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Journal of Cell Science Jul 2023During developmental and immune responses, cells move towards or away from some signals. Although much is known about chemoattraction, chemorepulsion (the movement of...
During developmental and immune responses, cells move towards or away from some signals. Although much is known about chemoattraction, chemorepulsion (the movement of cells away from a stimulus) remains poorly understood. Proliferating Dictyostelium discoideum cells secrete a chemorepellent protein called AprA. Examining existing knockout strains, we previously identified proteins required for AprA-induced chemorepulsion, and a genetic screen suggested that the enzyme phosphatidylinositol phosphate kinase A (PIPkinA, also known as Pik6) might also be needed for chemorepulsion. Here, we show that cells lacking PIPkinA are not repelled by AprA, and that this phenotype is rescued by expression of PIPkinA. To bias cell movement, AprA inhibits Ras activation at the side of the cell closest to the source of AprA, and we find that PIPkinA is required for AprA to inhibit Ras activation. PIPkinA decreases levels of phosphatidylinositol 4-phosphate [PI(4)P] and phosphatidylinositol (3,4,5)-trisphosphate [PI(3,4,5)P3], and possibly because of these effects, potentiates phagocytosis and inhibits cell proliferation. Cells lacking PIPkinA show normal AprA binding, suggesting that PIPkinA regulates chemorepulsion at a step between the AprA receptor and AprA inhibition of Ras activation.
Topics: Dictyostelium; Phosphates; Protozoan Proteins; Cell Proliferation; Genetic Testing
PubMed: 37259831
DOI: 10.1242/jcs.260541 -
IScience Jun 2023In chimeras, "cheaters" are strains that positively bias their contribution to the pool of spores, the reproductive cells resulting from development. On evolutionary...
In chimeras, "cheaters" are strains that positively bias their contribution to the pool of spores, the reproductive cells resulting from development. On evolutionary time scales, the selective advantage; thus, gained by cheaters is predicted to undermine collective functions whenever social behaviors are genetically determined. Genotypes; however, are not the sole determinant of spore bias, but the relative role of genetic and plastic differences in evolutionary success is unclear. Here, we study chimeras composed of cells harvested in different phases of population growth. We show that such heterogeneity induces frequency-dependent, plastic variation in spore bias. In genetic chimeras, the magnitude of such variation is not negligible and can even reverse the classification of a strain's social behavior. Our results suggest that differential cell mechanical properties can underpin, through biases emerging during aggregation, a "lottery" in strains' reproductive success that may counter the evolution of cheating.
PubMed: 37235054
DOI: 10.1016/j.isci.2023.106783 -
FEMS Microbiology Ecology May 2023Bacterial endosymbionts can provide benefits for their eukaryotic hosts, but it is often unclear if endosymbionts benefit from these relationships. The social amoeba...
Bacterial endosymbionts can provide benefits for their eukaryotic hosts, but it is often unclear if endosymbionts benefit from these relationships. The social amoeba Dictyostelium discoideum associates with three species of Paraburkholderia endosymbionts, including P. agricolaris and P. hayleyella. These endosymbionts can be costly to the host but are beneficial in certain contexts because they allow D. discoideum to carry prey bacteria through the dispersal stage. In experiments where no other species are present, P. hayleyella benefits from D. discoideum while P. agricolaris does not. However, the presence of other species may influence this symbiosis. We tested if P. agricolaris and P. hayleyella benefit from D. discoideum in the context of resource competition with Klebsiella pneumoniae, the typical laboratory prey of D. discoideum. Without D. discoideum, K. pneumoniae depressed the growth of both Paraburkholderia symbionts, consistent with competition. P. hayleyella was more harmed by interspecific competition than P. agricolaris. We found that P. hayleyella was rescued from competition by D. discoideum, while P. agricolaris was not. This may be because P. hayleyella is more specialized as an endosymbiont; it has a highly reduced genome compared to P. agricolaris and may have lost genes relevant for resource competition outside of its host.
