bluuebunny/crossref_metadata_2025_split huggingface dataset:
The Purified and Recombinant
Legionella pneumophila
Chaperonin Alters Mitochondrial Trafficking and Microfilament Organization
Authors: Chong Audrey , Lima Celia A. , Allan David S. , Nasrallah Gheyath K. , Garduño Rafael A. 
Published: August 2009

ABSTRACT

A portion of the total cellular pool of the
Legionella pneumophila
chaperonin, HtpB, is found on the bacterial cell surface, where it can mediate invasion of nonphagocytic cells. HtpB continues to be abundantly produced and released by internalized
L. pneumophila
and may thus have postinvasion functions. We used here two functional models (protein-coated beads and expression of recombinant proteins in CHO cells) to investigate the competence of HtpB in mimicking early intracellular trafficking events of
L. pneumophila
, including the recruitment of mitochondria, cytoskeletal alterations, the inhibition of phagosome-lysosome fusion, and association with the endoplasmic reticulum. Microscopy and flow cytometry studies indicated that HtpB-coated beads recruited mitochondria in CHO cells and U937-derived macrophages and induced transient changes in the organization of actin microfilaments in CHO cells. Ectopic expression of HtpB in the cytoplasm of transfected CHO cells also led to modifications in actin microfilaments similar to those produced by HtpB-coated beads but did not change the distribution of mitochondria. Association of phagosomes containing HtpB-coated beads with the endoplasmic reticulum was not consistently detected by either fluorescence or electron microscopy studies, and only a modest delay in the fusion of TrOv-labeled lysosomes with phagosomes containing HtpB-coated beads was observed. HtpB is the first
Legionella
protein and the first chaperonin shown to, by means of our functional models, induce mitochondrial recruitment and microfilament rearrangements, two postinternalization events that typify the early trafficking of virulent
L. pneumophila
.
Cryptosporidium parvumIsolate-Dependent Postinfectious Jejunal Hypersensitivity and Mast Cell Accumulation in an Immunocompetent Rat Model
Authors: Khaldi Samira , Gargala Gilles , Le Goff Laetitia , Parey Simon , Francois Arnaud , Fioramonti Jean , Ballet Jean-Jacques , Dupont Jean-Paul , Ducrotté Philippe , Favennec Loïc 
Published: August 2009

ABSTRACTCryptosporidiumspp. are a cause of self-limited diarrhea in immunocompetent hosts. In immunocompetent rats,Cryptosporidium parvuminfection induced digestive hypersensitivity, a key pathophysiological factor in functional digestive disorders such as irritable bowel syndrome (IBS). In such a rat model, we sought to document whether jejunal hypersensitivity depends onC. parvumisolate and is associated with a mast cell accumulation. Five-day-old rats were orally administered 105oocysts of either Nouzilly (NoI) or Iowa (IoI)C. parvumisolate. NoI-infected rats exhibited the lowest food intake on days 7 and 14 postinfection (p.i.). On day 7 p.i., small intestine villus atrophy, crypt hyperplasia, and inflammatory cell infiltration were prominent in NoI-infected rats, with higher numbers ofCryptosporidiumforms than in IoI-infected rats. Compared to uninfected control rats, jejunal intraepithelial lymphocytes (IELs) were increased only in NoI-infected rats on day 14 p.i. On day 50 p.i., jejunal hypersensitivity to distension was found only in NoI-infected rats; this hypersensitivity is associated with activated mast cell accumulation. The number of mast cells in the jejunal lamina propria was increased from day 36 p.i. in NoI-infected rats and only at day 120 p.i. in IoI-infected rats. Our data suggest that both the severity of infection (weight loss, reduced food intake, villus atrophy, and IEL accumulation) and the onset of a jejunal hypersensitivity after infection in association with an activated mast cell accumulation are isolate dependent and related to NoI infection. This cryptosporidiosis rat model is a relevant model for the study of underlying mechanisms of postinfectious IBS-like symptoms.
Roles for NK Cells and an NK Cell-Independent Source of Intestinal Gamma Interferon for Innate Immunity toCryptosporidium parvumInfection
Authors: Barakat Farah M. , McDonald Vincent , Di Santo James P. , Korbel Daniel S. 
Published: August 2009

