scholarly journals Interaction of the movement and coat proteins of Maize streak virus: implications for the transport of viral DNA

2001 ◽  
Vol 82 (1) ◽  
pp. 35-44 ◽  
Author(s):  
Huanting Liu ◽  
Margaret I. Boulton ◽  
Karl J. Oparka ◽  
Jeffrey W. Davies
Keyword(s):  
Cell Movement ◽  
Systemic Infection ◽  
Maize Streak Virus ◽  
Streak Virus ◽  
Cell Periphery ◽  
Coat Proteins ◽  
Viral Dna ◽  
Dna Complex

We have shown previously that the movement protein (MP) and coat protein (CP) of Maize streak virus (MSV) are both required for systemic infection. Towards understanding the roles of these two proteins in virus movement, each was expressed in E. coli and interactions of the MP with viral DNA or CP were investigated using south-western, gel overlay and immunoprecipitation assays. Unlike the CP, the MP did not bind to viral DNA but it interacted with the CP in vitro and an MP–CP complex was detected in extracts from MSV-infected maize, indicating the potential for an interaction in vivo. Microinjection showed that the MP could prevent the nuclear transport of an MSV CP–DNA complex in maize and tobacco cells. These results are consistent with a model in which the MP diverts a CP–DNA complex from the nucleus (where viral DNA replication takes place) to the cell periphery, and in co-operation with the CP, mediates the cell-to-cell movement of the viral DNA. In this respect, the MSV MP and CP have functional analogy with the BC1 and BV1 proteins, respectively, of the Begomovirus genus of the Geminiviridae.

scholarly journals Biological Activity of Two Tombusvirus Proteins Translated from Nested Genes Is Influenced by Dosage Control via Context-Dependent Leaky Scanning

1999 ◽  
Vol 12 (8) ◽  
pp. 670-679 ◽  
Author(s):  
Herman B. Scholthof ◽  
Bénédicte Desvoyes ◽  
Joan Kuecker ◽  
Elizabeth Whitehead
Keyword(s):  
Biological Activities ◽  
Cell Movement ◽  
Systemic Infection ◽  
Protein Accumulation ◽  
Leaky Scanning ◽  
Protein Ratio ◽  
Context Dependent ◽  
Translational Start

Tomato bushy stunt virus (TBSV) encodes a small gene (p19) nested within the cell-to-cell movement gene (p22), and their translation yields two proteins with separate activities for virus spread and symptom induction. The objective of this study was to determine the biological relevance associated with the translational mechanism responsible for expression of the nested p22 and p19 genes. Introduction of site-specific mutations to optimize the translational start site context of p22 caused a substantial shift in the ratio of the two proteins, mainly because it dramatically reduced the otherwise abundant levels of p19 protein accumulation in vitro and in vivo. Changes in the dosage or ratios of p22 and p19 proteins failed to noticeably affect virus replication or movement in Nicotiana spp. that support a systemic infection. In contrast, bio-assays with hypersensitive Nicotiana hosts illustrated that a substantially elevated p22/p19 protein ratio increased the size of p19 protein-mediated lesions whereas those induced by the p22 protein tended to be smaller. The reduced levels of p19 protein prevented the onset of a lethal apical necrosis in systemically infected Nicotiana benthamiana plants. Furthermore, the increased p22/p19 protein ratio impaired the ability of TBSV to systemically invade spinach plants. These results suggest that control of tombusvirus p22 and p19 protein ratios and dosage through context-dependent leaky scanning provides a co-translational mechanism to coordinate their biological activities.

scholarly journals Dissemination of Rat Cytomegalovirus through Infected Granulocytes and Monocytes In Vitro and In Vivo

2003 ◽  
Vol 77 (20) ◽  
pp. 11274-11278 ◽  
Author(s):  
B. W. A. van der Strate ◽  
J. L. Hillebrands ◽  
S. S. Lycklama à Nijeholt ◽  
L. Beljaars ◽  
C. A. Bruggeman ◽  
...  
Keyword(s):  
Intravenous Injection ◽  
Systemic Infection ◽  
Insight Into

ABSTRACT The role of leukocytes in the in vivo dissemination of cytomegalovirus was studied in this experiment. Rat cytomegalovirus (RCMV) could be transferred to rat granulocytes and monocytes by cocultivation with RCMV-infected fibroblasts in vitro. Intravenous injection of purified infected granulocytes or monocytes resulted in a systemic infection in rats, indicating that our model is a powerful tool to gain further insight into CMV dissemination and the development of new antivirals.

