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1.
mSphere ; 7(3): e0016022, 2022 06 29.
Artigo em Inglês | MEDLINE | ID: mdl-35587658

RESUMO

The protozoan Toxoplasma gondii is a highly successful obligate intracellular parasite that, upon invasion of its host cell, releases an array of host-modulating protein effectors to counter host defenses and further its own replication and dissemination. Early studies investigating the impact of T. gondii infection on host cell function revealed that this parasite can force normally quiescent cells to activate their cell cycle program. Prior reports by two independent groups identified the dense granule protein effector HCE1/TEEGR as being solely responsible for driving host cell transcriptional changes through its direct interaction with the cyclin E regulatory complex DP1 and associated transcription factors. Our group independently identified HCE1/TEEGR through the presence of distinct repeated regions found in a number of host nuclear targeted parasite effectors and verified its central role in initiating host cell cycle changes. Additionally, we report here the time-resolved kinetics of host cell cycle transition in response to HCE1/TEEGR, using the fluorescence ubiquitination cell cycle indicator reporter line (FUCCI), and reveal the existence of a block in S-phase progression and host DNA synthesis in several cell lines commonly used in the study of T. gondii. Importantly, we have observed that this S-phase block is not due to additional dense granule effectors but rather is dependent on the host cell line background and contact inhibition status of the host monolayer in vitro. This work highlights intriguing differences in the host response to reprogramming by the parasite and raises interesting questions regarding how parasite effectors differentially manipulate the host cell depending on the in vitro or in vivo context. IMPORTANCE Toxoplasma gondii chronically infects approximately one-third of the global population and can produce severe pathology in immunologically immature or compromised individuals. During infection, this parasite releases numerous host-targeted effector proteins that can dramatically alter the expression of a variety of host genes. A better understanding of parasite effectors and their host targets has the potential to not only provide ways to control infection but also inform us about our own basic biology. One host pathway that has been known to be altered by T. gondii infection is the cell cycle, and prior reports have identified a parasite effector, known as HCE1/TEEGR, as being responsible. In this report, we further our understanding of the kinetics of cell cycle transition induced by this effector and show that the capacity of HCE1/TEEGR to induce host cell DNA synthesis is dependent on both the cell type and the status of contact inhibition.


Assuntos
Toxoplasma , Inibição de Contato , DNA , Replicação do DNA , Humanos , Proteínas de Protozoários/genética , Proteínas de Protozoários/metabolismo , Toxoplasma/fisiologia
2.
J Cell Biol ; 221(4)2022 04 04.
Artigo em Inglês | MEDLINE | ID: mdl-35293964

RESUMO

Contact inhibition of locomotion (CIL) is a process that regulates cell motility upon collision with other cells. Improper regulation of CIL has been implicated in cancer cell dissemination. Here, we identify the cell adhesion molecule JAM-A as a central regulator of CIL in tumor cells. JAM-A is part of a multimolecular signaling complex in which tetraspanins CD9 and CD81 link JAM-A to αvß5 integrin. JAM-A binds Csk and inhibits the activity of αvß5 integrin-associated Src. Loss of JAM-A results in increased activities of downstream effectors of Src, including Erk1/2, Abi1, and paxillin, as well as increased activity of Rac1 at cell-cell contact sites. As a consequence, JAM-A-depleted cells show increased motility, have a higher cell-matrix turnover, and fail to halt migration when colliding with other cells. We also find that proper regulation of CIL depends on αvß5 integrin engagement. Our findings identify a molecular mechanism that regulates CIL in tumor cells and have implications on tumor cell dissemination.


Assuntos
Inibição de Contato , Fenofibrato , Adesão Celular , Moléculas de Adesão Celular/genética , Moléculas de Adesão Celular/metabolismo , Movimento Celular , Inibição de Contato/genética , Receptores de Vitronectina , Tetraspaninas
3.
Cells ; 11(3)2022 02 04.
Artigo em Inglês | MEDLINE | ID: mdl-35159360

RESUMO

Nonlimited proliferation is one of the most striking features of neoplastic cells. The basis of cell division is the sufficient presence of mass (amino acids) and energy (ATP and NADH). A sophisticated intracellular network permanently measures the mass and energy levels. Thus, in vivo restrictions in the form of amino acid, protein, or caloric restrictions strongly affect absolute lifespan and age-associated diseases such as cancer. The induction of permanent low energy metabolism (LEM) is essential in this process. The murine cell line L929 responds to methionine restriction (MetR) for a short time period with LEM at the metabolic level defined by a characteristic fingerprint consisting of the molecules acetoacetate, creatine, spermidine, GSSG, UDP-glucose, pantothenate, and ATP. Here, we used mass spectrometry (LC/MS) to investigate the influence of proliferation and contact inhibition on the energy status of cells. Interestingly, the energy status was essentially independent of proliferation or contact inhibition. LC/MS analyses showed that in full medium, the cells maintain active and energetic metabolism for optional proliferation. In contrast, MetR induced LEM independently of proliferation or contact inhibition. These results are important for cell behaviour under MetR and for the optional application of restrictions in cancer therapy.


