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1.
Nat Microbiol ; 7(3): 411-422, 2022 03.
Artigo em Inglês | MEDLINE | ID: mdl-35246664

RESUMO

Recent data support the hypothesis that Gram-positive bacteria (monoderms) arose from Gram-negative ones (diderms) through loss of the outer membrane (OM), but how this happened remains unknown. As tethering of the OM is essential for cell envelope stability in diderm bacteria, its destabilization may have been involved in this transition. In the present study, we present an in-depth analysis of the four known main OM-tethering systems across the Tree of Bacteria (ToB). We show that the presence of such systems follows the ToB with a bimodal distribution matching the deepest phylogenetic divergence between Terrabacteria and Gracilicutes. Whereas the lipoprotein peptidoglycan-associated lipoprotein (Pal) is restricted to the Gracilicutes, along with a more sporadic occurrence of OmpA, and Braun's lipoprotein is present only in a subclade of Gammaproteobacteria, diderm Terrabacteria display, as the main system, the OmpM protein. We propose an evolutionary scenario whereby OmpM represents a simple, ancestral OM-tethering system that was later replaced by one based on Pal after the emergence of the Lol machinery to deliver lipoproteins to the OM, with OmpA as a possible transition state. We speculate that the existence of only one main OM-tethering system in the Terrabacteria would have allowed the multiple OM losses specifically inferred in this clade through OmpM perturbation, and we provide experimental support for this hypothesis by inactivating all four ompM gene copies in the genetically tractable diderm Firmicute Veillonella parvula. High-resolution imaging and tomogram reconstructions reveal a non-lethal phenotype in which vast portions of the OM detach from the cells, forming huge vesicles with an inflated periplasm shared by multiple dividing cells. Together, our results highlight an ancient shift of OM-tethering systems in bacterial evolution and suggest a mechanism for OM loss and the multiple emergences of the monoderm phenotype from diderm ancestors.


Assuntos
Bactérias , Bactérias Gram-Positivas , Bactérias/genética , Bactérias Gram-Positivas/metabolismo , Lipoproteínas/genética , Lipoproteínas/metabolismo , Peptidoglicano/metabolismo , Periplasma/metabolismo , Filogenia
2.
Cell Host Microbe ; 26(6): 823-835.e11, 2019 12 11.
Artigo em Inglês | MEDLINE | ID: mdl-31761719

RESUMO

RNA-binding proteins (RBPs) perform key cellular activities by controlling the function of bound RNAs. The widely held assumption that RBPs are strictly intracellular has been challenged by the discovery of secreted RBPs. However, extracellular RBPs have been described in eukaryotes, while secreted bacterial RBPs have not been reported. Here, we show that the bacterial pathogen Listeria monocytogenes secretes a small RBP that we named Zea. We show that Zea binds a subset of L. monocytogenes RNAs, causing their accumulation in the extracellular medium. Furthermore, during L. monocytogenes infection, Zea binds RIG-I, the non-self-RNA innate immunity sensor, potentiating interferon-ß production. Mouse infection studies reveal that Zea affects L. monocytogenes virulence. Together, our results unveil that bacterial RNAs can be present extracellularly in association with RBPs, acting as "social RNAs" to trigger a host response during infection.


Assuntos
Proteína DEAD-box 58/metabolismo , Listeria monocytogenes/metabolismo , Proteínas de Ligação a RNA/metabolismo , Animais , Proteínas de Bactérias/metabolismo , Proteína DEAD-box 58/imunologia , Células HEK293 , Interações entre Hospedeiro e Microrganismos , Humanos , Imunidade Inata , Interferon beta/metabolismo , Listeria monocytogenes/imunologia , Listeria monocytogenes/patogenicidade , Camundongos , RNA Bacteriano/metabolismo , Transdução de Sinais/imunologia , Virulência/imunologia
3.
EMBO Rep ; 17(6): 858-73, 2016 06.
Artigo em Inglês | MEDLINE | ID: mdl-27215606

