bluuebunny/crossref_metadata_2025 huggingface dataset:
Expansion of Human Cytomegalovirus (HCMV) Immediate-Early 1-Specific CD8+T Cells and Control of HCMV Replication after Allogeneic Stem Cell Transplantation
Authors: Sacre Karim , Nguyen Stéphanie , Deback Claire , Carcelain Guislaine , Vernant Jean-Paul , Leblond Véronique , Autran Brigitte , Dhedin Nathalie
Published: August 2008

Recovery of human cytomegalovirus (HCMV)-specific T immunity is critical for protection against HCMV disease in the early phase after allogeneic stem cell transplantation (SCT). Using an enzyme-linked immunospot assay with overlapping 15-mer peptides spanning pp65 and immediate-early 1 HCMV proteins, we investigated which HCMV-specific CD8+gamma interferon-positive (IFN-γ+) T-cell responses against pp65 and IE-1 were associated with control of HCMV replication in 48 recipients of unmanipulated HLA-matched allografts at 3 months (M3) and 6 months (M6) after SCT and in 23 donors. At M3 after SCT, the magnitude of the pp65-specific IFN-γ-producing CD8+T-cell response was greater in recipients than in donors, regardless of HCMV status. In contrast, expansion of IE-1-specific CD8+T cells at M3 was associated with protection against HCMV, and no patient with this expansion had HCMV replication at M3. At M6, the number of HCMV-specific CD8+T cells against both pp65 and IE-1 had expanded in all recipients, regardless of their previous levels of HCMV replication. The recipients' HCMV-specific CD8+T cells already detectable in related donors were predominantly targeting pp65. In contrast, in 40% of the cases, the HCMV-specific CD8+T cells in recipients involved new CD8+T-cell specificities undetectable in their related donors and preferentially targeting IE-1. Taken together, these results showed that the delay in reconstituting IE-1-specific CD8+T cells is correlated with the lack of protection against HCMV in the first 3 months after SCT. They also show that IE-1 is a major antigenic determinant of the early restoration of protective immunity to HCMV after SCT.
Gene Expression Analysis of Host Innate Immune Responses during Lethal H5N1 Infection in Ferrets
Authors: Cameron Cheryl M. , Cameron Mark J. , Bermejo-Martin Jesus F. , Ran Longsi , Xu Luoling , Turner Patricia V. , Ran Ran , Danesh Ali , Fang Yuan , Chan Pak-Kei M. , Mytle Nutan , Sullivan Timothy J. , Collins Tassie L. , Johnson Michael G. , Medina Julio C. , Rowe Thomas , Kelvin David J.
Published: August 2008

How viral and host factors contribute to the severe pathogenicity of the H5N1 subtype of avian influenza virus infection in humans is poorly understood. We identified three clusters of differentially expressed innate immune response genes in lungs from H5N1 (A/Vietnam/1203/04) influenza virus-infected ferrets by oligonucleotide microarray analysis. Interferon response genes were more strongly expressed in H5N1-infected ferret lungs than in lungs from ferrets infected with the less pathogenic H3N2 subtype. In particular, robust CXCL10 gene expression in H5N1-infected ferrets led us to test the pathogenic role of signaling via CXCL10's cognate receptor, CXCR3, during H5N1 influenza virus infection. Treatment of H5N1-infected ferrets with the drug AMG487, a CXCR3 antagonist, resulted in a reduction of symptom severity and delayed mortality compared to vehicle treatment. We contend that unregulated host interferon responses are at least partially responsible for the severity of H5N1 infection and provide evidence that attenuating the CXCR3 signaling pathway improves the clinical course of H5N1 infection in ferrets.
Regulatory Role of CD1d in Neurotropic Virus Infection
Authors: Tsunoda Ikuo , Tanaka Tomoko , Fujinami Robert S.
Published: August 2008