Assuntos
Metionina , Neoplasias , Trifosfato de Adenosina , Aminoácidos , Animais , Proliferação de Células , Inibição de Contato , Metionina/metabolismo , Camundongos
4.
Int J Mol Sci ; 23(2)2022 Jan 10.
Artigo em Inglês | MEDLINE | ID: mdl-35054901

RESUMO

Mesenchymal stem cells (MSCs) can differentiate into endoderm lineages, especially parathyroid-hormone (PTH)-releasing cells. We have previously reported that tonsil-derived MSC (T-MSC) can differentiate into PTH-releasing cells (T-MSC-PTHCs), which restored the parathyroid functions in parathyroidectomy (PTX) rats. In this study, we demonstrate quality optimization by standardizing the differentiation rate for a better clinical application of T-MSC-PTHCs to overcome donor-dependent variation of T-MSCs. Quantitation results of PTH mRNA copy number in the differentiated cells and the PTH concentration in the conditioned medium confirmed that the differentiation efficiency largely varied depending on the cells from each donor. In addition, the differentiation rate of the cells from all the donors greatly improved when differentiation was started at a high cell density (100% confluence). The large-scale expression profiling of T-MSC-PTHCs by RNA sequencing indicated that those genes involved in exiting the differentiation and the cell cycle were the major pathways for the differentiation of T-MSC-PTHCs. Furthermore, the implantation of the T-MSC-PTHCs, which were differentiated at a high cell density embedded in hyaluronic acid, resulted in a higher serum PTH in the PTX model. This standardized efficiency of differentiation into PTHC was achieved by initiating differentiation at a high cell density. Our findings provide a potential solution to overcome the limitations due to donor-dependent variation by establishing a standardized differentiation protocol for the clinical application of T-MSC therapy in treating hypoparathyroidism.


Assuntos
Diferenciação Celular , Células-Tronco Mesenquimais/metabolismo , Tonsila Palatina/citologia , Hormônio Paratireóideo/biossíntese , Biomarcadores , Cálcio/metabolismo , Técnicas de Cultura de Células , Células Cultivadas , Inibição de Contato , Espaço Extracelular/metabolismo , Perfilação da Expressão Gênica , Redes Reguladoras de Genes , Sequenciamento de Nucleotídeos em Larga Escala , Humanos , Células-Tronco Mesenquimais/citologia
5.
Chem Biol Interact ; 351: 109763, 2022 Jan 05.
Artigo em Inglês | MEDLINE | ID: mdl-34852269

RESUMO

Air pollution has been linked to emphysema in chronic obstruction pulmonary disease (COPD). However, the underlying mechanisms in the development of emphysema due to air pollution remain unclear. The objective of this study was to investigate the role of components of the Hippo signaling pathway for E-cadherin-mediated contact inhibition of proliferation in the lungs after air pollution exposure. E-Cadherin-mediated contact inhibition of proliferation via the Hippo signaling pathway was investigated in Sprague-Dawley (SD) rats whole-body exposed to air pollution, and in alveolar epithelial A549 cells exposed to diesel exhaust particles (DEPs), E-cadherin-knockdown, and high-mobility group box 1 (HMGB1) treatment. Underlying epithelial differentiation, apoptosis, and senescence were also examined, and the interaction network among these proteins was examined. COPD lung sections were used to confirm the observations in rats. Expressions of HMGB1 and E-cadherin were negatively regulated in the lungs and A549 cells by air pollution, and this was confirmed by knockdown of E-cadherin and by treating A549 cells with HMGB1. Depletion of phosphorylated (p)-Yap occurred after exposure to air pollution and E-cadherin-knockdown, which resulted in decreases of SPC and T1α. Exposure to air pollution and E-cadherin-knockdown respectively downregulated p-Sirt1 and increased p53 levels in the lungs and in A549 cells. Moreover, the protein interaction network suggested that E-cadherin is a key activator in regulating Sirt1 and p53, as well as alveolar epithelial cell differentiation by SPC and T1α. Consistently, downregulation of E-cadherin, p-Yap, SPC, and T1α was observed in COPD alveolar regions with particulate matter (PM) deposition. In conclusion, our results indicated that E-cadherin-mediated cell-cell contact directly regulates the Hippo signaling pathway to control differentiation, cell proliferation, and senescence due to air pollution. Exposure to air pollution may initiate emphysema in COPD patients.