RESUMO

Mitochondria are essential eukaryotic organelles often forming intricate networks. The overall network morphology is determined by mitochondrial fusion and fission. Among the multiple mechanisms that appear to regulate mitochondrial fission, the ER and actin have recently been shown to play an important role by mediating mitochondrial constriction and promoting the action of a key fission factor, the dynamin-like protein Drp1. Here, we report that the cytoskeletal component septin 2 is involved in Drp1-dependent mitochondrial fission in mammalian cells. Septin 2 localizes to a subset of mitochondrial constrictions and directly binds Drp1, as shown by immunoprecipitation of the endogenous proteins and by pulldown assays with recombinant proteins. Depletion of septin 2 reduces Drp1 recruitment to mitochondria and results in hyperfused mitochondria and delayed FCCP-induced fission. Strikingly, septin depletion also affects mitochondrial morphology in Caenorhabditis elegans, strongly suggesting that the role of septins in mitochondrial dynamics is evolutionarily conserved.


Assuntos
GTP Fosfo-Hidrolases/metabolismo , Proteínas Associadas aos Microtúbulos/metabolismo , Mitocôndrias/metabolismo , Dinâmica Mitocondrial , Proteínas Mitocondriais/metabolismo , Septinas/metabolismo , Actomiosina/metabolismo , Evolução Biológica , Proteínas de Caenorhabditis elegans/genética , Proteínas de Caenorhabditis elegans/metabolismo , Dinaminas , Técnicas de Silenciamento de Genes , Inativação Gênica , Células HeLa , Humanos , Mitocôndrias/genética , Proteínas Mitocondriais/genética , Septinas/genética
4.
Science ; 331(6022): 1319-21, 2011 Mar 11.
Artigo em Inglês | MEDLINE | ID: mdl-21252314

RESUMO

Intracellular pathogens such as Listeria monocytogenes subvert cellular functions through the interaction of bacterial effectors with host components. Here we found that a secreted listerial virulence factor, LntA, could target the chromatin repressor BAHD1 in the host cell nucleus to activate interferon (IFN)-stimulated genes (ISGs). IFN-λ expression was induced in response to infection of epithelial cells with bacteria lacking LntA; however, the BAHD1-chromatin associated complex repressed downstream ISGs. In contrast, in cells infected with lntA-expressing bacteria, LntA prevented BAHD1 recruitment to ISGs and stimulated their expression. Murine listeriosis decreased in BAHD1(+/-) mice or when lntA was constitutively expressed. Thus, the LntA-BAHD1 interplay may modulate IFN-λ-mediated immune response to control bacterial colonization of the host.


Assuntos
Cromatina/metabolismo , Proteínas Cromossômicas não Histona/metabolismo , Interferons/metabolismo , Interleucinas/metabolismo , Listeria monocytogenes/patogenicidade , Listeriose/imunologia , Fatores de Virulência/metabolismo , Sequência de Aminoácidos , Animais , Linhagem Celular , Linhagem Celular Tumoral , Núcleo Celular/metabolismo , Regulação para Baixo , Perfilação da Expressão Gênica , Regulação da Expressão Gênica , Interações Hospedeiro-Patógeno , Humanos , Interferons/genética , Interferons/imunologia , Interleucinas/genética , Interleucinas/imunologia , Listeria monocytogenes/genética , Listeria monocytogenes/metabolismo , Listeriose/microbiologia , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Dados de Sequência Molecular , Transdução de Sinais , Fatores de Virulência/química , Fatores de Virulência/genética
5.
Cell Host Microbe ; 8(5): 433-44, 2010 Nov 18.
Artigo em Inglês | MEDLINE | ID: mdl-21075354

RESUMO

Actin-based motility is used by various pathogens for dissemination within and between cells. Yet host factors restricting this process have not been identified. Septins are GTP-binding proteins that assemble as filaments and are essential for cell division. However, their role during interphase has remained elusive. Here, we report that septin assemblies are recruited to different bacteria that polymerize actin. We observed that intracytosolic Shigella either become compartmentalized in septin cage-like structures or form actin tails. Inactivation of septin caging increases the number of Shigella with actin tails and enhances cell-to-cell spread. TNF-α, a host cytokine produced upon Shigella infection, stimulates septin caging and restricts actin tail formation and cell-to-cell spread. Finally, we show that septin cages entrap bacteria targeted to autophagy. Together, these results reveal an unsuspected mechanism of host defense that restricts dissemination of invasive pathogens.