ABSTRACTA gamma interferon (IFN-γ)-dependent innate immune response operates against the intestinal parasiteCryptosporidium parvumin T- and B-cell-deficient SCID mice. Although NK cells are a major source of IFN-γ in innate immunity, their protective role againstC. parvumhas been unclear. The role of NK cells in innate immunity was investigated using Rag2−/−mice, which lack T and B cells, and Rag2−/−γc−/−mice, which, in addition, lack NK cells. Adult mice of both knockout lines developed progressive chronic infections; however, on most days the level of oocyst excretion was higher in Rag2−/−γc−/−mice and these animals developed morbidity and died, whereas within the same period the Rag2−/−mice appeared healthy. Neonatal mice of both mouse lines survived a rapid onset of infection that reached a higher intensity in Rag2−/−γc−/−mice. Significantly, similar levels of intestinal IFN-γ mRNA were expressed in Rag2−/−and Rag2−/−γc−/−mice. Also, infections in each mouse line were exacerbated by treatment with anti-IFN-γ neutralizing antibodies. These results support a protective role for NK cells and IFN-γ in innate immunity againstC. parvum. In addition, the study implies that an intestinal cell type other than NK cells may be an important source of IFN-γ during infection and that NK cells may have an IFN-γ-independent protective role.
Microbial Quorum-Sensing Molecules Induce Acrosome Loss and Cell Death in Human Spermatozoa
Authors: Rennemeier Claudia , Frambach Torsten , Hennicke Florian , Dietl Johannes , Staib Peter 
Published: August 2009

ABSTRACT

Infertility in men and women is frequently associated with genital contamination by various commensal or uropathogenic microbes. Since many microorganisms are known to release quorum-sensing signals in substantial amounts, we raised the question whether such molecules can directly affect human spermatozoa. Here we show that farnesol and 3-oxododecanoyl-
l
-homoserine lactone, employed by the opportunistic pathogenic yeast
Candida albicans
and the gram-negative bacterium
Pseudomonas aeruginosa
, respectively, induce multiple damage in spermatozoa. A reduction in the motility of spermatozoa coincided in a dose-dependent manner with apoptosis and necrosis at concentrations which were nondeleterious for dendritic cell-like immune cells. Moreover, sublethal doses of both signaling molecules induced premature loss of the acrosome, a cap-like structure of the sperm head which is essential for fertilization. Addressing their mechanism of action, we found that the bacterial molecule, but not the fungal molecule, actively induced the acrosome reaction via a calcium-dependent mechanism. This work uncovers a new facet in the interaction of microorganisms with human gametes and suggests a putative link between microbial communication systems and host infertility.
Ehrlichia chaffeensis
Infections in
Drosophila melanogaster
Authors: Luce-Fedrow Alison , Von Ohlen Tonia , Chapes Stephen K. 
Published: August 2009

ABSTRACT

Ehrlichia chaffeensis
is an obligate, intracellular bacterium, transmitted by the tick
Amblyomma americanum
, and is the causative agent of human monocytic ehrlichiosis infections. We previously demonstrated that
E. chaffeensis
is capable of growing in
Drosophila
S2 cells. Therefore, we tested the hypothesis that
E. chaffeensis
can infect adult
Drosophila melanogaster
. Adult
Drosophila
organisms were experimentally challenged with intra-abdominal injections of bacteria.
Ehrlichia
-infected flies showed decreased survival compared to wild-type flies, and bacteria isolated from flies could reinfect mammalian macrophages.
Ehrlichia
infections activated both the cellular and humoral immune responses in the fly. Hemocytes phagocytosed bacteria after injection, and antimicrobial peptide pathways were induced following infection. Increased pathogenicity in flies carrying mutations in genes in both the Toll and Imd pathways suggests that both immune defense pathways participate in host defense. Induction of
Drosophila
cellular and humoral responses and the in vivo replication of
E. chaffeensis
suggests that
D. melanogaster
is a suitable host for
E. chaffeensis
. In the future, it will be a useful tool to unlock some of the in vivo mysteries of this arthropod-borne bacterium.
The NS1 Protein of the 1918 Pandemic Influenza Virus Blocks Host Interferon and Lipid Metabolism Pathways
Authors: Billharz Rosalind , Zeng Hui , Proll Sean C. , Korth Marcus J. , Lederer Sharon , Albrecht Randy , Goodman Alan G. , Rosenzweig Elizabeth , Tumpey Terrence M. , García-Sastre Adolfo , Katze Michael G. 
Published: August 2009