Assuntos
Poluição do Ar/efeitos adversos , Caderinas/metabolismo , Proliferação de Células/fisiologia , Inibição de Contato/fisiologia , Enfisema/metabolismo , /fisiologia , Células A549 , Animais , Apoptose/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Enfisema/induzido quimicamente , Proteína HMGB1/metabolismo , Humanos , Masculino , Mapas de Interação de Proteínas , Doença Pulmonar Obstrutiva Crônica/induzido quimicamente , Doença Pulmonar Obstrutiva Crônica/metabolismo , Ratos Sprague-Dawley , /metabolismo
6.
Nat Commun ; 12(1): 6998, 2021 12 01.
Artigo em Inglês | MEDLINE | ID: mdl-34853317

RESUMO

Bacteria have evolved toxins to outcompete other bacteria or to hijack host cell pathways. One broad family of bacterial polymorphic toxins gathers multidomain proteins with a modular organization, comprising a C-terminal toxin domain fused to a N-terminal domain that adapts to the delivery apparatus. Polymorphic toxins include bacteriocins, contact-dependent growth inhibition systems, and specialized Hcp, VgrG, PAAR or Rhs Type VI secretion (T6SS) components. We recently described and characterized Tre23, a toxin domain fused to a T6SS-associated Rhs protein in Photorhabdus laumondii, Rhs1. Here, we show that Rhs1 forms a complex with the T6SS spike protein VgrG and the EagR chaperone. Using truncation derivatives and cross-linking mass spectrometry, we demonstrate that VgrG-EagR-Rhs1 complex formation requires the VgrG C-terminal ß-helix and the Rhs1 N-terminal region. We then report the cryo-electron-microscopy structure of the Rhs1-EagR complex, demonstrating that the Rhs1 central region forms a ß-barrel cage-like structure that encapsulates the C-terminal toxin domain, and provide evidence for processing of the Rhs1 protein through aspartyl autoproteolysis. We propose a model for Rhs1 loading on the T6SS, transport and delivery into the target cell.


Assuntos
Toxinas Bacterianas/química , Toxinas Bacterianas/metabolismo , Photorhabdus/metabolismo , Sistemas de Secreção Tipo VI/química , Adaptação Fisiológica , Proteínas de Bactérias/química , Toxinas Bacterianas/classificação , Toxinas Bacterianas/genética , Bacteriocinas/química , Inibição de Contato , Microscopia Crioeletrônica , Espectrometria de Massas , Modelos Moleculares , Photorhabdus/genética , Sistemas de Secreção Tipo VI/genética , Sistemas de Secreção Tipo VI/metabolismo
7.
Eur Phys J E Soft Matter ; 44(11): 137, 2021 Nov 15.
Artigo em Inglês | MEDLINE | ID: mdl-34782959

RESUMO

Epithelial cell clusters often move collectively on a substrate. Mechanical signals play a major role in organizing this behavior. There are a number of experimental observations in these systems which await a comprehensive explanation. These include: the internal strains are tensile even for clusters that expand by proliferation; the tractions on the substrate are often confined to the edges of the cluster; there can exist density waves within the cluster; and for cells in an annulus, there is a transition between expanding clusters with proliferation and the case where cells fill the annulus and rotate around it. We formulate a mechanical model to examine these effects. We use a molecular clutch picture which allows "stalling"-inhibition of cell contraction by external forces. Stalled cells are passive from a physical point of view and the un-stalled cells are active. By attaching cells to the substrate and to each other, and taking into account contact inhibition of locomotion, we get a simple picture for many of these findings as well as predictions that could be tested.


Assuntos
Inibição de Contato , Modelos Biológicos , Movimento Celular
8.
mBio ; 12(5): e0253021, 2021 10 26.
Artigo em Inglês | MEDLINE | ID: mdl-34634941