Assuntos
Colo do Útero/microbiologia , Colo/microbiologia , Citosol/microbiologia , Interações Hospedeiro-Patógeno , Septinas/metabolismo , Shigella flexneri/patogenicidade , Actinas/metabolismo , Células CACO-2/imunologia , Células CACO-2/microbiologia , Células CACO-2/ultraestrutura , Colo do Útero/citologia , Colo/citologia , Feminino , Células HeLa/imunologia , Células HeLa/microbiologia , Células HeLa/ultraestrutura , Humanos , Shigella flexneri/metabolismo , Fator de Necrose Tumoral alfa/metabolismo
6.
Infect Immun ; 78(1): 204-9, 2010 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-19901060

RESUMO

Listeria monocytogenes is an intracellular bacterial pathogen that invades epithelial cells by subverting two cellular receptors, E-cadherin and Met. We recently identified type II phosphatidylinositol 4-kinases alpha and beta (PI4KIIalpha and PI4KIIbeta) as being required for bacterial entry downstream of Met. In this work, we investigated whether tetraspanins CD9, CD63, and CD81, which figure among the few described molecular partners of PI4KIIalpha, function as molecular adaptors recruiting PI4KIIalpha to the bacterial entry site. We observed by fluorescence microscopy that CD9, CD63, and CD81 are expressed and detected at the cellular surface and also within intracellular compartments, particularly in the case of CD63. In resting cells, colocalization of tetraspanins and PI4KIIalpha is detectable only in restricted areas of the perinuclear region. Upon infection with Listeria, endogenous CD9, CD63, and CD81 were recruited to the bacterial entry site but did not colocalize strictly with endogenous PI4KIIalpha. Live-cell imaging confirmed that tetraspanins and PI4KIIalpha do not follow the same recruitment dynamics to the Listeria entry site. Depletion of CD9, CD63, and CD81 levels by small interfering RNA demonstrated that CD81 is required for bacterial internalization, identifying for the first time a role for a member of the tetraspanin family in the entry of Listeria into target cells. Moreover, depletion of CD81 inhibits the recruitment of PI4KIIalpha but not that of the Met receptor to the bacterial entry site, suggesting that CD81 may act as a membrane organizer required for the integrity of signaling events occurring at Listeria entry sites.


Assuntos
Antígenos CD/metabolismo , Listeria monocytogenes/fisiologia , Antígenos CD/genética , Células Epiteliais , Regulação da Expressão Gênica/imunologia , Células HeLa , Humanos , Glicoproteínas de Membrana/genética , Glicoproteínas de Membrana/metabolismo , Antígenos de Histocompatibilidade Menor , Fosfotransferases (Aceptor do Grupo Álcool)/genética , Fosfotransferases (Aceptor do Grupo Álcool)/metabolismo , Glicoproteínas da Membrana de Plaquetas/genética , Glicoproteínas da Membrana de Plaquetas/metabolismo , Transporte Proteico , Tetraspanina 28 , Tetraspanina 29 , Tetraspanina 30
7.
Proc Natl Acad Sci U S A ; 106(33): 13826-31, 2009 Aug 18.
Artigo em Inglês | MEDLINE | ID: mdl-19666599