The GDVII strain of Theiler's murine encephalomyelitis virus (TMEV) causes an acute fatal polioencephalomyelitis in mice. Infection of susceptible mice with the DA strain of TMEV results in an acute polioencephalomyelitis followed by chronic immune-mediated demyelination with virus persistence in the central nervous system (CNS); DA virus infection is used as an animal model for multiple sclerosis. CD1d-restricted natural killer T (NKT) cells can contribute to viral clearance and regulation of autoimmune responses. To investigate the role of CD1d in TMEV infection, we first infected CD1d-deficient mice (CD1−/−) and wild-type BALB/c mice with GDVII virus. Wild-type mice were more resistant to virus than CD1−/−mice (50% lethal dose titers: wild-type mice, 10 PFU; CD1−/−mice, 1.6 PFU). Wild-type mice had fewer viral antigen-positive cells with greater inflammation in the CNS than CD1−/−mice. Second, an analysis of DA virus infection in CD1−/−mice was conducted. Although both wild-type and CD1−/−mice had similar clinical signs during the first 2 weeks after infection, CD1−/−mice had an increase in neurological deficits over those observed in wild-type mice at 3 to 5 weeks after infection. Although wild-type mice had no demyelination, 20 and 60% of CD1−/−mice developed demyelination at 3 and 5 weeks after infection, respectively. TMEV-specific lymphoproliferative responses, interleukin-4 (IL-4) production, and IL-4/gamma interferon ratios were higher in CD1−/−mice than in wild-type mice. Thus, CD1d-restricted NKT cells may play a protective role in TMEV-induced neurological disease by alteration of the cytokine profile and virus-specific immune responses.
Vaccinia Virus WR53.5/F14.5 Protein Is a New Component of Intracellular Mature Virus and Is Important for Calcium-Independent Cell Adhesion and Vaccinia Virus Virulence in Mice
Authors: Izmailyan Roza , Chang Wen
Published: August 2008

The vaccinia virus WR53.5L/F14.5L gene encodes a small conserved protein that was not detected previously. However, additional proteomic analyses of different vaccinia virus isolates and strains revealed that the WR53.5 protein was incorporated into intracellular mature virus (IMV). The WR53.5 protein contains a putative N-terminal transmembrane region and a short C-terminal region. Protease digestion removed the C terminus of WR53.5 protein from IMV particles, suggesting a similar topology to that of the IMV type II transmembrane protein. We generated a recombinant vaccinia virus, vi53.5L, that expressed WR53.5 protein under isopropyl-β-
d
-thiogalactopyranoside (IPTG) regulation and found that the vaccinia virus life cycle proceeded normally with or without IPTG, suggesting that WR53.5 protein is not essential for vaccinia virus growth in cell cultures. Interestingly, the C-terminal region of WR53.5 protein was exposed on the cell surface of infected cells and mediated calcium-independent cell adhesion. Finally, viruses with inactivated WR53.5L gene expression exhibited reduced virulence in mice when animals were inoculated intranasally, demonstrating that WR53.5 protein was required for virus virulence in vivo. In summary, we identified a new vaccinia IMV envelope protein, WR53.5, that mediates cell adhesion and is important for virus virulence in vivo.
Kinesin KIF4 Regulates Intracellular Trafficking and Stability of the Human Immunodeficiency Virus Type 1 Gag Polyprotein
Authors: Martinez Nathaniel W. , Xue Xiaoxiao , Berro Reem G. , Kreitzer Geri , Resh Marilyn D.
Published: August 2008

Retroviral Gag proteins are synthesized as soluble, myristoylated precursors that traffic to the plasma membrane and promote viral particle production. The intracellular transport of human immunodeficiency virus type 1 (HIV-1) Gag to the plasma membrane remains poorly understood, and cellular motor proteins responsible for Gag movement are not known. Here we show that disrupting the function of KIF4, a kinesin family member, slowed temporal progression of Gag through its trafficking intermediates and inhibited virus-like particle production. Knockdown of KIF4 also led to increased Gag degradation, resulting in reduced intracellular Gag protein levels; this phenotype was rescued by reintroduction of KIF4. When KIF4 function was blocked, Gag transiently accumulated in discrete, perinuclear, nonendocytic clusters that colocalized with endogenous KIF4, with Ubc9, an E2 SUMO-1 conjugating enzyme, and with SUMO. These studies identify a novel transit station through which Gag traffics en route to particle assembly and highlight the importance of KIF4 in regulating HIV-1 Gag trafficking and stability.
Transactivation of Cellular Genes Involved in Nucleotide Metabolism by the Regulatory IE1 Protein of Murine Cytomegalovirus Is Not Critical for Viral Replicative Fitness in Quiescent Cells and Host Tissues
Authors: Wilhelmi Vanessa , Simon Christian O. , Podlech Jürgen , Böhm Verena , Däubner Torsten , Emde Simone , Strand Dennis , Renzaho Angélique , Lemmermann Niels A. W. , Seckert Christof K. , Reddehase Matthias J. , Grzimek Natascha K. A.
Published: August 2008