RESUMO

Contact-dependent growth inhibition (CDI) systems enable the direct transfer of protein toxins between competing Gram-negative bacteria. CDI+ strains produce cell surface CdiA effector proteins that bind specific receptors on neighboring bacteria to initiate toxin delivery. Three classes of CdiA effectors that recognize different outer membrane protein receptors have been characterized in Escherichia coli to date. Here, we describe a fourth effector class that uses the lipopolysaccharide (LPS) core as a receptor to identify target bacteria. Selection for CDI-resistant target cells yielded waaF and waaP "deep-rough" mutants, which are unable to synthesize the full LPS core. The CDI resistance phenotypes of other waa mutants suggest that phosphorylated inner-core heptose residues form a critical CdiA recognition epitope. Class IV cdi loci also encode putative lysyl acyltransferases (CdiC) that are homologous to enzymes that lipidate repeats-in-toxin (RTX) cytolysins. We found that catalytically active CdiC is required for full target cell killing activity, and we provide evidence that the acyltransferase appends 3-hydroxydecanoate to a specific Lys residue within the CdiA receptor-binding domain. We propose that the lipid moiety inserts into the hydrophobic leaflet of lipid A to anchor CdiA interactions with the core oligosaccharide. Thus, LPS-binding CDI systems appear to have co-opted an RTX toxin-activating acyltransferase to increase the affinity of CdiA effectors for the target cell outer membrane. IMPORTANCE Contact-dependent growth inhibition (CDI) is a common form of interbacterial competition in which cells use CdiA effectors to deliver toxic proteins into their neighbors. CdiA recognizes target bacteria through specific receptor molecules on the cell surface. Here, we describe a new family of CdiA proteins that use lipopolysaccharide as a receptor to identify target bacteria. Target cell recognition is significantly enhanced by a unique fatty acid that is appended to the receptor-binding region of CdiA. We propose that the linked fatty acid inserts into the target cell outer membrane to stabilize the interaction. The CdiA receptor-binding region appears to mimic the biophysical properties of polymyxins, which are potent antibiotics used to disrupt the outer membranes of Gram-negative bacteria.


Assuntos
Inibição de Contato/fisiologia , Proteínas de Escherichia coli/metabolismo , Escherichia coli/crescimento & desenvolvimento , Escherichia coli/metabolismo , Proteínas de Membrana/metabolismo , Inibição de Contato/genética , Escherichia coli/genética , Proteínas de Escherichia coli/genética , Lipídeos , Proteínas de Membrana/genética , Ligação Proteica
9.
Sci Rep ; 11(1): 17759, 2021 09 07.
Artigo em Inglês | MEDLINE | ID: mdl-34493746

RESUMO

Inducing cardiac myocytes to proliferate is considered a potential therapy to target heart disease, however, modulating cardiac myocyte proliferation has proven to be a technical challenge. The Hippo pathway is a kinase signaling cascade that regulates cell proliferation during the growth of the heart. Inhibition of the Hippo pathway increases the activation of the transcription factors YAP/TAZ, which translocate to the nucleus and upregulate transcription of pro-proliferative genes. The Hippo pathway regulates the proliferation of cancer cells, pluripotent stem cells, and epithelial cells through a cell-cell contact-dependent manner, however, it is unclear if cell density-dependent cell proliferation is a consistent feature in cardiac myocytes. Here, we used cultured human iPSC-derived cardiac myocytes (hiCMs) as a model system to investigate this concept. hiCMs have a comparable transcriptome to the immature cardiac myocytes that proliferate during heart development in vivo. Our data indicate that a dense syncytium of hiCMs can regain cell cycle activity and YAP expression and activity when plated sparsely or when density is reduced through wounding. We found that combining two small molecules, XMU-MP-1 and S1P, increased YAP activity and further enhanced proliferation of low-density hiCMs. Importantly, these compounds had no effect on hiCMs within a dense syncytium. These data add to a growing body of literature that link Hippo pathway regulation with cardiac myocyte proliferation and demonstrate that regulation is restricted to cells with reduced contact inhibition.


Assuntos
Células-Tronco Pluripotentes Induzidas/citologia , Miócitos Cardíacos/citologia , Transdução de Sinais/fisiologia , Proteínas Adaptadoras de Transdução de Sinal/fisiologia , Sequência de Bases , Contagem de Células , Ciclo Celular/efeitos dos fármacos , Diferenciação Celular , Divisão Celular/efeitos dos fármacos , Células Cultivadas , Inibição de Contato/efeitos dos fármacos , Humanos , Células-Tronco Pluripotentes Induzidas/metabolismo , Lisofosfolipídeos/farmacologia , Miócitos Cardíacos/efeitos dos fármacos , Miócitos Cardíacos/metabolismo , Polimorfismo de Nucleotídeo Único , RNA/biossíntese , RNA/genética , Transdução de Sinais/efeitos dos fármacos , Esfingosina/análogos & derivados , Esfingosina/farmacologia , Sulfonamidas/farmacologia , Fatores de Transcrição/fisiologia
10.
Int J Mol Sci ; 22(17)2021 Aug 28.
Artigo em Inglês | MEDLINE | ID: mdl-34502237