Topics: Dictyostelium; Amoeba; Burkholderiaceae; Bacteria; Ecology
PubMed: 37226596
DOI: 10.1093/femsec/fiad055 -
MicroLife 2023Vesicular trafficking and membrane fusion are well-characterized, versatile, and sophisticated means of 'long range' intracellular protein and lipid delivery. Membrane... (Review)
Review
Vesicular trafficking and membrane fusion are well-characterized, versatile, and sophisticated means of 'long range' intracellular protein and lipid delivery. Membrane contact sites (MCS) have been studied in far less detail, but are crucial for 'short range' (10-30 nm) communication between organelles, as well as between pathogen vacuoles and organelles. MCS are specialized in the non-vesicular trafficking of small molecules such as calcium and lipids. Pivotal MCS components important for lipid transfer are the VAP receptor/tether protein, oxysterol binding proteins (OSBPs), the ceramide transport protein CERT, the phosphoinositide phosphatase Sac1, and the lipid phosphatidylinositol 4-phosphate (PtdIns(4)). In this review, we discuss how these MCS components are subverted by bacterial pathogens and their secreted effector proteins to promote intracellular survival and replication.
PubMed: 37223745
DOI: 10.1093/femsml/uqad018 -
Cell Death Discovery May 2023Glioblastomas are a highly aggressive cancer type which respond poorly to current pharmaceutical treatments, thus novel therapeutic approaches need to be investigated....
Glioblastomas are a highly aggressive cancer type which respond poorly to current pharmaceutical treatments, thus novel therapeutic approaches need to be investigated. One such approach involves the use of the bioactive natural product Tanshinone IIA (T2A) derived from the Chinese herb Danshen, where mechanistic insight for this anti-cancer agent is needed to validate its use. Here, we employ a tractable model system, Dictyostelium discoideum, to provide this insight. T2A potently inhibits cellular proliferation of Dictyostelium, suggesting molecular targets in this model. We show that T2A rapidly reduces phosphoinositide 3 kinase (PI3K) and protein kinase B (PKB) activity, but surprisingly, the downstream complex mechanistic target of rapamycin complex 1 (mTORC1) is only inhibited following chronic treatment. Investigating regulators of mTORC1, including PKB, tuberous sclerosis complex (TSC), and AMP-activated protein kinase (AMPK), suggests these enzymes were not responsible for this effect, implicating an additional molecular mechanism of T2A. We identify this mechanism as the increased expression of sestrin, a negative regulator of mTORC1. We further show that combinatory treatment using a PI3K inhibitor and T2A gives rise to a synergistic inhibition of cell proliferation. We then translate our findings to human and mouse-derived glioblastoma cell lines, where both a PI3K inhibitor (Paxalisib) and T2A reduces glioblastoma proliferation in monolayer cultures and in spheroid expansion, with combinatory treatment significantly enhancing this effect. Thus, we propose a new approach for cancer treatment, including glioblastomas, through combinatory treatment with PI3K inhibitors and T2A.
PubMed: 37202382
DOI: 10.1038/s41420-023-01462-6 -
Microbiology Spectrum Jun 2023
Erratum for Densi et al., "Synonymous and Nonsynonymous Substitutions in Dictyostelium discoideum Ammonium Transporter Are Necessary for Functional Complementation in Saccharomyces cerevisiae".
PubMed: 37199617
DOI: 10.1128/spectrum.01824-23 -
Cellular Signalling Aug 2023Protein kinases are major regulators of cellular processes, but the roles of most kinases remain unresolved. Dictyostelid social amoebas have been useful in identifying...
Protein kinases are major regulators of cellular processes, but the roles of most kinases remain unresolved. Dictyostelid social amoebas have been useful in identifying functions for 30% of its kinases in cell migration, cytokinesis, vesicle trafficking, gene regulation and other processes but their upstream regulators and downstream effectors are mostly unknown. Comparative genomics can assist to distinguish between genes involved in deeply conserved core processes and those involved in species-specific innovations, while co-expression of genes as evident from comparative transcriptomics can provide cues to the protein complement of regulatory networks. Genomes and developmental and cell-type specific transcriptomes are available for species that span the 0.5 billion years of evolution of Dictyostelia from their unicellular ancestors. In this work we analysed conservation and change in the abundance, functional domain architecture and developmental regulation of protein kinases across the 4 major taxon groups of Dictyostelia. All data are summarized in annotated phylogenetic trees of the kinase subtypes and accompanied by functional information of all kinases that were experimentally studied. We detected 393 different protein kinase domains across the five studied genomes, of which 212 were fully conserved. Conservation was highest (71%) in the previously defined AGC, CAMK, CK1, CMCG, STE and TKL groups and lowest (26%) in the "other" group of typical protein kinases. This was mostly due to species-specific single gene amplification of "other" kinases. Apart from the AFK and α-kinases, the atypical protein kinases, such as the PIKK and histidine kinases were also almost fully conserved. The phylogeny-wide developmental and cell-type specific expression profiles of the protein kinase genes were combined with profiles from the same transcriptomic experiments for the families of G-protein coupled receptors, small GTPases and their GEFs and GAPs, the transcription factors and for all genes that upon lesion generate a developmental defect. This dataset was subjected to hierarchical clustering to identify clusters of co-expressed genes that potentially act together in a signalling network. The work provides a valuable resource that allows researchers to identify protein kinases and other regulatory proteins that are likely to act as intermediates in a network of interest.