Despite its high coding capacity, murine CMV (mCMV) does not encode functional enzymes for nucleotide biosynthesis. It thus depends on cellular enzymes, such as ribonucleotide reductase (RNR) and thymidylate synthase (TS), to be supplied with deoxynucleoside triphosphates (dNTPs) for its DNA replication. Viral transactivation of these cellular genes in quiescent cells of host tissues is therefore a parameter of viral fitness relevant to pathogenicity. Previous work has shown that the IE1, but not the IE3, protein of mCMV transactivates RNR and TS gene promoters and has revealed an in vivo attenuation of the mutant virus mCMV-ΔIE1. It was attractive to propose the hypothesis that lack of transactivation by IE1 and a resulting deficiency in the supply of dNTPs are the reasons for growth attenuation. Here, we have tested this hypothesis with the mutant virus mCMV-IE1-Y165C expressing an IE1 protein that selectively fails to transactivate RNR and TS in quiescent cells upon transfection while maintaining the capacity to disperse repressive nuclear domains (ND10). Our results confirm in vivo attenuation of mCMV-ΔIE1, as indicated by a longer doubling time in host organs, whereas mCMV-IE1-Y165C replicated like mCMV-WT and the revertant virus mCMV-IE1-C165Y. Notably, the mutant virus transactivated RNR and TS upon infection of quiescent cells, thus indicating that IE1 is not the only viral transactivator involved. We conclude that transactivation of cellular genes of dNTP biosynthesis is ensured by redundancy and that attenuation of mCMV-ΔIE1 results from the loss of other critical functions of IE1, with its function in the dispersal of ND10 being a promising candidate.
Herpes Simplex Virus Type 2-Induced Mortality following Genital Infection Is Blocked by Anti-Tumor Necrosis Factor Alpha Antibody in CXCL10-Deficient Mice
Authors: Thapa Manoj , Carr Daniel J. J.
Published: August 2008

The role of tumor necrosis factor alpha (TNF-α) was evaluated for CXCL10-deficient (CXCL10−/−) mice which succumbed to genital herpes simplex virus type 2 (HSV-2) infection and possessed elevated levels of virus and TNF-α but not other cytokines in the central nervous system (CNS) and vaginal tissue within the first 7 days following virus exposure. Anti-TNF-α but not control antibody treatment offsets the elevated mortality rate of CXCL10−/−mice, despite increased CNS viral titers. In addition, TNF-α neutralization suppressed recruitment of leukocyte subpopulations into the CNS, which is associated with reduced CCL2 and CXCL9 expression. Collectively, the results implicate TNF-α as the principal mediator of mortality in response to genital HSV-2 infection.
Neonatal Neural Progenitor Cells and Their Neuronal and Glial Cell Derivatives Are Fully Permissive for Human Cytomegalovirus Infection
Authors: Luo Min Hua , Schwartz Philip H. , Fortunato Elizabeth A.
Published: August 2008

Congenital human cytomegalovirus (HCMV) infection causes central nervous system structural abnormalities and functional disorders, affecting both astroglia and neurons with a pathogenesis that is only marginally understood. To better understand HCMV's interactions with such clinically important cell types, we utilized neural progenitor cells (NPCs) derived from neonatal autopsy tissue, which can be differentiated down either glial or neuronal pathways. Studies were performed using two viral isolates, Towne (laboratory adapted) and TR (a clinical strain), at a multiplicity of infection of 3. NPCs were fully permissive for both strains, expressing the full range of viral antigens (Ags) and producing relatively large numbers of infectious virions. NPCs infected with TR showed delayed development of cytopathic effects (CPE) and replication centers and shed less virus. This pattern of delay for TR infections held true for all cell types tested. Differentiation of NPCs was carried out for 21 days to obtain either astroglia (>95% GFAP
+
) or a 1:5 mixed neuron/astroglia population (β-tubulin III
+
/GFAP
+
). We found that both of these differentiated populations were fully permissive for HCMV infection and produced substantial numbers of infectious virions. Utilizing a difference in plating efficiencies, we were able to enrich the neuron population to ∼80% β-tubulin III
+
cells. These β-tubulin III
+
-enriched populations remained fully permissive for infection but were very slow to develop CPE. These infected enriched neurons survived longer than either NPCs or astroglia, and a small proportion were alive until at least 14 days postinfection. These surviving cells were all β-tubulin III
+
and showed viral Ag expression. Surprisingly, some cells still exhibited extended processes, similar to mock-infected neurons. Our findings strongly suggest neurons as reservoirs for HCMV within the developing brain.
Different Types of nsP3-Containing Protein Complexes in Sindbis Virus-Infected Cells
Authors: Gorchakov Rodion , Garmashova Natalia , Frolova Elena , Frolov Ilya
Published: August 2008