RESUMO

Neural crest (NC) cells are highly migratory cells that contribute to various vertebrate tissues, and whose migratory behaviors resemble cancer cell migration and invasion. Information exchange via dynamic NC cell-cell contact is one mechanism by which the directionality of migrating NC cells is controlled. One transmembrane protein that is most likely involved in this process is protein tyrosine kinase 7 (PTK7), an evolutionary conserved Wnt co-receptor that is expressed in cranial NC cells and several tumor cells. In Xenopus, Ptk7 is required for NC migration. In this study, we show that the Ptk7 protein is dynamically localized at cell-cell contact zones of migrating Xenopus NC cells and required for contact inhibition of locomotion (CIL). Using deletion constructs of Ptk7, we determined that the extracellular immunoglobulin domains of Ptk7 are important for its transient accumulation and that they mediate homophilic binding. Conversely, we found that ectopic expression of Ptk7 in non-NC cells was able to prevent NC cell invasion. However, deletion of the extracellular domains of Ptk7 abolished this effect. Thus, Ptk7 is sufficient at protecting non-NC tissue from NC cell invasion, suggesting a common role of PTK7 in contact inhibition, cell invasion, and tissue integrity.


Assuntos
Moléculas de Adesão Celular/metabolismo , Diferenciação Celular , Movimento Celular , Inibição de Contato , Neoplasias Pulmonares/metabolismo , Crista Neural/metabolismo , Receptores Proteína Tirosina Quinases/metabolismo , Animais , Polaridade Celular , Humanos , Neoplasias Pulmonares/patologia , Xenopus laevis
11.
PLoS One ; 16(8): e0254697, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34424918

RESUMO

The PAF complex (PAFC) coordinates transcription elongation and mRNA processing and its CDC73/parafibromin subunit functions as a tumour suppressor. The NF2/Merlin tumour suppressor functions both at the cell cortex and nucleus and is a key mediator of contact inhibition but the molecular mechanisms remain unclear. In this study we have used affinity proteomics to identify novel Merlin interacting proteins and show that Merlin forms a complex with multiple proteins involved in RNA processing including the PAFC and the CHD1 chromatin remodeller. Tumour-derived inactivating mutations in both Merlin and the CDC73 PAFC subunit mutually disrupt their interaction and growth suppression by Merlin requires CDC73. Merlin interacts with the PAFC in a cell density-dependent manner and we identify a role for FAT cadherins in regulating the Merlin-PAFC interaction. Our results suggest that in addition to its function within the Hippo pathway, Merlin is part of a tumour suppressor network regulated by cell-cell adhesion which coordinates post-initiation steps of the transcription cycle of genes mediating contact inhibition.


Assuntos
Adesão Celular/genética , DNA Helicases/genética , Proteínas de Ligação a DNA/genética , Neoplasias/genética , Neurofibromina 2/genética , Proteínas Supressoras de Tumor/genética , Proliferação de Células/genética , Cromatina/genética , Montagem e Desmontagem da Cromatina/genética , Inibição de Contato/genética , Regulação da Expressão Gênica/genética , Células HEK293 , Humanos , Neoplasias/patologia , Ligação Proteica/genética , Mapas de Interação de Proteínas/genética , Transdução de Sinais/genética
12.
Life Sci Alliance ; 4(10)2021 10.
Artigo em Inglês | MEDLINE | ID: mdl-34404733

RESUMO

Contact inhibition is a key cellular phenomenon that prevents cells from hyper-proliferating upon reaching confluence. Although not fully characterized, a critical driver of this process is the Hippo signaling pathway, whose downstream effector yes-associated protein plays pivotal roles in cell growth and differentiation. Here, we provide evidence that the E3 ligase WWP1 (WW-domain containing protein 1) mono-ubiquitinates AMOTL2 (angiomotin-like 2) at K347 and K408. Mono-ubiquitinated AMOTL2, in turn, interacts with the kinase LATS2, which facilitates recruitment of the upstream Hippo pathway component SAV1 and ultimately promotes yes-associated protein phosphorylation and subsequent cytoplasmic sequestration and/or degradation. Furthermore, contact inhibition induced by high cell density promoted the localization and stabilization of WWP1 at cell junctions, where it interacted with Crumbs polarity proteins. Notably, the Crumbs complex was functionally important for AMOTL2 mono-ubiquitination and LATS activation under high cell density conditions. These findings delineate a functionally important molecular mechanism in which AMOTL2 mono-ubiquitination by WWP1 at cell junctions and LATS activation are tightly coupled to upstream cell density cues.