scholarly journals Antimicrobial Photodynamic Inactivation Inhibits Candida albicans Virulence Factors and ReducesIn VivoPathogenicity

2012 ◽  
Vol 57 (1) ◽  
pp. 445-451 ◽  
Author(s):  
Ilka Tiemy Kato ◽  
Renato Araujo Prates ◽  
Caetano Padial Sabino ◽  
Beth Burgwyn Fuchs ◽  
George P. Tegos ◽  
...  
Keyword(s):  
Candida Albicans ◽  
Virulence Factors ◽  
Sodium Dodecyl ◽  
Systemic Infection ◽  
Tube Formation ◽  
Photodynamic Inactivation ◽  
Content Type ◽  
Daughter Cells

ABSTRACTThe objective of this study was to evaluate whetherCandida albicansexhibits altered pathogenicity characteristics following sublethal antimicrobial photodynamic inactivation (APDI) and if such alterations are maintained in the daughter cells.C. albicanswas exposed to sublethal APDI by using methylene blue (MB) as a photosensitizer (0.05 mM) combined with a GaAlAs diode laser (λ 660 nm, 75 mW/cm2, 9 to 27 J/cm2).In vitro, we evaluated APDI effects onC. albicansgrowth, germ tube formation, sensitivity to oxidative and osmotic stress, cell wall integrity, and fluconazole susceptibility.In vivo, we evaluatedC. albicanspathogenicity with a mouse model of systemic infection. Animal survival was evaluated daily. Sublethal MB-mediated APDI reduced the growth rate and the ability ofC. albicansto form germ tubes compared to untreated cells (P< 0.05). Survival of mice systemically infected withC. albicanspretreated with APDI was significantly increased compared to mice infected with untreated yeast (P< 0.05). APDI increasedC. albicanssensitivity to sodium dodecyl sulfate, caffeine, and hydrogen peroxide. The MIC for fluconazole forC. albicanswas also reduced following sublethal MB-mediated APDI. However, none of those pathogenic parameters was altered in daughter cells ofC. albicanssubmitted to APDI. These data suggest that APDI may inhibit virulence factors and reducein vivopathogenicity ofC. albicans. The absence of alterations in daughter cells indicates that APDI effects are transitory. The MIC reduction for fluconazole following APDI suggests that this antifungal could be combined with APDI to treatC. albicansinfections.

scholarly journals SpoVID Guides SafA to the Spore Coat inBacillus subtilis

2001 ◽  
Vol 183 (10) ◽  
pp. 3041-3049 ◽  
Author(s):  
Amanda J. Ozin ◽  
Craig S. Samford ◽  
Adriano O. Henriques ◽  
Charles P. Moran
Keyword(s):  
Direct Interaction ◽  
Spore Coat ◽  
Coat Proteins ◽  
Yeast Two Hybrid System ◽  
Spore Coat Proteins ◽  
Basement Layer ◽  
Two Hybrid ◽  
Spore Coat Protein

ABSTRACT Bacteria assemble complex structures by targeting proteins to specific subcellular locations. The protein coat that encasesBacillus subtilis spores is an example of a structure that requires coordinated targeting and assembly of more than 24 polypeptides. The earliest stages of coat assembly require the action of three morphogenetic proteins: SpoIVA, CotE, and SpoVID. In the first steps, a basement layer of SpoIVA forms around the surface of the forespore, guiding the subsequent positioning of a ring of CotE protein about 75 nm from the forespore surface. SpoVID localizes near the forespore membrane where it functions to maintain the integrity of the CotE ring and to anchor the nascent coat to the underlying spore structures. However, it is not known which spore coat proteins interact directly with SpoVID. In this study we examined the interaction between SpoVID and another spore coat protein, SafA, in vivo using the yeast two-hybrid system and in vitro. We found evidence that SpoVID and SafA directly interact and that SafA interacts with itself. Immunofluorescence microscopy showed that SafA localized around the forespore early during coat assembly and that this localization of SafA was dependent on SpoVID. Moreover, targeting of SafA to the forespore was also dependent on SpoIVA, as was targeting of SpoVID to the forespore. We suggest that the localization of SafA to the spore coat requires direct interaction with SpoVID.

scholarly journals Macrophage oxygen-dependent antimicrobial activity. III. Enhanced oxidative metabolism as an expression of macrophage activation.