Alphaviruses represent a serious public health threat and cause a wide variety of diseases, ranging from severe encephalitis, which can result in death or neurological sequelae, to mild infection, characterized by fever, skin rashes, and arthritis. In the infected cells, alphaviruses express only four nonstructural proteins, which function in the synthesis of virus-specific RNAs and in modification of the intracellular environment. The results of our study suggest that Sindbis virus (SINV) infection in BHK-21 cells leads to the formation of at least two types of nsP3-containing complexes, one of which was found in association with the plasma membrane and endosome-like vesicles, while the second was coisolated with cell nuclei. The latter complexes could be solubilized only with the cytoskeleton-destabilizing detergent. Besides viral nsPs, in the mammalian cells, both complexes contained G3BP1 and G3BP2 (which were found in different ratios), YBX1, and HSC70. Rasputin, an insect cell-specific homolog of G3BP1, was found in the nsP3-containing complexes isolated from mosquito cells, which was suggestive of a high conservation of the complexes in the cells of both vertebrate and invertebrate origin. The endosome- and plasma membrane-associated complexes contained a high concentration of double-stranded RNAs (dsRNAs), which is indicative of their function in viral-RNA synthesis. The dsRNA synthesis is likely to efficiently proceed on the plasma membrane, and at least some of the protein-RNA complexes would then be transported into the cytosol in association with the endosome-like vesicular organelles. These findings provide new insight into the mechanism of SINV replication and virus-host cell interactions.
Human Immunodeficiency Virus Type 1 Replication and Regulation of APOBEC3G by Peptidyl Prolyl Isomerase Pin1
Authors: Watashi Koichi , Khan Mohammad , Yedavalli Venkat R. K. , Yeung Man Lung , Strebel Klaus , Jeang Kuan-Teh
Published: August 2008

APOBEC3G (A3G) is a cytidine deaminase that restricts human immunodeficiency virus type 1 (HIV-1) replication. HIV-1 synthesizes a viral infectivity factor (Vif) to counter A3G restriction. Currently, it is poorly understood how A3G expression/activity is regulated by cellular factors. Here, we show that the prolyl isomerase Pin1 protein modulates A3G expression. Pin1 was found to be an A3G-interacting protein that reduces A3G expression and its incorporation into HIV-1 virion, thereby limiting A3G-mediated restriction of HIV-1. Intriguingly, HIV-1 infection modulates the phosphorylation state of Pin1, enhancing its ability to moderate A3G activity. These new findings suggest a potential Vif-independent way for HIV-1 to moderate the cellular action of A3G.
Construction of a Fully Retargeted Herpes Simplex Virus 1 Recombinant Capable of Entering Cells Solely via Human Epidermal Growth Factor Receptor 2
Authors: Menotti Laura , Cerretani Arianna , Hengel Hartmut , Campadelli-Fiume Gabriella
Published: August 2008

A novel frontier in the treatment of tumors that are difficult to treat is oncolytic virotherapy, in which a replication-competent virus selectively infects and destroys tumor cells. Herpes simplex virus (HSV) represents a particularly attractive system. Effective retargeting to tumor-specific receptors has been achieved by insertion in gD of heterologous ligands. Previously, our laboratory generated an HSV retargeted to human epidermal growth factor receptor 2 (HER2), a receptor overexpressed in about one-third of mammary tumors and in some ovarian tumors. HER2 overexpression correlates with increased metastaticity and poor prognosis. Because HER2 has no natural ligand, the inserted ligand was a single-chain antibody to HER2. The objective of this work was to genetically engineer an HSV that selectively targets the HER2-expressing tumor cells and that has lost the ability to enter cells through the natural gD receptors, HVEM and nectin1. Detargeting from nectin1 was attempted by two different strategies, point mutations and insertion of the single-chain antibody at a site in gD different from previously described sites of insertion. We report that point mutations at gD amino acids 34, 215, 222, and 223 failed to generate a nectin1-detargeted HSV. An HSV simultaneously detargeted from nectin1 and HVEM and retargeted to HER2 was successfully engineered by moving the site of single-chain antibody insertion at residue 39, i.e., in front of the nectin1-interacting surface and not lateral to it, and by deleting amino acid residues 6 to 38. The resulting recombinant, R-LM113, entered cells and spread from cell to cell solely via HER2.
Small-Ruminant Lentivirus Enhances PrP
Sc
Accumulation in Cultured Sheep Microglial Cells
Authors: Stanton James B. , Knowles Donald P. , O'Rourke Katherine I. , Herrmann-Hoesing Lynn M. , Mathison Bruce A. , Baszler Timothy V.
Published: August 2008