Assuntos
/metabolismo , Inibição de Contato , Ubiquitina-Proteína Ligases/metabolismo , /genética , Inibição de Contato/genética , Ativação Enzimática , Humanos , Fosforilação , Ligação Proteica , Transporte Proteico , Proteínas Supressoras de Tumor/genética , Proteínas Supressoras de Tumor/metabolismo , Ubiquitina-Proteína Ligases/genética , Ubiquitinação
13.
FASEB J ; 35(9): e21742, 2021 09.
Artigo em Inglês | MEDLINE | ID: mdl-34403506

RESUMO

Withdrawal from contact inhibition is necessary for epithelial cancer precursor cells to initiate cell growth and motility. Nevertheless, little is understood about the mechanism for the sudden initiation of cell growth under static conditions. We focused on cellular junctions as one region where breaking out of contact inhibition occurs. In well-differentiated endometrial cancer cells, Sawano, the ligand administration for tricellular tight junction protein LSR, which transiently decreased the robust junction property, caused an abrupt increase in cell motility and consequent excessive multilayered cell growth despite being under contact inhibition conditions. We observed that macropinocytosis essentially and temporarily occurred as an antecedent event for the above process at intercellular junctions without disruption of the junction apparatus but not at the apical plasma membrane. Collectively, we concluded that the formation of macropinocytosis, which is derived from tight junction-mediated signaling, was triggered for the initiation of cell growth in static precancerous epithelium.


Assuntos
Adesão Celular , Inibição de Contato , Pinocitose , Receptores de Lipoproteínas/metabolismo , Fatores de Transcrição/metabolismo , Toxinas Bacterianas/farmacologia , Sítios de Ligação , Processos de Crescimento Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Movimento Celular/efeitos dos fármacos , Humanos , Junções Intercelulares/efeitos dos fármacos , Junções Intercelulares/metabolismo , Proteínas Quinases JNK Ativadas por Mitógeno/metabolismo , Fenótipo , Pinocitose/efeitos dos fármacos , Transporte Proteico , Vacúolos/efeitos dos fármacos , Vacúolos/metabolismo , Proteínas rac de Ligação ao GTP/metabolismo
14.
Gut Microbes ; 13(1): 1959841, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34455923

RESUMO

Complex inter-bacterial interactions largely influence the structure and function of the gut microbial community. Though several host-associated phenomena have often been shown to be involved in the stability, structure, and function of the gut microbial community, the implication of contact-dependent and contact-independent inter-bacterial interactions has been overlooked. Such interactions are tightly governed at multiple layers through several extracellular organelles, including contact-dependent inhibition (CDI), nanotubes, type VI secretion system (T6SS), and membrane vesicles (MVs). Recent advancements in molecular techniques have revealed that such extracellular organelles function beyond exhibiting competitive behavior and are also involved in manifesting cooperative behaviors. Cooperation between bacteria occurs through the sharing of several beneficial molecules including nucleic acids, proteins, metabolites, and nutrients among the members of the community, while competition occurs by means of multiple toxins. Intrinsic coordination between contact-dependent and contact-independent mechanisms collectively provides a fitness advantage and increased colonization resistance to the gut microbiota, where molecular trafficking plays a key role. This review is intended to provide a comprehensive view of the salient features of the different bacterial interactions and to highlight how microbiota deploy multifaceted organelles, for exerting both cooperative and competitive behaviors. We discuss the current knowledge of bacterial molecular trafficking and its impact on shaping the gut microbial community.


Assuntos
Bactérias/metabolismo , Microbioma Gastrointestinal/fisiologia , Organelas/fisiologia , Percepção de Quorum/fisiologia , Transdução de Sinais/fisiologia , Bactérias/classificação , Bactérias/genética , Inibição de Contato/fisiologia , Interações Microbianas/fisiologia , Nanotubos , Sistemas de Secreção Tipo VI/fisiologia
15.
Proc Natl Acad Sci U S A ; 118(12)2021 03 23.
Artigo em Inglês | MEDLINE | ID: mdl-33737392

RESUMO

Contact inhibition of locomotion (CIL), in which cells repolarize and move away from contact, is now established as a fundamental driving force in development, repair, and disease biology. Much of what we know of CIL stems from studies on two-dimensional (2D) substrates that do not provide an essential biophysical cue-the curvature of extracellular matrix fibers. We discover rules controlling outcomes of cell-cell collisions on suspended nanofibers and show them to be profoundly different from the stereotyped CIL behavior on 2D substrates. Two approaching cells attached to a single fiber do not repolarize upon contact but rather usually migrate past one another. Fiber geometry modulates this behavior; when cells attach to two fibers, reducing their freedom to reorient, only one cell repolarizes on contact, leading to the cell pair migrating as a single unit. CIL outcomes also change when one cell has recently divided and moves with high speed-cells more frequently walk past each other. Our computational model of CIL in fiber geometries reproduces the core qualitative results of the experiments robustly to model parameters. Our model shows that the increased speed of postdivision cells may be sufficient to explain their increased walk-past rate. We also identify cell-cell adhesion as a key mediator of collision outcomes. Our results suggest that characterizing cell-cell interactions on flat substrates, channels, or micropatterns is not sufficient to predict interactions in a matrix-the geometry of the fiber can generate entirely new behaviors.