ABSTRACT
The “Spanish influenza” of 1918 claimed an unprecedented number of lives, yet the determinants of virulence for this virus are still not fully understood. Here, we used functional genomics and an in vitro human lung epithelial cell infection model to define the global host transcriptional response to the eight-gene 1918 virus. To better understand the role of the 1918 virus NS1 gene, we also evaluated the host response to a reassortant 1918 virus containing the NS1 gene from A/Texas/36/91 (a seasonal isolate of human influenza virus), as well as the host response to a reassortant of A/Texas/36/91 containing the 1918 NS1 gene. Genomic analyses revealed that the 1918 virus blocked the transcription of multiple interferon-stimulated genes and also downregulated a network of genes associated with lipid metabolism. In contrast, the expression of genes encoding chemokines and cytokines, which serve to attract infiltrating immune cells, was upregulated. Viruses containing the NS1 gene from A/Texas/36/91 induced a significant increase in type I interferon signaling but did not repress lipid metabolism. The 1918 NS1 gene may therefore have contributed to the virulence of the 1918 pandemic virus by disrupting the innate immune response, inducing hypercytokinemia, and by blocking the transcription of certain lipid-based proinflammatory mediators that function as part of the host antiviral response.
Host Genetic Variation Affects Resistance to Infection with a Highly Pathogenic H5N1 Influenza A Virus in Mice
Authors: Boon Adrianus C. M. , deBeauchamp Jennifer , Hollmann Anna , Luke Jennifer , Kotb Malak , Rowe Sarah , Finkelstein David , Neale Geoffrey , Lu Lu , Williams Robert W. , Webby Richard J. 
Published: August 2009

ABSTRACTDespite the prevalence of H5N1 influenza viruses in global avian populations, comparatively few cases have been diagnosed in humans. Although viral factors almost certainly play a role in limiting human infection and disease, host genetics most likely contribute substantially. To model host factors in the context of influenza virus infection, we determined the lethal dose of a highly pathogenic H5N1 virus (A/Hong Kong/213/03) in C57BL/6J and DBA/2J mice and identified genetic elements associated with survival after infection. The lethal dose in these hosts varied by 4 logs and was associated with differences in replication kinetics and increased production of proinflammatory cytokines CCL2 and tumor necrosis factor alpha in susceptible DBA/2J mice. Gene mapping with recombinant inbred BXD strains revealed five loci orQivr(quantitative trait loci forinfluenzavirusresistance) located on chromosomes 2, 7, 11, 15, and 17 associated with resistance to H5N1 virus. In conjunction with gene expression profiling, we identified a number of candidate susceptibility genes. One of the validated genes, the hemolytic complement gene, affected virus titer 7 days after infection. We conclude that H5N1 influenza virus-induced pathology is affected by a complex and multigenic host component.
Human Immunodeficiency Virus Type 1 Protease-Correlated Cleavage Site Mutations Enhance Inhibitor Resistance
Authors: Kolli Madhavi , Stawiski Eric , Chappey Colombe , Schiffer Celia A. 
Published: August 2009

ABSTRACT
Drug resistance is an important cause of antiretroviral therapy failure in human immunodeficiency virus (HIV)-infected patients. Mutations in the protease render the virus resistant to protease inhibitors (PIs). Gag cleavage sites also mutate, sometimes correlating with resistance mutations in the protease, but their contribution to resistance has not been systematically analyzed. The present study examines mutations in Gag cleavage sites that associate with protease mutations and the impact of these associations on drug susceptibilities. Significant associations were observed between mutations in the nucleocapsid-p1 (NC-p1) and p1-p6 cleavage sites and various PI resistance-associated mutations in the protease. Several patterns were frequently observed, including mutations in the NC-p1 cleavage site in combination with I50L, V82A, and I84V within the protease and mutations within the p1-p6 cleavage site in combination with D30N, I50V, and I84V within the protease. For most patterns, viruses with mutations both in the protease and in either cleavage site were significantly less susceptible to specific PIs than viruses with mutations in the protease alone. Altered PI resistance in HIV-1 was found to be associated with the presence of Gag cleavage site mutations. These studies suggest that associated cleavage site mutations may contribute to PI susceptibility in highly specific ways depending on the particular combinations of mutations and inhibitors. Thus, cleavage site mutations should be considered when assessing the level of PI resistance.
Preinfection Human Immunodeficiency Virus (HIV)-Specific Cytotoxic T Lymphocytes Failed To Prevent HIV Type 1 Infection from Strains Genetically Unrelated to Viruses in Long-Term Exposed Partners
Authors: Liu Yi , Woodward Amanda , Zhu Haiying , Andrus Thomas , McNevin John , Lee Jean , Mullins James I. , Corey Lawrence , McElrath M. Juliana , Zhu Tuofu 
Published: August 2009