Topics: Dictyostelium; Phylogeny; Protein Kinases; Genome; Transcription Factors
PubMed: 37187217
DOI: 10.1016/j.cellsig.2023.110714 -
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 2023The amoeba-resistant bacterium causes Legionnaires' disease and employs a type IV secretion system (T4SS) to replicate in the unique, ER-associated -containing vacuole...
The amoeba-resistant bacterium causes Legionnaires' disease and employs a type IV secretion system (T4SS) to replicate in the unique, ER-associated -containing vacuole (LCV). The large fusion GTPase Sey1/atlastin is implicated in ER dynamics, ER-derived lipid droplet (LD) formation, and LCV maturation. Here, we employ cryo-electron tomography, confocal microscopy, proteomics, and isotopologue profiling to analyze LCV-LD interactions in the genetically tractable amoeba . Dually fluorescence-labeled producing LCV and LD markers revealed that Sey1 as well as the T4SS and the Ran GTPase activator LegG1 promote LCV-LD interactions. In vitro reconstitution using purified LCVs and LDs from parental or Δ mutant indicated that Sey1 and GTP promote this process. Sey1 and the fatty acid transporter FadL were implicated in palmitate catabolism and palmitate-dependent intracellular growth. Taken together, our results reveal that Sey1 and LegG1 mediate LD- and FadL-dependent fatty acid metabolism of intracellular .
Topics: Humans; Legionella pneumophila; GTP Phosphohydrolases; Macrophages; Dictyostelium; Lipid Droplets; Vacuoles; Legionella; Legionnaires' Disease; Bacterial Proteins
PubMed: 37158597
DOI: 10.7554/eLife.85142 -
Infection and Immunity May 2023For many years, Streptococcus anginosus has been considered a commensal colonizing the oral cavity, as well as the gastrointestinal and genitourinary tracts. However,...
For many years, Streptococcus anginosus has been considered a commensal colonizing the oral cavity, as well as the gastrointestinal and genitourinary tracts. However, recent epidemiological and clinical data designate this bacterium as an emerging opportunistic pathogen. Despite the reported pathogenicity of S. anginosus, the molecular mechanism underpinning its virulence is poorly described. Therefore, our goal was to develop and optimize efficient and simple infection models that can be applied to examine the virulence of S. anginosus and to study host-pathogen interactions. Using 23 S. anginosus isolates collected from different infections, including severe and superficial infections, as well as an attenuated strain devoid of CppA, we demonstrate for the first time that Dictyostelium discoideum is a suitable model for initial, fast, and large-scale screening of virulence. Furthermore, we found that another nonvertebrate animal model, Galleria mellonella, can be used to study the pathogenesis of S. anginosus infection, with an emphasis on the interactions between the pathogen and host innate immunity. Examining the profile of immune defense genes, including antimicrobial peptides, opsonins, regulators of nodulation, and inhibitors of proteases, by quantitative PCR (qPCR) we identified different immune response profiles depending on the S. anginosus strain. Using these models, we show that S. anginosus is resistant to the bactericidal activity of phagocytes, a phenomenon confirmed using human neutrophils. Notably, since we found that the data from these models corresponded to the clinical severity of infection, we propose their further application to studies of the virulence of S. anginosus.
Topics: Animals; Humans; Virulence; Dictyostelium; Streptococcus anginosus; Moths; Virulence Factors; Disease Models, Animal; Larva
PubMed: 37097148
DOI: 10.1128/iai.00016-23