1980 ◽  
Vol 152 (6) ◽  
pp. 1596-1609 ◽  
Author(s):  
H W Murray ◽  
Z A Cohn
Keyword(s):  
Antimicrobial Activity ◽  
Oxidative Metabolism ◽  
Macrophage Activation ◽  
Systemic Infection ◽  
Peritoneal Macrophages ◽  
H2o2 Production ◽  
Oxygen Intermediates ◽  
Cultivation In Vitro

The capacity of 15 separate populations of mouse peritoneal macrophages to generate and release H2O2 (an index of oxidative metabolism) was compared with their ability to inhibit the intracellular replication of virulent Toxoplasma gondii. Resident macrophages and those elicited by inflammatory agents readily supported toxoplasma multiplication and released 4-20X less H2O2 than macrophages activated in vivo by systemic infection with Bacille Calmette-Guérin or T. gondii, or by immunization with Corynebacterium parvum. Immunologically activated cells consistently displayed both enhanced H2O2 production and antitoxoplasma activity. Exposure to lymphokines generated from cultures of spleen cells from T. gondii immune mice and toxoplasma antigen preserved both the antitoxoplasma activity and the heightened H2O2 release of toxoplasma immune and immune-boosted macrophages, which otherwise were lost after 48-72 h of cultivation. In vitro activation of resident and chemically-elicited cells by 72 h of exposure to mitogen- and antigen-prepared lymphokines, conditions that induce trypanocidal (5) and leishmanicidal activity (14), stimulated O2- and H2O2 release, and enhanced nitroblue tetrazolium reduction in response to toxoplasma ingestion. Such treatment, however, failed to confer any antitoxoplasma activity, indicating that intracellular pathogens may vary in their susceptibility to macrophage microbicidal mechanisms, including specific oxygen intermediates. In contrast, cocultivating normal macrophages with lymphokine plus heart infusion broth for 18H rendered these cells toxoplasmastatic. This in vitro-acquired activity was inhibited by scavengers of O2-, H2O2, OH., and 1O2, demonstrating a role for oxidative metabolites in lymphokine-induced enhancement of macrophage antimicrobial activity. These findings indicate that augmented oxidative metabolism is an consistent marker of macrophage activation, and that oxygen intermediates participate in the resistance of both in vivo- and vitro-activated macrophages toward the intracellular parasite, T. gondii.

The mechanism of mouse egg-cylinder morphogenesis in vitro

Development ◽  
1981 ◽  
Vol 61 (1) ◽  
pp. 277-287
Author(s):  
A. J. Copp
Keyword(s):  
Giant Cell ◽  
Cell Formation ◽  
Cell Movement ◽  
Giant Cells ◽  
Cell Number ◽  
Dependent Change ◽  
Morphogenesis In Vitro ◽  
Trophoblast Giant Cells

The number of trophoblast giant cells in outgrowths of mouse blastocysts was determined before, during and after egg-cylinder formation in vitro. Giant-cell numbers rose initially but reached a plateau 12 h before the egg cylinder appeared. A secondary increase began 24 h after egg-cylinder formation. Blastocysts whose mural trophectoderm cells were removed before or shortly after attachment in vitro formed egg cylinders at the same time as intact blastocysts but their trophoblast outgrowths contained fewer giant cells at this time. The results support the idea that egg-cylinder formation in vitro is accompanied by a redirection of the polar to mural trophectoderm cell movement which characterizes blastocysts before implantation. The resumption of giant-cell number increase in trophoblast outgrowths after egg-cylinder formation may correspond to secondary giant-cell formation in vivo. It is suggested that a time-dependent change in the strength of trophoblast cell adhesion to the substratum occurs after blastocyst attachment in vitro which restricts the further entry of polar cells into the outgrowth and therefore results in egg-cylinder formation.

scholarly journals Smurf1 regulates tumor cell plasticity and motility through degradation of RhoA leading to localized inhibition of contractility

2006 ◽  
Vol 176 (1) ◽  
pp. 35-42 ◽  
Author(s):  
Erik Sahai ◽  
Raquel Garcia-Medina ◽  
Jacques Pouysségur ◽  
Emmanuel Vial
Keyword(s):  
Cell Migration ◽  
Tumor Cell ◽  
Rho Gtpases ◽  
Rho Kinase ◽  
Cell Movement ◽  
Leading Edge ◽  
Cell Periphery ◽  
Tumor Cell Migration ◽  
Tumor Cell Motility

