Fusion of Chlamydia trachomatis-Containing Inclusions Is Inhibited at Low Temperatures and Requires Bacterial Protein Synthesis

ABSTRACT The human pathogen Chlamydia trachomatis is an obligate intracellular bacterium with a unique developmental cycle. Within the host cell cytoplasm, it resides within a membrane-bound compartment, the inclusion. A distinguishing characteristic of the C. trachomatis life cycle is the fusion of the chlamydia-containing inclusions with each other in the host cell cytoplasm. We report that fusion of inclusions does not occur at 32°C in multiple mammalian cell lines and with three different serovars of C. trachomatis. The inhibition of fusion was inclusion specific; the fusion with sphingolipid-containing secretory vesicles and the interaction with early endosomes were unaffected by incubation at 32°C. The inhibition of fusion of the inclusions was not primarily the result of delayed maturation of the inclusion, as infectious progeny was produced in host cells incubated at 32°C, and the unfused inclusions remained competent to fuse up to 48 h postinfection. The ability to reverse the inhibition of fusion by shifting the infected cells from 32 to 37°C allowed the measurement of the rate and the time of fusion of the inclusions after entry of the bacteria. Most significantly, we demonstrate that fusion of inclusions with each other requires bacterial protein synthesis and that the required bacterial protein(s) is present, but inactive or not secreted, at 32°C.

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Localization of Chlamydia trachomatis hypothetical protein CT311 in host cell cytoplasm

Microbial Pathogenesis ◽

10.1016/j.micpath.2011.05.002 ◽

2011 ◽

Vol 51 (3) ◽

pp. 101-109 ◽

Cited By ~ 18

Author(s):

Lei Lei ◽

Manli Qi ◽

Nicole Budrys ◽

Robert Schenken ◽

Guangming Zhong

Keyword(s):

Chlamydia Trachomatis ◽

Host Cell ◽

Hypothetical Protein ◽

Cell Cytoplasm ◽

Host Cell Cytoplasm

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Chlamydia trachomatis GlgA Is Secreted into Host Cell Cytoplasm

PLoS ONE ◽

10.1371/journal.pone.0068764 ◽

2013 ◽

Vol 8 (7) ◽

pp. e68764 ◽

Cited By ~ 27

Author(s):

Chunxue Lu ◽

Lei Lei ◽

Bo Peng ◽

Lingli Tang ◽

Honglei Ding ◽

...

Keyword(s):

Chlamydia Trachomatis ◽

Host Cell ◽

Cell Cytoplasm ◽

Host Cell Cytoplasm

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Toxoplasma gondii reorganizes the host cell architecture during spontaneous cyst formation in vitro

Parasitology ◽

10.1017/s0031182017002050 ◽

2017 ◽

Vol 145 (8) ◽

pp. 1027-1038 ◽

Cited By ~ 5

Author(s):

T. C. Paredes-Santos ◽

E. S. Martins-Duarte ◽

W. de Souza ◽

M. Attias ◽

R. C. Vommaro

Keyword(s):

Toxoplasma Gondii ◽

Host Cell ◽

Cyst Wall ◽

Cyst Formation ◽

Acute Stage ◽

Host Cells ◽

Cell Cytoplasm ◽

Chronic Stage ◽

Host Cell Cytoplasm

AbstractToxoplasma gondii is an intracellular protozoan parasite that causes toxoplasmosis, a prevalent infection related to abortion, ocular diseases and encephalitis in immuno-compromised individuals. In the untreatable (and life-long) chronic stage of toxoplasmosis, parasitophorous vacuoles (PVs, containing T. gondii tachyzoites) transform into tissue cysts, containing slow-dividing bradyzoite forms. While acute-stage infection with tachyzoites involves global rearrangement of the host cell cytoplasm, focused on favouring tachyzoite replication, the cytoplasmic architecture of cells infected with cysts had not been described. Here, we characterized (by fluorescence and electron microscopy) the redistribution of host cell structures around T. gondii cysts, using a T. gondii strain (EGS) with high rates of spontaneous cystogenesis in vitro. Microtubules and intermediate filaments (but not actin microfilaments) formed a ‘cage’ around the cyst, and treatment with taxol (to inhibit microtubule dynamics) favoured cystogenesis. Mitochondria, which appeared adhered to the PV membrane, were less closely associated with the cyst wall. Endoplasmic reticulum (ER) profiles were intimately associated with folds in the cyst wall membrane. However, the Golgi complex was not preferentially localized relative to the cyst, and treatment with tunicamycin or brefeldin A (to disrupt Golgi or ER function, respectively) had no significant effect on cystogenesis. Lysosomes accumulated around cysts, while early and late endosomes were more evenly distributed in the cytoplasm. The endocytosis tracer HRP (but not BSA or transferrin) reached bradyzoites after uptake by infected host cells. These results suggest that T. gondii cysts reorganize the host cell cytoplasm, which may fulfil specific requirements of the chronic stage of infection.