RESUMO

Gene silencing via heterochromatin formation plays a major role in cell differentiation and maintenance of homeostasis. Here we report the identification and characterization of a novel heterochromatinization factor in vertebrates, bromo adjacent homology domain-containing protein 1 (BAHD1). This nuclear protein interacts with HP1, MBD1, HDAC5, and several transcription factors. Through electron and immunofluorescence microscopy studies, we show that BAHD1 overexpression directs HP1 to specific nuclear sites and promotes the formation of large heterochromatic domains, which lack acetyl histone H4 and are enriched in H3 trimethylated at lysine 27 (H3K27me3). Furthermore, ectopically expressed BAHD1 colocalizes with the heterochromatic inactive X chromosome (Xi). The BAH domain is required for BAHD1 colocalization with H3K27me3, but not with the Xi chromosome. As highlighted by whole genome microarray analysis of BAHD1 knockdown cells, BAHD1 represses several proliferation and survival genes, in particular the insulin-like growth factor II gene (IGF2). When overexpressed, BAHD1 specifically binds the CpG-rich P3 promoter of IGF2, which increases MBD1 and HDAC5 targeting at this locus. This region contains DNA-binding sequences for the transcription factor SP1, with which BAHD1 coimmunoprecipitates. Collectively, these findings provide evidence that BAHD1 acts as a silencer by recruiting at specific promoters a set of proteins that coordinate heterochromatin assembly.


Assuntos
Proteínas Cromossômicas não Histona/fisiologia , Inativação Gênica , Heterocromatina/química , Proteínas Nucleares/genética , Proteínas Nucleares/fisiologia , Cromatina/química , Mapeamento Cromossômico , Ilhas de CpG , Heterocromatina/metabolismo , Histonas/química , Humanos , Fator de Crescimento Insulin-Like II/metabolismo , Lisina/química , Microscopia de Fluorescência/métodos , Modelos Genéticos , Ligação Proteica , Estrutura Terciária de Proteína , Transcrição Gênica
8.
J Biol Chem ; 284(17): 11613-21, 2009 Apr 24.
Artigo em Inglês | MEDLINE | ID: mdl-19234302

RESUMO

Septins are filament-forming GTPases implicated in several cellular functions, including cytokinesis. We previously showed that SEPT2, SEPT9, and SEPT11 colocalize with several bacteria entering into mammalian non-phagocytic cells, and SEPT2 was identified as essential for this process. Here, we investigated the function of SEPT11, an interacting partner of SEPT9 whose function is still poorly understood. In uninfected HeLa cells, SEPT11 depletion by siRNA increased cell size but surprisingly did not affect actin filament formation or the colocalization of SEPT9 with actin filaments. SEPT11 depletion increased Listeria invasion, and incubating SEPT11-depleted cells with beads coated with the Listeria surface protein InlB also led to increased entry as compared with control cells. Strikingly, as shown by fluorescence resonance energy transfer, the InlB-mediated stimulation of Met signaling remained intact in SEPT11-depleted cells. Taken together, our results show that SEPT11 is not required for the bacterial entry process and rather restricts its efficacy. Because SEPT2 is essential for the InlB-mediated entry of Listeria, but SEPT11 is not, our findings distinguish the roles of different mammalian septins.


Assuntos
Proteínas de Bactérias/metabolismo , Proteínas de Ciclo Celular/metabolismo , Listeria monocytogenes/patogenicidade , Proteínas de Membrana/metabolismo , Proteínas de Ciclo Celular/fisiologia , Membrana Celular/metabolismo , Transferência Ressonante de Energia de Fluorescência , GTP Fosfo-Hidrolases/metabolismo , Células HeLa , Humanos , Cinética , Listeria monocytogenes/metabolismo , Microscopia Confocal , Microscopia de Fluorescência/métodos , Modelos Biológicos , Monoéster Fosfórico Hidrolases/metabolismo , Septinas , Transdução de Sinais
9.
Proc Natl Acad Sci U S A ; 104(33): 13467-72, 2007 Aug 14.
Artigo em Inglês | MEDLINE | ID: mdl-17675409

RESUMO

Upon infection, pathogens reprogram host gene expression. In eukaryotic cells, genetic reprogramming is induced by the concerted activation/repression of transcription factors and various histone modifications that control DNA accessibility in chromatin. We report here that the bacterial pathogen Listeria monocytogenes induces a dramatic dephosphorylation of histone H3 as well as a deacetylation of histone H4 during early phases of infection. This effect is mediated by the major listerial toxin listeriolysin O in a pore-forming-independent manner. Strikingly, a similar effect also is observed with other toxins of the same family, such as Clostridium perfringens perfringolysin and Streptococcus pneumoniae pneumolysin. The decreased levels of histone modifications correlate with a reduced transcriptional activity of a subset of host genes, including key immunity genes. Thus, control of epigenetic regulation emerges here as an unsuspected function shared by several bacterial toxins, highlighting a common strategy used by intracellular and extracellular pathogens to modulate the host response early during infection.