Rho GTPases participate in various cellular processes, including normal and tumor cell migration. It has been reported that RhoA is targeted for degradation at the leading edge of migrating cells by the E3 ubiquitin ligase Smurf1, and that this is required for the formation of protrusions. We report that Smurf1-dependent RhoA degradation in tumor cells results in the down-regulation of Rho kinase (ROCK) activity and myosin light chain 2 (MLC2) phosphorylation at the cell periphery. The localized inhibition of contractile forces is necessary for the formation of lamellipodia and for tumor cell motility in 2D tissue culture assays. In 3D invasion assays, and in in vivo tumor cell migration, the inhibition of Smurf1 induces a mesenchymal–amoeboid–like transition that is associated with a more invasive phenotype. Our results suggest that Smurf1 is a pivotal regulator of tumor cell movement through its regulation of RhoA signaling.

Lactoferrin protects neonatal rats from gut-related systemic infection

2001 ◽  
Vol 281 (5) ◽  
pp. G1140-G1150 ◽  
Author(s):  
Lynn Edde ◽  
Ronaldo B. Hipolito ◽  
Freda F. Y. Hwang ◽  
Denis R. Headon ◽  
Robert A. Shalwitz ◽  
...  
Keyword(s):  
Systemic Infection ◽  
In Vitro Studies ◽  
Peptide Antibiotics ◽  
Neonatal Rats ◽  
Vitro Assay ◽  
E Coli ◽  
Severity Scores ◽  
Factor Α

Lactoferrin is a milk protein that reportedly protects infants from gut-related, systemic infection. Proof for this concept is limited and was addressed during in vivo and in vitro studies. Neonatal rats pretreated orally with recombinant human lactoferrin (rh-LF) had less bacteremia and lower disease severity scores ( P < 0.001) after intestinal infection with Escherichia coli. Control animals had 1,000-fold more colony-forming units of E. coli per milliliter of blood than treated animals ( P < 0.001). Liver cultures from control animals had a twofold increase in bacterial counts compared with cultures from rh-LF-treated pups ( P < 0.02). Oral therapy with rh-LF + FeSO4did not alter the protective effect. In vitro studies confirmed that rh-LF interacted with the infecting bacterium and rat macrophages. An in vitro assay showed that rh-LF did not kill E. coli, but a combination of rh-LF + lysozyme was microbicidal. In vitro studies showed that rat macrophages released escalating amounts of nitric oxide and tumor necrosis factor-α when stimulated with increasing concentrations of rh-LF. The in vitro studies suggest that rh-LF may act with other “natural peptide antibiotics” or may prime macrophages to kill E. coli in vivo.

scholarly journals An In Vitro Model for Assessing Corneal Keratocyte Spreading and Migration on Aligned Fibrillar Collagen

2018 ◽  
Vol 9 (4) ◽  
pp. 54 ◽  
Author(s):  
Pouriska Kivanany ◽  
Kyle Grose ◽  
Nihan Yonet-Tanyeri ◽  
Sujal Manohar ◽  
Yukta Sunkara ◽  
...  
Keyword(s):  
Growth Factor ◽  
Transforming Growth Factor Beta ◽  
Transforming Growth Factor ◽  
Cell Movement ◽  
Time Lapse ◽  
Collagen Fibrils ◽  
Fibrillar Collagen ◽  
Freeze Injury

Background: Corneal stromal cells (keratocytes) are responsible for developing and maintaining normal corneal structure and transparency, and for repairing the tissue after injury. Corneal keratocytes reside between highly aligned collagen lamellae in vivo. In addition to growth factors and other soluble biochemical factors, feedback from the extracellular matrix (ECM) itself has been shown to modulate corneal keratocyte behavior. Methods: In this study, we fabricate aligned collagen substrates using a microfluidics approach and assess their impact on corneal keratocyte morphology, cytoskeletal organization, and patterning after stimulation with platelet derived growth factor (PDGF) or transforming growth factor beta 1 (TGFβ). We also use time-lapse imaging to visualize the dynamic interactions between cells and fibrillar collagen during wound repopulation following an in vitro freeze injury. Results: Significant co-alignment between keratocytes and aligned collagen fibrils was detected, and the degree of cell/ECM co-alignment further increased in the presence of PDGF or TGFβ. Freeze injury produced an area of cell death without disrupting the collagen. High magnification, time-lapse differential interference contrast (DIC) imaging allowed cell movement and subcellular interactions with the underlying collagen fibrils to be directly visualized. Conclusions: With continued development, this experimental model could be an important tool for accessing how the integration of multiple biophysical and biochemical signals regulate corneal keratocyte differentiation.

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