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The Hydrophilic Translocator for Vibrio parahaemolyticus, T3SS2, Is Also Translocated

Infection and Immunity ◽

10.1128/iai.00402-12 ◽

2012 ◽

Vol 80 (8) ◽

pp. 2940-2947 ◽

Cited By ~ 12

Author(s):

Xiaohui Zhou ◽

Jennifer M. Ritchie ◽

Hirotaka Hiyoshi ◽

Tetsuya Iida ◽

Brigid M. Davis ◽

...

Keyword(s):

Host Cell ◽

Vibrio Parahaemolyticus ◽

Diarrheal Disease ◽

Host Cells ◽

Cell Cytoplasm ◽

Host Cell Membrane ◽

Membrane Pore ◽

Content Type ◽

Bacterial Proteins ◽

Host Cell Cytoplasm

ABSTRACTThe pathogenesis of the diarrheal disease caused byVibrio parahaemolyticus, a leading cause of seafood-associated enteritis worldwide, is dependent upon a type III secretion system, T3SS2. This apparatus enables the pathogen to inject bacterial proteins (effectors) into the cytosol of host cells and thereby modulate host processes. T3SS effector proteins transit into the host cell via a membrane pore (translocon) typically formed by 3 bacterial proteins. We have identified the third translocon protein for T3SS2: VopW, which was previously classified as an effector protein for a homologous T3SS inV. cholerae. VopW is a hydrophilic translocon protein; like other such proteins, it is not inserted into the host cell membrane but is required for insertion of the two hydrophobic translocators, VopB2 and VopD2, that constitute the membrane channel. VopW is not required for secretion of T3SS2 effectors into the bacterial culture medium; however, it is essential for transfer of these proteins into the host cell cytoplasm. Consequently, deletion ofvopWabrogates the virulence ofV. parahaemolyticusin several animal models of diarrheal disease. Unlike previously described hydrophilic translocators, VopW is itself translocated into the host cell cytoplasm, raising the possibility that it functions as both a translocator and an effector.

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Expression of the EspB Protein of EnteropathogenicEscherichia coli within HeLa Cells Affects Stress Fibers and Cellular Morphology

Infection and Immunity ◽

10.1128/iai.67.1.120-125.1999 ◽

1999 ◽

Vol 67 (1) ◽

pp. 120-125 ◽

Cited By ~ 55

Author(s):

Kathleen A. Taylor ◽

Paul W. Luther ◽

Michael S. Donnenberg

Keyword(s):

Host Cell ◽

Hela Cells ◽

Type Iii Secretion ◽

Cell Clone ◽

Host Cells ◽

Stress Fibers ◽

Cell Cytoplasm ◽

Eukaryotic Promoter ◽

Host Cell Cytoplasm ◽

Attached Bacteria

ABSTRACT The EspB protein of enteropathogenic Escherichia coli(EPEC) is essential for the signaling events that lead to the accumulation of actin beneath intimately attached bacteria, a process that is known as the attaching and effacing effect. EspB is targeted to the host cell cytoplasm by a type III secretion apparatus. To determine the effect of intracellular EspB on the host cell cytoskeleton, we transfected HeLa cells with a plasmid containing the espBgene under the control of an inducible eukaryotic promoter. A HeLa cell clone that expressed espB mRNA and EspB protein after induction was selected for further study. The expression of EspB in these cells caused a dramatic change in cell morphology and a marked reduction in actin stress fibers. Cells expressing EspB were significantly impaired in their ability to support invasion by EPEC andSalmonella typhimurium. However, the expression of EspB within host cells could not compensate for the lack of EspB expression by an espB mutant strain of EPEC to restore attaching and effacing activity. These studies suggest that EspB is a cytoskeletal toxin that is translocated to the host cell cytoplasm, where it causes a redistribution of actin.