Sheep scrapie is the prototypical transmissible spongiform encephalopathy (prion disease), which has a fundamental pathogenesis involving conversion of normal cellular prion protein (PrP
C
[C superscript stands for cellular]) to disease-associated prion protein (PrP
Sc
[Sc superscript stands for sheep scrapie]). Sheep microglial cell cultures, derived from a
prnp
136VV/171QQ near-term fetal brain, were developed to study sheep scrapie in the natural host and to investigate potential cofactors in the prion conversion process. Two culture systems, a primary cell culture and a cell line transformed with the large T antigen of simian virus 40, were developed, and both were identified as microglial in origin as indicated by expression of several microglial phenotype markers. Following exposure to PrP
Sc
, sheep microglial cells demonstrated relatively low levels (transformed cell line) to high levels (primary cell line) of PrP
Sc
accumulation over time. The accumulated PrP
Sc
demonstrated protease resistance, an inferred beta-sheet conformation (as determined by a commercial enzyme-linked immunosorbent assay), specific inhibition by anti-PrP antibodies, and was transmissible in a dose-dependent manner. Primary microglia coinfected with a small-ruminant lentivirus (caprine arthritis encephalitis virus-Cork strain) and PrP
Sc
demonstrated an approximately twofold increase in PrP
Sc
accumulation compared to that of primary microglia infected with PrP
Sc
alone. The results demonstrate the in vitro utility of PrP
Sc
-permissive sheep microglial cells in investigating the biology of natural prion diseases and show that small-ruminant lentiviruses enhance prion conversion in cultured sheep microglia.
Block to the Production of Full-Length B19 Virus Transcripts by Internal Polyadenylation Is Overcome by Replication of the Viral Genome
Authors: Guan Wuxiang , Cheng Fang , Yoto Yuko , Kleiboeker Steve , Wong Susan , Zhi Ning , Pintel David J. , Qiu Jianming
Published: August 2008

The pre-mRNA processing strategy of the B19 virus is unique among parvoviruses. B19 virus-generated pre-mRNAs are transcribed from a single promoter and are extensively processed by alternative splicing and alternative polyadenylation to generate 12 transcripts. Blockage of the production of full-length B19 virus transcripts at the internal polyadenylation site [(pA)p] was previously reported to be a limiting step in B19 virus permissiveness. We show here that in the absence of genome replication, internal polyadenylation of B19 virus RNAs at (pA)p is favored in cells which are both permissive and nonpermissive for B19 viral replication. Replication of the B19 virus genome, however, introduced either by viral infection or by transfection of an infectious clone into permissive cells or forced by heterologous replication systems in nonpermissive cells, enhanced readthrough of (pA)p and the polyadenylation of B19 virus transcripts at the distal site [(pA)d]. Therefore, replication of the genome facilitates the generation of sufficient full-length transcripts that encode the viral capsid proteins and the essential 11-kDa nonstructural protein. Furthermore, we show that polyadenylation of B19 viral RNA at (pA)p likely competes with splicing at the second intron. Thus, we conclude that replication of the B19 virus genome is the primary limiting step governing B19 virus tropism.
Retroviral Nucleocapsid Proteins Display Nonequivalent Levels of Nucleic Acid Chaperone Activity
Authors: Stewart-Maynard Kristen M. , Cruceanu Margareta , Wang Fei , Vo My-Nuong , Gorelick Robert J. , Williams Mark C. , Rouzina Ioulia , Musier-Forsyth Karin
Published: August 2008

Human immunodeficiency virus type 1 (HIV-1) nucleocapsid protein (NC) is a nucleic acid chaperone that facilitates the remodeling of nucleic acids during various steps of the viral life cycle. Two main features of NC's chaperone activity are its abilities to aggregate and to destabilize nucleic acids. These functions are associated with NC's highly basic character and with its zinc finger domains, respectively. While the chaperone activity of HIV-1 NC has been extensively studied, less is known about the chaperone activities of other retroviral NCs. In this work, complementary experimental approaches were used to characterize and compare the chaperone activities of NC proteins from four different retroviruses: HIV-1, Moloney murine leukemia virus (MLV), Rous sarcoma virus (RSV), and human T-cell lymphotropic virus type 1 (HTLV-1). The different NCs exhibited significant differences in their overall chaperone activities, as demonstrated by gel shift annealing assays, decreasing in the order HIV-1 ∼ RSV > MLV ≫ HTLV-1. In addition, whereas HIV-1, RSV, and MLV NCs are effective aggregating agents, HTLV-1 NC, which exhibits poor overall chaperone activity, is unable to aggregate nucleic acids. Measurements of equilibrium binding to single- and double-stranded oligonucleotides suggested that all four NC proteins have moderate duplex destabilization capabilities. Single-molecule DNA-stretching studies revealed striking differences in the kinetics of nucleic acid dissociation between the NC proteins, showing excellent correlation between nucleic acid dissociation kinetics and overall chaperone activity.
Bap31 Is a Novel Target of the Human Papillomavirus E5 Protein
Authors: Regan Jennifer A. , Laimins Laimonis A.
Published: August 2008

