OBSERVATIONS ON THE MODE OF RELEASE OF HERPES VIRUS FROM INFECTED HELA CELLS

The development of a well adapted strain of herpes virus has been studied in HeLa cells using thin sectioning techniques for electron microscopy. Particular attention was directed to events in the cytoplasm and certain new features were observed. Profuse immature particles with a nucleoid and single limiting membrane were present in the nuclei of infected cells, often in crystalline array; morphologically indistinguishable immature particles were also found very frequently in the cytoplasm. Cells with such particles were intact and well preserved, and contained smooth vacuoles apparently derived from the Golgi component of the endoplasmic reticulum. The cytoplasmic particles escaped from the cells by bulging out as buds through the cell membrane or through that of the cytoplasmic vacuoles until they were attached only by a pedicle and then became free. During this process the particles were gradually enclosed by the membrane through which they passed and carried a coat of it with them as they matured. After permanganate fixation the triple-layered structure of the cell membrane and vacuolar membranes was evident and was identical with that of the outer coat of the mature virus. These findings are discussed both in relation to different types of virus structure and to function in the endoplasmic reticulum and cell membrane.

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THE ENTRY AND DISTRIBUTION OF HERPES VIRUS AND COLLOIDAL GOLD IN HELA CELLS AFTER CONTACT IN SUSPENSION

Journal of Experimental Medicine ◽

10.1084/jem.119.2.291 ◽

1964 ◽

Vol 119 (2) ◽

pp. 291-302 ◽

Cited By ~ 24

Author(s):

M. A. Epstein ◽

K. Hummeler ◽

A. Berkaloff

Keyword(s):

Hela Cells ◽

Colloidal Gold ◽

Herpes Virus ◽

Amorphous Layer ◽

Cytoplasmic Matrix ◽

Virus Particles ◽

Thin Sectioning ◽

Outer Aspect ◽

Herpès Virus ◽

Mature Virus

The way in which herpes virus of a well adapted strain penetrates susceptible HeLa cells has been investigated using thin sectioning techniques for electron microscopy. Mature virus particles and cells were mixed together in suspension cultures for 15, 30, 60, or 120 minutes so that the stages in virus uptake could be followed in sequence. The ingestion of particles of colloidal gold by HeLa cells under similar conditions was studied for comparison in parallel experiments. After 15 minutes' contact, the mature virus was found adsorbed on the surface of the cells but separated from them by a narrow gap in which phosphotungstic acid staining was sometimes able to reveal an extraneous coat which appeared as an amorphous layer on the outer aspect of the plasma membrane. When mixing continued for longer the particles were present in deep invaginations or actual cytoplasmic vacuoles, with their outer layers in various stages of stripping and digestion. The stripped, naked, central portion of the virus was occasionally found in these vacuoles but was more commonly free in the cytoplasmic matrix; the mode of transition between these sites could not be determined. Where contact continued for 2 hours these phenomena were much less frequently observed. The larger particles of colloidal gold were ingested in the same way as the virus, but smaller ones were taken up in micropinocytosis vesicles. The gold passed through membrane-bounded cytoplasmic spaces to accumulate in vacuoles from which, in contrast to herpes particles, it did not escape. These findings are discussed, and considered with particular reference to their bearing on the initiation of infection, the uptake and disposal of particles by cells, and the influence on the latter of virus morphology.

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Brucella abortus Transits through the Autophagic Pathway and Replicates in the Endoplasmic Reticulum of Nonprofessional Phagocytes

Infection and Immunity ◽

10.1128/iai.66.12.5711-5724.1998 ◽

1998 ◽

Vol 66 (12) ◽

pp. 5711-5724 ◽

Cited By ~ 284

Author(s):

Javier Pizarro-Cerdá ◽

Stéphane Méresse ◽

Robert G. Parton ◽

Gisou van der Goot ◽

Alberto Sola-Landa ◽

...