Assuntos
Técnicas de Cultura de Células , Movimento Celular , Fenômenos Fisiológicos Celulares , Inibição de Contato , Nanofibras , Matriz Extracelular/metabolismo
16.
mBio ; 12(1)2021 02 02.
Artigo em Inglês | MEDLINE | ID: mdl-33531386

RESUMO

The C-terminal (CT) toxin domains of contact-dependent growth inhibition (CDI) CdiA proteins target Gram-negative bacteria and must breach both the outer and inner membranes of target cells to exert growth inhibitory activity. Here, we examine two CdiA-CT toxins that exploit the bacterial general protein secretion machinery after delivery into the periplasm. A Ser281Phe amino acid substitution in transmembrane segment 7 of SecY, the universally conserved channel-forming subunit of the Sec translocon, decreases the cytotoxicity of the membrane depolarizing orphan10 toxin from enterohemorrhagic Escherichia coli EC869. Target cells expressing secYS281F and lacking either PpiD or YfgM, two SecY auxiliary factors, are fully protected from CDI-mediated inhibition either by CdiA-CTo10 EC869 or by CdiA-CTGN05224, the latter being an EndoU RNase CdiA toxin from Klebsiella aerogenes GN05224 that has a related cytoplasm entry domain. RNase activity of CdiA-CTGN05224 was reduced in secYS281F target cells and absent in secYS281F ΔppiD or secYS281F ΔyfgM target cells during competition co-cultures. Importantly, an allele-specific mutation in secY (secYG313W ) renders ΔppiD or ΔyfgM target cells specifically resistant to CdiA-CTGN05224 but not to CdiA-CTo10 EC869, further suggesting a direct interaction between SecY and the CDI toxins. Our results provide genetic evidence of a unique confluence between the primary cellular export route for unfolded polypeptides and the import pathways of two CDI toxins.IMPORTANCE Many bacterial species interact via direct cell-to-cell contact using CDI systems, which provide a mechanism to inject toxins that inhibit bacterial growth into one another. Here, we find that two CDI toxins, one that depolarizes membranes and another that degrades RNA, exploit the universally conserved SecY translocon machinery used to export proteins for target cell entry. Mutations in genes coding for members of the Sec translocon render cells resistant to these CDI toxins by blocking their movement into and through target cell membranes. This work lays the foundation for understanding how CDI toxins interact with the protein export machinery and has direct relevance to development of new antibiotics that can penetrate bacterial cell envelopes.


Assuntos
Proteínas de Escherichia coli/metabolismo , Proteínas de Membrana/metabolismo , Canais de Translocação SEC/genética , Inibição de Contato , Mutação , Transporte Proteico , Canais de Translocação SEC/fisiologia
17.
Biochem Biophys Res Commun ; 546: 65-73, 2021 03 26.
Artigo em Inglês | MEDLINE | ID: mdl-33571906

RESUMO

The occurrence and development of tumors cannot be separated from the influence of differentiation at different stages and levels. Our study found that E-cadherin was significantly increased in cell model induced by sodium butyrate and cell density, while METTL3, METTL16 and WTAP were decreased during the differentiation of cells. In the clinicopathological tissues, E-cadherin was low expressed in poorly differentiated tumor tissues and above three regulators were highly expressed in poorly differentiated tissues. At the levels of clinicopathological differentiation, tissue differentiation and cell differentiation, the result indicated that the poor prognosis of colorectal cancer (CRC) may be closely related to high expression of total m6A level and high expression of METTL3, METTL16 and WTAP.