The E5 proteins of human papillomaviruses (HPVs) are small hydrophobic proteins that are expressed in the early and late stages of the viral life cycle; however, their role in HPV pathogenesis is not clearly understood. In this study, a split-ubiquitin yeast (
Saccharomyces cerevisiae
) two-hybrid system was used to identify B-cell-associated protein 31 (Bap31) as a binding partner of HPV E5 proteins. The association of these proteins was confirmed by coimmunoprecipitation of complexes of Bap31 with either HPV type 16 (HPV16) or HPV31 E5. In addition, Bap31 and E5 were found to colocalize in perinuclear patterns consistent with localization to the endoplasmic reticulum. Mutational analysis of E5 identified amino acids in the extreme C terminus as important for stabilizing the interaction with Bap31. Deletion of these C-terminal amino acids of E5 in the context of complete HPV31 genomes resulted in impaired proliferative capacity of HPV-positive keratinocytes following differentiation. When small interfering RNAs were used to reduce the levels of Bap31, the proliferative ability of HPV-positive keratinocytes upon differentiation was also reduced, implicating Bap31 as a regulator of this process. These studies identify a novel binding partner of the high-risk HPV E5 proteins and provide insight into how the E5 proteins may modulate the life cycle in differentiating cells.
In Vivo Competence of Murine Cytomegalovirus under the Control of the Human Cytomegalovirus Major Immediate-Early Enhancer in the Establishment of Latency and Reactivation
Authors: Gustems Montse , Busche Andreas , Messerle Martin , Ghazal Peter , Angulo Ana
Published: August 2008

The human cytomegalovirus (HCMV) major immediate-early enhancer has been postulated to play a pivotal role in the control of latency and reactivation. However, the absence of an animal model has obstructed a direct test of this hypothesis. Here we report on the establishment of an in vivo, experimentally tractable system for quantitatively investigating physiological functions of the HCMV enhancer. Using a neonate BALB/c mouse model, we show that a chimeric murine CMV under the control of the HCMV enhancer is competent in vivo, replicating in key organs of mice with acute CMV infection and exhibiting latency/reactivation features comparable for the most part to those of the parental and revertant viruses.
Integration Site Preference of Xenotropic Murine Leukemia Virus-Related Virus, a New Human Retrovirus Associated with Prostate Cancer
Authors: Kim Sanggu , Kim Namshin , Dong Beihua , Boren David , Lee Serena A. , Das Gupta Jaydip , Gaughan Christina , Klein Eric A. , Lee Christopher , Silverman Robert H. , Chow Samson A.
Published: August 2008

Xenotropic murine leukemia virus-related virus (XMRV) is a new human gammaretrovirus identified in prostate cancer tissue from patients homozygous for a reduced-activity variant of the antiviral enzyme RNase L. Neither a casual relationship between XMRV infection and prostate cancer nor a mechanism of tumorigenesis has been established. To determine the integration site preferences of XMRV and the potential risk of proviral insertional mutagenesis, we carried out a genome-wide analysis of viral integration sites in the prostate cell line DU145 after an acute XMRV infection and compared the integration site pattern of XMRV with those found for murine leukemia virus and two human retroviruses, human immunodeficiency virus type 1 and human T-cell leukemia virus type 1. Among all retroviruses analyzed, XMRV has the strongest preference for transcription start sites, CpG islands, DNase-hypersensitive sites, and gene-dense regions; all are features frequently associated with structurally open transcription regulatory regions of a chromosome. Analyses of XMRV integration sites in tissues from prostate cancer patients found a similar preference for the aforementioned chromosomal features. Additionally, XMRV integration sites in cancer tissues were associated with cancer breakpoints, common fragile sites, microRNA, and cancer-related genes, suggesting a selection process that favors certain chromosomal integration sites. In both acutely infected cells and cancer tissues, no common integration site was detected within or near proto-oncogenes or tumor suppressor genes. These results are consistent with a model in which XMRV may contribute to tumorigenicity via a paracrine mechanism.
CD20, CD3, and CD40 Ligand Microclusters Segregate Three-Dimensionally In Vivo at B-Cell-T-Cell Immunological Synapses after Viral Immunity in Primate Brain
Authors: Barcia Carlos , Gomez Aurora , de Pablos Vicente , Fernández-Villalba Emiliano , Liu Chunyan , Kroeger Kurt M. , Martín Javier , Barreiro Andrés Fernández , Castro Maria G. , Lowenstein Pedro R. , Herrero Maria-Trinidad
Published: August 2008