ABSTRACT

Understanding the mechanisms underlying potential altered susceptibility to human immunodeficiency virus type 1 (HIV-1) infection in highly exposed seronegative (ES) individuals and the later clinical consequences of breakthrough infection can provide insight into strategies to control HIV-1 with an effective vaccine. From our Seattle ES cohort, we identified one individual (LSC63) who seroconverted after over 2 years of repeated unprotected sexual contact with his HIV-1-infected partner (P63) and other sexual partners of unknown HIV-1 serostatus. The HIV-1 variants infecting LSC63 were genetically unrelated to those sequenced from P63. This may not be surprising, since viral load measurements in P63 were repeatedly below 50 copies/ml, making him an unlikely transmitter. However, broad HIV-1-specific cytotoxic T-lymphocyte (CTL) responses were detected in LSC63 before seroconversion. Compared to those detected after seroconversion, these responses were of lower magnitude and half of them targeted different regions of the viral proteome. Strong HLA-B27-restricted CTLs, which have been associated with disease control, were detected in LSC63 after but not before seroconversion. Furthermore, for the majority of the protein-coding regions of the HIV-1 variants in LSC63 (except
gp41
,
nef
, and the 3′ half of
pol
), the genetic distances between the infecting viruses and the viruses to which he was exposed through P63 (termed the exposed virus) were comparable to the distances between random subtype B HIV-1 sequences and the exposed viruses. These results suggest that broad preinfection immune responses were not able to prevent the acquisition of HIV-1 infection in LSC63, even though the infecting viruses were not particularly distant from the viruses that may have elicited these responses.
Human Immunodeficiency Virus Type 1 Mediates Global Disruption of Innate Antiviral Signaling and Immune Defenses within Infected Cells
Authors: Doehle Brian P. , Hladik Florian , McNevin John P. , McElrath M. Juliana , Gale Michael 
Published: August 2009

ABSTRACTInterferon regulatory factor 3 (IRF-3) is essential for innate intracellular immune defenses that limit virus replication, but these defenses fail to suppress human immunodeficiency virus (HIV) infection, which can ultimately associate with opportunistic coinfections and the progression to AIDS. Here, we examined antiviral defenses in CD4+cells during virus infection and coinfection, revealing that HIV type 1 (HIV-1) directs a global disruption of innate immune signaling and supports a coinfection model through suppression of IRF-3. T cells responded to paramyxovirus infection to activate IRF-3 and interferon-stimulated gene expression, but they failed to mount a response against HIV-1. The lack of response associated with a marked depletion of IRF-3 but not IRF-7 in HIV-1-infected cells, which supported robust viral replication, whereas ectopic expression of active IRF-3 suppressed HIV-1 infection. IRF-3 depletion was dependent on a productive HIV-1 replication cycle and caused the specific disruption of Toll-like receptor and RIG-I-like receptor innate immune signaling that rendered cells permissive to secondary virus infection. IRF-3 levels were reduced in vivo within CD4+T cells from patients with acute HIV-1 infection but not from long-term nonprogressors. Our results indicate that viral suppression of IRF-3 promotes HIV-1 infection by disrupting IRF-3-dependent signaling pathways and innate antiviral defenses of the host cell. IRF-3 may direct an innate antiviral response that regulates HIV-1 replication and viral set point while governing susceptibility to opportunistic virus coinfections.
Drastic Reduction in the Production of Subviral Particles Does Not Impair Hepatitis B Virus Virion Secretion
Authors: Garcia Tamako , Li Jisu , Sureau Camille , Ito Kiyoaki , Qin Yanli , Wands Jack , Tong Shuping 
Published: August 2009

