unique, non-canonical, localization adjacent to the inner nuclear membrane. The verified nucleolar proteins of this eukaryotic... (Review)
Review
The nucleoli of have a comparatively unique, non-canonical, localization adjacent to the inner nuclear membrane. The verified nucleolar proteins of this eukaryotic microbe are detailed while other potential proteins are introduced. Heat shock protein 32 (Hsp32), eukaryotic translation initiation factor 6 (eIF6), and tumour necrosis factor receptor-associated protein 1 (TRAP1) are essential for cell survival. NumA1, a breast cancer type 1 susceptibility protein-C Terminus domain-containing protein linked to cell cycle, functions in the regulation of nuclear number. The cell cycle checkpoint kinase 2 homologue forkhead-associated kinase A (FhkA) and BRG1-associated factor 60a homologue Snf12 are also discussed. While nucleoli appear homogeneous ultrastructurally, evidence for nucleolar subcompartments exists. Nucleolar localization sequences (NoLS) have been defined that target proteins to either the general nucleolar area or to a specific intranucleolar domain. Protein translocations during mitosis are protein-specific and support the multiple functions of the nucleolus. To enrich the picture, binding partners of NumA1, the most well-characterized nucleolar protein, are examined: nucleolar Ca-binding protein 4a (CBP4a), nuclear puromycin-sensitive aminopeptidase A (PsaA) and Snf12. The role of as a model for understanding the contribution of nucleolar proteins to various diseases and cellular stress is discussed throughout the review.
Topics: Amino Acid Sequence; Cell Nucleolus; Dictyostelium; Nuclear Proteins; Prions; Protein Binding; Protein Transport
PubMed: 30781559
DOI: 10.3390/cells8020167 -
Protist Feb 2018Dictyostelia is a monophyletic group of transiently multicellular (sorocarpic) amoebae, whose study is currently limited to laboratory culture. This tends to favour...
Dictyostelia is a monophyletic group of transiently multicellular (sorocarpic) amoebae, whose study is currently limited to laboratory culture. This tends to favour faster growing species with robust sorocarps, while species with smaller more delicate sorocarps constitute most of the group's taxonomic breadth. The number of known species is also small (∼150) given Dictyostelia's molecular depth and apparent antiquity (>600 myr). Nonetheless, dictyostelid sequences are rarely recovered in culture independent sampling (ciPCR) surveys. We developed ciPCR primers to specifically target dictyostelid small subunit (SSU or 18S) rDNA and tested them on total DNAs extracted from a wide range of soils from five continents. The resulting clone libraries show mostly dictyostelid sequences (∼90%), and phylogenetic analyses of these sequences indicate novel lineages in all four dictyostelid families and most genera. This is especially true for the species-rich Heterostelium and Dictyosteliaceae but also the less species-rich Raperosteliaceae. However, the most novel deep branches are found in two very species-poor taxa, including the deepest branch yet seen in the highly divergent Cavenderiaceae. These results confirm a deep hidden diversity of Dictyostelia, potentially including novel morphologies and developmental schemes. The primers and protocols presented here should also enable more comprehensive studies of dictyostelid ecology.
Topics: Biodiversity; DNA Primers; DNA, Protozoan; DNA, Ribosomal; Dictyostelium; Phylogeny; Polymerase Chain Reaction
PubMed: 29427837
DOI: 10.1016/j.protis.2017.12.005 -
Cells Mar 2019Cell-derived extracellular vesicles (EVs) are newly uncovered messengers for intercellular communication. They are released by almost all cell types in the three... (Review)
Review
Cell-derived extracellular vesicles (EVs) are newly uncovered messengers for intercellular communication. They are released by almost all cell types in the three kingdoms, Archeabacteria, Bacteria and Eukaryotes. They are known to mediate important biological functions and to be increasingly involved in cell physiology and in many human diseases, especially in oncology. The aim of this review is to recapitulate the current knowledge about EVs and to summarize our pioneering work about EVs. However, many challenges remain unsolved in the EV research field, before any EV application for theranostics (diagnosis, prognosis, and therapy) of human cancers, can be efficiently implemented in the clinics. might be an outstanding eukaryotic cell model for deciphering the utmost challenging problem of EV heterogeneity, and for unraveling the still mostly unknown mechanisms of their specific functions as mediators of intercellular communication.
Topics: Animals; Dictyostelium; Disease; Drug Delivery Systems; Extracellular Vesicles; Health; Humans; Models, Biological
PubMed: 30857191
DOI: 10.3390/cells8030225 -
Cells Feb 2019Nuclear lamins are nucleus-specific intermediate filaments (IF) found at the inner nuclear membrane (INM) of the nuclear envelope (NE). Together with nuclear envelope...
Nuclear lamins are nucleus-specific intermediate filaments (IF) found at the inner nuclear membrane (INM) of the nuclear envelope (NE). Together with nuclear envelope transmembrane proteins, they form the nuclear lamina and are crucial for gene regulation and mechanical robustness of the nucleus and the whole cell. Recently, we characterized NE81 as an evolutionarily conserved lamin-like protein, both on the sequence and functional level. Here, we show on the structural level that the NE81 is also capable of assembling into filaments, just as metazoan lamin filament assemblies. Using field-emission scanning electron microscopy, we show that NE81 expressed in oocytes forms filamentous structures with an overall appearance highly reminiscent of lamin B2. The in vitro assembly properties of recombinant His-tagged NE81 purified from extracts are very similar to those of metazoan lamins. Super-resolution stimulated emission depletion (STED) and expansion microscopy (ExM), as well as transmission electron microscopy of negatively stained purified NE81, demonstrated its capability of forming filamentous structures under low-ionic-strength conditions. These results recommend as a non-mammalian model organism with a well-characterized nuclear envelope involving all relevant protein components known in animal cells.
Topics: Animals; Cell Membrane Permeability; Dictyostelium; Nuclear Envelope; Oocytes; Protozoan Proteins; Xenopus
PubMed: 30781468
DOI: 10.3390/cells8020162