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A Chlamydia trachomatis OmcB C-Terminal Fragment Is Released into the Host Cell Cytoplasm and Is Immunogenic in Humans

Infection and Immunity ◽

10.1128/iai.00003-11 ◽

2011 ◽

Vol 79 (6) ◽

pp. 2193-2203 ◽

Cited By ~ 10

Author(s):

Manli Qi ◽

Siqi Gong ◽

Lei Lei ◽

Quanzhong Liu ◽

Guangming Zhong

Keyword(s):

Chlamydia Trachomatis ◽

Host Cell ◽

Host Cells ◽

Content Type ◽

Membrane Complex ◽

Host Cell Cytoplasm ◽

Complex Protein ◽

Infected Cells ◽

Cell Cytosol ◽

Host Cell Cytosol

ABSTRACTTheChlamydia trachomatisouter membrane complex protein B (OmcB) is an antigen with diagnostic and vaccine relevance. To further characterize OmcB, we generated antibodies against OmcB C-terminal (OmcBc) and N-terminal (OmcBn) fragments. Surprisingly, the anti-OmcBc antibody detected dominant signals in the host cell cytosol, while the anti-OmcBn antibody exclusively labeled intrainclusion signals inC. trachomatis-infected cells permeabilized with saponin. Western blot analyses revealed that OmcB was partially processed into OmcBc and OmcBn fragments. The processed OmcBc was released into host cell cytosol, while the OmcBn and remaining full-length OmcB were retained within the chlamydial inclusions. The organism-associated OmcB epitopes became detectable only after theC. trachomatis-infected cells were permeabilized with strong detergents such as SDS. However, the harsh permeabilization conditions also led to the leakage of the already secreted OmcBc and chlamydia-secreted protease (CPAF) out of the host cells. The OmcBc processing and release occurred in all biovars ofC. trachomatis. Moreover, the released OmcBc but not the retained OmcBn was highly immunogenic inC. trachomatis-infected women, which is consistent with the concept that exposure of chlamydial proteins to host cell cytosol is accompanied by increased immunogenicity. These observations have provided important information for further exploring/optimizing OmcB as a target for the development of diagnosis methods and vaccines.

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Brucella abortus Rough Mutants Induce Macrophage Oncosis That Requires Bacterial Protein Synthesis and Direct Interaction with the Macrophage

Infection and Immunity ◽

10.1128/iai.74.5.2667-2675.2006 ◽

2006 ◽

Vol 74 (5) ◽

pp. 2667-2675 ◽

Cited By ~ 34

Author(s):

Jianwu Pei ◽

Joshua E. Turse ◽

Qingmin Wu ◽

Thomas A. Ficht

Keyword(s):

Cell Death ◽

Protein Synthesis ◽

Brucella Abortus ◽

Direct Interaction ◽

Host Cells ◽

Target Cells ◽

Bacterial Protein ◽

Necrosis Factor Alpha ◽

Bacterial Protein Synthesis ◽

Rough Mutant

ABSTRACT Previous studies suggest that smooth Brucella organisms inhibit macrophage apoptosis. In contrast, necrotic cell death of macrophages infected with rough Brucella organisms in vitro has been reported, which may in part explain the failure of some rough organisms to thrive. To characterize these potential macrophage killing mechanisms, J774.A1 murine macrophages were infected with Brucella abortus S2308-derived rough mutant CA180. Electron microscopic analysis and polyethylene glycol protection assays revealed that the cells were killed as a result of necrosis and oncosis. This killing was shown to be unaffected by treatment with carbenicillin, an inhibitor of bacterial cell wall biosynthesis and, indirectly, replication. In contrast, chloramphenicol treatment of macrophages infected at multiplicities of infection exceeding 10,000 prevented cell death, despite internalization of large numbers of bacteria. Similarly, heat-killed and gentamicin-killed CA180 did not induce cytopathic effects in the macrophage. These results suggested that killing of infected host cells requires active bacterial protein synthesis. Cytochalasin D treatment revealed that internalization of the bacteria was necessary to initiate killing. Transwell experiments demonstrated that cell death is not mediated by a diffusible product, including tumor necrosis factor alpha and nitric oxide, but does require direct contact between host and pathogen. Furthermore, macrophages preinfected with B. abortus S2308 or pretreated with B. abortus O polysaccharide did not prevent rough CA180-induced cell death. In conclusion, Brucella rough mutant infection induces necrotic and oncotic macrophage cell death that requires bacterial protein synthesis and direct interaction of bacteria with the target cells.

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