The clearance of virally infected cells from the brain is mediated by T cells that engage antigen-presenting cells to form supramolecular activation clusters at the immunological synapse. However, after clearance, the T cells persist at the infection site and remain activated locally. In the present work the long-term interactions of immune cells in brains of monkeys were imaged in situ 9 months after the viral inoculation. After viral immunity, the persistent infiltration of T cells and B cells was observed at the infection sites. T cells showed evidence of T-cell receptor signaling as a result of contacts with B cells. Three-dimensional analysis of B-cell-T-cell synapses showed clusters of CD3 in T cells and the segregation of CD20 in B cells, involving the recruitment of CD40 ligand at the interface. These results demonstrate that immunological synapses between B cells and T cells forming three-dimensional microclusters occur in vivo in the central nervous system and suggest that these interactions may be involved in the lymphocyte activation after viral immunity at the original infection site.
Small-Molecule Effectors of Hepatitis B Virus Capsid Assembly Give Insight into Virus Life Cycle
Authors: Bourne Christina , Lee Sejin , Venkataiah Bollu , Lee Angela , Korba Brent , Finn M. G. , Zlotnick Adam
Published: August 2008

The relationship between the physical chemistry and biology of self-assembly is poorly understood, but it will be critical to quantitatively understand infection and for the design of antivirals that target virus genesis. Here we take advantage of heteroaryldihydropyrimidines (HAPs), which affect hepatitis B virus (HBV) assembly, to gain insight and correlate in vitro assembly with HBV replication in culture. Based on a low-resolution crystal structure of a capsid-HAP complex, a closely related series of HAPs were designed and synthesized. These differentially strengthen the association between neighboring capsid proteins, alter the kinetics of assembly, and give rise to aberrant structures incompatible with a functional capsid. The chemical nature of the HAP variants correlated well with the structure of the HAP binding pocket. The thermodynamics and kinetics of in vitro assembly had strong and predictable effects on product morphology. However, only the kinetics of in vitro assembly had a strong correlation with inhibition of HBV replication in HepG2.2.15 cells; there was at best a weak correlation between assembly thermodynamics and replication. The correlation between assembly kinetics and virus suppression implies a competition between successful assembly and misassembly, small molecule induced or otherwise. This is a predictive and testable model for the mechanism of action of assembly effectors.
Functions of Varicella-Zoster Virus ORF23 Capsid Protein in Viral Replication and the Pathogenesis of Skin Infection
Authors: Chaudhuri Vaishali , Sommer Marvin , Rajamani Jaya , Zerboni Leigh , Arvin Ann M.
Published: August 2008

The assembly of herpesvirus capsids is a complex process involving interactions of multiple proteins in the cytoplasm and in the nucleus. Based on comparative genome analyses, varicella-zoster virus (VZV) open reading frame 23 (ORF23) encodes a conserved capsid protein, referred to as VP26 (UL35) in other alphaherpesviruses. Mutagenesis using a VZV bacterial artificial chromosome system showed that ORF23 was dispensable for replication in vitro. However, the absence of ORF23 disrupted capsid assembly in a melanoma cell line. Expression of ORF23 as a red fluorescent protein (RFP) fusion protein appeared to have a dominant negative effect on replication that was rescued by ORF23 expression from a nonnative site in the VZV genome. In contrast to its VP26 homolog, ORF23 has an intrinsic nuclear localization capacity that was mapped to an SRSRVV motif at residues 229 to 234 in the extreme C terminus of ORF23. In addition, coexpression with ORF23 resulted in nuclear import of the major capsid protein, ORF40. VZV ORF33.5 also translocated ORF40, which may provide a redundant mechanism in vitro but appears insufficient to overcome the dominant negative effect of the monomeric RFP-ORF23 (mRFP23) fusion protein. ORF23 was required for VZV infection of human skin xenografts, indicating that ORF33.5 does not compensate for lack of ORF23 in vivo. These observations suggest a model of VZV capsid assembly in which nuclear transport of the major capsid protein and associated proteins requires ORF23 during VZV replication in the human host. If so, ORF23 expression could be a target for a novel antiviral drug against VZV.
SOME OBSERVATIONS ON THE FORMATION OF GLYCERIC ACID DURING PHOTOSYNTHESIS EXPERIMENTS
Authors: Mortimer D. C.
Published: August 2008