Assuntos
Adenosina/análogos & derivados , Diferenciação Celular , Neoplasias Colorretais/metabolismo , Neoplasias Colorretais/patologia , Adenosina/metabolismo , Ácido Butírico/farmacologia , Proteínas de Ciclo Celular , Diferenciação Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Neoplasias Colorretais/genética , Inibição de Contato/efeitos dos fármacos , Feminino , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Humanos , Masculino , Metiltransferases , Pessoa de Meia-Idade , Biossíntese de Proteínas/efeitos dos fármacos , Fatores de Processamento de RNA , Transcrição Genética/efeitos dos fármacos
18.
Int J Mol Sci ; 21(21)2020 Oct 27.
Artigo em Inglês | MEDLINE | ID: mdl-33121148

RESUMO

Over millions of years of evolution, bacteria have developed complex strategies for intra-and interspecies interactions and competition for ecological niches and resources. Contact-dependent growth inhibition systems (CDI) are designed to realize a direct physical contact of one bacterial cell with other cells in proximity via receptor-mediated toxin delivery. These systems are found in many microorganisms including clinically important human pathogens. The main purpose of these systems is to provide competitive advantages for the growth of the population. In addition, non-competitive roles for CDI toxin delivery systems including interbacterial signal transduction and mediators of bacterial collaboration have been suggested. In this review, our goal was to systematize the recent findings on the structure, mechanisms, and purpose of CDI systems in bacterial populations and discuss the potential biological and evolutionary impact of CDI-mediated interbacterial competition and/or cooperation.


Assuntos
Bactérias/crescimento & desenvolvimento , Bactérias/metabolismo , Proteínas de Bactérias/metabolismo , Biofilmes/crescimento & desenvolvimento , Inibição de Contato , Interações Microbianas
19.
Elife ; 92020 04 30.
Artigo em Inglês | MEDLINE | ID: mdl-32352381

RESUMO

Biophysical mechanisms underlying collective cell migration of eukaryotic cells have been studied extensively in recent years. One mechanism that induces cells to correlate their motions is contact inhibition of locomotion, by which cells migrating away from the contact site. Here, we report that tail-following behavior at the contact site, termed contact following locomotion (CFL), can induce a non-trivial collective behavior in migrating cells. We show the emergence of a traveling band showing polar order in a mutant Dictyostelium cell that lacks chemotactic activity. We find that CFL is the cell-cell interaction underlying this phenomenon, enabling a theoretical description of how this traveling band forms. We further show that the polar order phase consists of subpopulations that exhibit characteristic transversal motions with respect to the direction of band propagation. These findings describe a novel mechanism of collective cell migration involving cell-cell interactions capable of inducing traveling band with polar order.


The cells of animals and many other living things are able to migrate together in groups. This collective cell migration plays crucial roles in many processes in animals such as forming organs and limbs, and healing wounds. A soil-dwelling amoeba called Dictyostelium discoideum ­ or just Dicty for short ­ is commonly used as a model to study how groups of cells migrate collectively. Individual Dicty cells may live alone but sometimes many cells come together to form a larger mobile structure called a "slug". Chemical signals coordinate how the cells collectively migrate to form the multicellular slug. Mutant Dicty cells that lack these chemical signal processes can still move together as a band that travels across a surface. This movement resembles a type of collective motion that has previously been observed in physics experiments using self-propelled particles. However, it remains unclear how this collective behavior works. Hayakawa et al. have now combined genetics, cell biology and computational approaches to study how groups of the mutant Dicty cells migrate together. The experiments showed that the traveling band is dynamically maintained by cells joining or leaving, and that this turnover is caused by simple interactions between the cells known as "contact following locomotion". Contact following locomotion has been also reported in mammalian cells so the findings of Hayakawa et al. may aid research into how animals develop and how errors in cell migration may lead to diseases. Further studies are required to find out whether other cells showing contact following locomotion also travel in a band.


Assuntos
Comunicação Celular , Movimento Celular , Inibição de Contato , Dictyostelium/fisiologia , Dictyostelium/genética , Microscopia de Vídeo , Modelos Biológicos , Mutação , Análise de Célula Única , Fatores de Tempo
20.
Sci Rep ; 10(1): 6713, 2020 04 21.
Artigo em Inglês | MEDLINE | ID: mdl-32317692

RESUMO

Contact inhibition is a cell property that limits the migration and proliferation of cells in crowded environments. Here we investigate the growth dynamics of a cell colony composed of migrating and proliferating cells on a substrate using a minimal model that incorporates the mechanisms of contact inhibition of locomotion and proliferation. We find two distinct regimes. At early times, when contact inhibition is weak, the colony grows exponentially in time, fully characterised by the proliferation rate. At long times, the colony boundary moves at a constant speed, determined only by the migration speed of a single cell and independent of the proliferation rate. Further, the model demonstrates how cell-cell alignment speeds up colony growth. Our model illuminates how simple local mechanical interactions give rise to contact inhibition, and from this, how cell colony growth is self-organised and controlled on a local level.


Assuntos
Inibição de Contato , Animais , Adesão Celular , Ciclo Celular , Movimento Celular , Proliferação de Células , Forma Celular , Simulação por Computador , Modelos Biológicos
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