ABSTRACT

Hepatitis B virus (HBV) contains three coterminal envelope proteins on the virion surface: large (L), middle (M), and small (S). The M and S proteins are also secreted as empty “subviral particles,” which exceed virions by at least 1,000-fold. The S protein serves as the morphogenic factor for both types of particles, while the L protein is required only for virion formation. We found that cotransfecting replication constructs with a small dose of the expression construct for the missing L, M, and S proteins reconstituted efficient virion secretion but only 5 to 10% of subviral particles. The L protein inhibited secretion of subviral particles in a dose-dependent manner, whereas a too-high or too-low L/S protein ratio inhibited virion secretion. Consistent with the results of cotransfection experiments, a point mutation at the −3 position of the S gene AUG codon reduced HBsAg secretion by 60 to 70% but maintained efficient virion secretion. Surprisingly, ablating M protein expression reduced virion secretion but markedly increased the maturity of virion-associated genomes, which could be reversed by providing
in trans
both L and M proteins but not just M protein. M protein stability was dependent on the coexpression of S protein. Our findings suggest that efficient HBV virion secretion could be maintained despite drastic reduction in subviral particle production, which supports the recent demonstration of separate secretion pathways adopted by the two types of particles. The M protein appears to facilitate core particle envelopment, thus shortening the window of plus strand DNA elongation.
Two-Step Conformational Changes in a Coronavirus Envelope Glycoprotein Mediated by Receptor Binding and Proteolysis
Authors: Matsuyama Shutoku , Taguchi Fumihiro 
Published: August 2009

ABSTRACT
The coronaviruses mouse hepatitis virus type 2 (MHV-2) and severe acute respiratory syndrome coronavirus (SARS-CoV) utilize proteases to enter host cells. Upon receptor binding, the spike (S) proteins of both viruses are activated for membrane fusion by proteases, such as trypsin, present in the environment, facilitating virus entry from the cell surface. In contrast, in the absence of extracellular proteases, these viruses can enter cells via an endosomal pathway and utilize endosomal cathepsins for S protein activation. We demonstrate that the MHV-2 S protein uses multistep conformational changes for membrane fusion. After interaction with a soluble form of the MHV receptor (CEACAM1a), the metastable form of S protein is converted to a stable trimer, as revealed by mildly denaturing sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Liposome-binding assays indicate that the receptor-bound virions are associated with the target membrane through hydrophobic interactions. The exposure of receptor-bound S protein to trypsin or cathepsin L (CPL) induces the formation of six-helix bundles (6HB), the final conformation. This trypsin- or CPL-mediated conversion to 6HB can be blocked by a heptad repeat peptide known to block the formation of 6HB. Although trypsin treatment enabled receptor-bound MHV-2 to enter from the cell surface, CPL failed to do so. Interestingly, consecutive treatment with CPL and then chlorpromazine enabled a portion of the virus to enter from cell surface. These results suggest that trypsin suffices for the induction of membrane fusion of receptor-primed S protein, but an additional unidentified cellular factor is required to trigger membrane fusion by CPL.
Peptides That Mimic the Amino-Terminal End of the Rabies Virus Phosphoprotein Have Antiviral Activity
Authors: Castel Guillaume , Chtéoui Mohamed , Caignard Grégory , Préhaud Christophe , Méhouas Stéphanie , Réal Eléonore , Jallet Corinne , Jacob Yves , Ruigrok Rob W. H. , Tordo Noël 
Published: August 2009

ABSTRACT

We wanted to develop a therapeutic approach against rabies disease by targeting the lyssavirus transcription/replication complex. Because this complex (nucleoprotein N-RNA template processed by the L polymerase and its cofactor, the phosphoprotein P) is similar to that of other negative-strand RNA viruses, we aimed to design broad-spectrum antiviral drugs that could be used as a complement to postexposure vaccination and immunotherapy. Recent progress in understanding the structure/function of the rabies virus P, N, and L proteins predicts that the amino-terminal end of P is an excellent target for destabilizing the replication complex because it interacts with both L (for positioning onto the N-RNA template) and N (for keeping N soluble, as needed for viral RNA encapsidation). Thus, peptides mimicking various lengths of the amino-terminal end of P have been evaluated, as follows: (i) for binding properties to the N-P-L partners by the two-hybrid method; (ii) for their capacity to inhibit the transcription/replication of a rabies virus minigenome encoding luciferase in BHK-21-T7 cells; and (iii) for their capacity to inhibit rabies virus infection of BHK-21-T7 cells and of two derivatives of the neuronal SK-N-SH cell line. Peptides P60 and P57 (the first 60 and first 57 NH
2
residues of P, respectively) exhibited a rapid, strong, and long-lasting inhibitory potential on luciferase expression (>95% from 24 h to 55 h). P42 was less efficient in its inhibition level (75% for 18 to 30 h) and duration (40% after 48 h). The most promising peptides were synthesized in tandem with the Tat sequence, allowing cell penetration. Their inhibitory effects were observed on BHK-21-T7 cells infected with rabies virus and Lagos bat virus but not with vesicular stomatitis virus. In neuronal cells, a significant inhibition of both nucleocapsid inclusions and rabies virus release was observed.