Keyword(s):

Endoplasmic Reticulum ◽

Hela Cells ◽

Brucella Abortus ◽

Cathepsin D ◽

Bacterial Strains ◽

Infected Cells ◽

Bacterial Replication ◽

Protein Disulfide ◽

Intracellular Pathway ◽

Replication Compartment

ABSTRACT Brucella abortus is an intracellular pathogen that replicates within a membrane-bounded compartment. In this study, we have examined the intracellular pathway of the virulent B. abortus strain 2308 (S2308) and the attenuated strain 19 (S19) in HeLa cells. At 10 min after inoculation, both bacterial strains are transiently detected in phagosomes characterized by the presence of early endosomal markers such as the early endosomal antigen 1. At ∼1 h postinoculation, bacteria are located within a compartment positive for the lysosome-associated membrane proteins (LAMPs) and the endoplasmic reticulum (ER) marker sec61β but negative for the mannose 6-phosphate receptors and cathepsin D. Interestingly, this compartment is also positive for the autophagosomal marker monodansylcadaverin, suggesting that S2308 and S19 are located in autophagic vacuoles. At 24 h after inoculation, attenuated S19 is degraded in lysosomes, while virulent S2308 multiplies within a LAMP- and cathepsin D-negative but sec61β- and protein disulfide isomerase-positive compartment. Furthermore, treatment of infected cells with the pore-forming toxin aerolysin from Aeromonas hydrophila causes vacuolation of the bacterial replication compartment. These results are compatible with the hypothesis that pathogenic B. abortus exploits the autophagic machinery of HeLa cells to establish an intracellular niche favorable for its replication within the ER.

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Compartment-specific immunolocalization of conserved epitopes of the glycoprotein gB of herpes simplex virus type 1 and bovine herpes virus type 2 in infected cells

Archives of Virology ◽

10.1007/bf01313738 ◽

1989 ◽

Vol 108 (1-2) ◽

pp. 1-17 ◽

Cited By ~ 6

Author(s):

Sylvia M. Pietschmann ◽

H. R. Gelderblom ◽

G. Pauli

Keyword(s):

Herpes Simplex Virus ◽

Herpes Simplex ◽

Virus Type ◽

Herpes Simplex Virus Type ◽

Herpes Virus ◽

Infected Cells ◽

Herpès Virus ◽

Simplex Virus

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Possible associationo virus-induced deoxyribonucleic acid polymerase and deoxyribonucleic acid exonuclease in herpes-virus-infected cells

Biochemical Journal ◽

10.1042/bj1140039p ◽

1969 ◽

Vol 114 (2) ◽

pp. 39P-39P

Author(s):

R D Paton ◽

J M Morrison

Keyword(s):

Herpes Virus ◽

Deoxyribonucleic Acid ◽

Infected Cells ◽

Herpès Virus

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Role of High-Mobility Group Box 1 Protein and Poly(ADP-Ribose) Polymerase 1 Degradation in Chlamydia trachomatis-Induced Cytopathicity

Infection and Immunity ◽

10.1128/iai.01404-09 ◽

2010 ◽

Vol 78 (7) ◽

pp. 3288-3297 ◽

Cited By ~ 19

Author(s):

Hangxing Yu ◽

Katja Schwarzer ◽

Martin Förster ◽

Olaf Kniemeyer ◽

Vera Forsbach-Birk ◽

...

Keyword(s):

Chlamydia Trachomatis ◽

Cell Membrane ◽

Hela Cells ◽

High Mobility ◽

Host Cells ◽

High Mobility Group ◽

Cell Membrane Permeability ◽

Dependent Manner ◽

Infected Cells ◽

Parp 1

ABSTRACT As intracellular bacteria, chlamydiae block the apoptotic pathways of their host cells. However, the infection of epithelial cells causes the loss of cell membrane integrity and can result in nonapoptotic death. Normally, cells undergoing necrosis release high-mobility group box 1 protein (HMGB1) that acts as an important proinflammatory mediator. Here, we show that in Chlamydia trachomatis-infected HeLa cells HMGB1 is not translocated from the nucleus to the cytosol and not released from injured cells in increased amounts. At 48 h after infection, degradation of HMGB1 was observed. In infected cells, poly(ADP-ribose) polymerase 1 (PARP-1), a DNA repair enzyme that also regulates HMGB1 translocation, was found to be cleaved into fragments that correspond to a necrosislike pattern of PARP-1 degradation. Cell-free cleavage assays and immunoprecipitation using purified proteolytic fractions from infected cells demonstrated that the chlamydial-protease-like activity factor (CPAF) is responsible for the cleavage of both HMGB1 and PARP-1. Proteolytic cleavage of PARP-1 was accompanied by a significant decrease in the enzymatic activity in a time-dependent manner. The loss of PARP-1 function obviously affects the viability of Chlamydia-infected cells because silencing of PARP-1 in uninfected HeLa cells with specific small interfering RNA results in increased cell membrane permeability. Our findings suggest that the Chlamydia-specific protease CPAF interferes with necrotic cell death pathways. By the degradation of HMGB1 and PARP-1, the pathogen may have evolved a strategy to reduce the inflammatory response to membrane-damaged cells in vivo.

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