Assuntos
Toxinas Bacterianas/química , Histonas/química , Acetilação , Células HeLa , Humanos , Listeria monocytogenes/fisiologia , Fosforilação , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Transcrição Gênica
10.
Mol Cell Neurosci ; 27(2): 151-62, 2004 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-15485771

RESUMO

Polysialic acid (PSA) on NCAM is an important modulator of cell-cell interactions during development and regeneration. Here we investigated whether PSA overexpression influences neural cell migration and myelination. We stably expressed a GFP-tagged polysialytransferase, PSTGFP, in mouse neurospheres and induced prolonged PSA synthesis. Using a chick xenograft assay for migration, we show that PSA can instruct precursor migration along the ventral pathway. PSA persistence did not change neural precursor multipotentiality in vitro but induced a delay in oligodendrocyte differentiation. PSTGFP+ precursors showed widespread engraftment in shiverer brain, closely similar to that observed with control precursors expressing a fluorescent protein. Initially, myelination by oligodendrocytes was delayed but, eventually, down-regulation of PSTGFP occurred, allowing myelination to proceed. Thus down-regulation of polysialyltransferases takes place even in cells where its RNA is under the control of a heterologous promoter and engineering PSA overexpression in neural precursors does not cause irreversible unphysiological effects.


Assuntos
Movimento Celular/fisiologia , Fibras Nervosas Mielinizadas/metabolismo , Molécula L1 de Adesão de Célula Nervosa/biossíntese , Neurônios/metabolismo , Ácidos Siálicos/biossíntese , Células-Tronco/metabolismo , Células 3T3 , Animais , Movimento Celular/efeitos dos fármacos , Células Cultivadas , Embrião de Galinha , Regulação da Expressão Gênica/fisiologia , Humanos , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Fibras Nervosas Mielinizadas/transplante , Molécula L1 de Adesão de Célula Nervosa/genética , Neurônios/transplante , Engenharia de Proteínas/métodos , Ácidos Siálicos/genética
11.
Glia ; 42(2): 139-48, 2003 Apr 15.
Artigo em Inglês | MEDLINE | ID: mdl-12655598

RESUMO

alpha-chemokines, which control the activation and directed migration of leukocytes, participate in the inflammatory processes in host defense response. One of the alpha-chemokines, CXCL12 or stromal cell-derived factor 1 (SDF-1), not only regulates cell growth and migration of hematopoietic stem cells but may also play a central role in brain development as we discuss here. SDF-1 indeed activates the CXCR4 receptor expressed in a variety of neural cells, and this signaling results in diverse biological effects. It enhances migration and proliferation of cerebellar granule cells, chemoattracts microglia, and stimulates cytokine production and glutamate release by astrocytes. Moreover, it elicits postsynaptic currents in Purkinje cells, triggers migration of cortical neuron progenitors, and produces pain by directly exciting nociceptive neurons. By modulating cell signaling and survival during neuroinflammation, SDF-1 may also play a role in the pathogenesis of brain tumors, experimental allergic encephalitis, and the nervous system dysfunction associated with acquired immunodeficiency syndrome.


Assuntos
Diferenciação Celular/fisiologia , Sistema Nervoso Central/embriologia , Sistema Nervoso Central/crescimento & desenvolvimento , Quimiocinas CXC/metabolismo , Neuroglia/metabolismo , Neurônios/metabolismo , Células-Tronco/metabolismo , Animais , Sistema Nervoso Central/metabolismo , Quimiocina CXCL12 , Quimiotaxia/fisiologia , Encefalite/metabolismo , Humanos , Neuroglia/citologia , Neurônios/citologia , Receptores CXCR4/metabolismo , Transdução de Sinais/fisiologia , Células-Tronco/citologia
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