The dephosphorylation of phosphoglyceric acid was tested under a variety of conditions. Hydrolysis of the ester by leaf homogenates was relatively slow, even when conditions favored enzymatic activity, and was not detected when frozen homogenate was extracted by boiling 80% ethanol. The results suggest that glyceric acid recovered in ethanol-soluble extracts of leaves following short-term photosynthesis experiments is not normally derived from phosphoglyceric acid. The route proposed for the synthesis of glyceric acid is related, with particular reference to soybean leaf, to an alternate pathway for photosynthetic assimilation which does not utilize phosphoglyceric acid.
CYTOTAXONOMY, PHYLOGENY, AND CANADIAN DISTRIBUTION OF CIRSIUM UNDULATUM AND CIRSIUM FLODMANII
Authors: Frankton C. , Moore R. J.
Published: August 2008

The morphology and specific differences of Cirsium undulatum (Nutt.) Spreng. and of C. flodmanii (Rydb.) Arthur are described and their Canadian distributions are reported in detail. The chromosome numbers are C. undulatum f. undulatum and f. album Farwell, 2n = 26; C. flodmanii f. flodmanii and f. albiflorum D. Löve, 2n = 22. The origin of four North American species of Cirsium that do not follow the world-wide base number 17 is discussed; it is postulated that reduction in number has occurred by translocations. The chromosomes of species with reduced numbers are larger than those of the unreduced species but the total length of the chromosomes of both groups is approximately the same.
A PRELIMINARY STUDY OF DIFFERENTIAL POLLEN GRAIN PRESERVATION
Authors: Sangster A. G. , Dale H. M.
Published: August 2008

Samples of pollen of three genera (Populus, Pinus, and Typha) which are normally shed at different times during the growing season were placed concurrently in four habitats (a pond, a lake, a swamp, and a bog). Their decomposition was traced through the summer and autumn of two seasons. The study shows that there is considerable variability in fossilization of pollen under identical conditions, and that the decomposition of the same pollen is dependent on the habitat, which affects not only the rate of decomposition but also the processes involved.
CYTOLOGICAL STUDIES OF THE DEVELOPMENT OF METAPHASE I IN HYBRIDS BETWEEN TRITICUM TIMOPHEEVI ZHUK. AND T. DURUM DESF.
Authors: Wagenaar E. B.
Published: August 2008

In two hybrids between Triticum timopheevi Zhuk. and T. durum Desf., which have irregular meioses, metaphase I was subdivided into four developmental stages, early, medium, late, and very late. This subdivision was based on the presence in the anthers of other stages that occurred together with metaphase I. It was then discovered that in metaphase I cell populations there was a progressive increase of univalents from the early and medium stages to the very late stage. This phenomenon can be explained on the assumption that metaphase I is of shorter duration in the less irregular cells which pass into anaphase I earlier than the more irregular cells. As a consequence of this developmental phenomenon at metaphase I, the anaphase I and telophase I cells in the late anthers contained fewer lagging chromosomes than the anaphase I and telophase I cells in the very late anthers. Despite these numerical differences, the degrees of lagging were remarkably similar in both stages; approximately 70% of these univalents lagged at late and very late stages in both hybrids.During metaphase I many univalents of the irregular cells moved towards the equatorial plate, became oriented, and lagged at anaphase I and telophase I. It was found that the univalents of the least irregular cells accumulated somewhat faster at the plates than those of the more irregular cells.Considering the relationships between all of the available data, the hypothesis is advanced that when a certain number of univalents have accumulated at the equatorial plate a state of equilibrium is established and anaphase I is initiated. On the basis of this hypothesis an explanation of the trends observed at metaphase I is given.
CHROMOSOME NUMBERS AND TAXONOMIC NOTES ON NORTHERN GRASSES: IV. TRIBE FESTUCEAE: POA, AND PUCCINELLIA
Authors: Bowden Wray M.
Published: August 2008

Chromosome numbers and voucher specimens are recorded for some Canadian and Alaskan collections and a few mainland American collections of Poa and Puccinellia which belong to the tribe FESTUCEAE. The somatic chromosome numbers of the following are recorded: (1) Poa agassizensis, 2n = ca. 56; P. alpina, 2n = 32, 33, 39, and 42; P. ampla, 2n = 62; P. annua, 2n = 28; P. arctica, 2n = 56, 63, 70, ca. 75, 78, 82, 85, 86, and 106; P. arida, 2n = ca. 76; P. canbyi, 2n = 56, 70, and ca. 98; P. cusickii, 2n = 56, and 59; P. eminens, 2n = 42; P. fernaldiana, 2n = ca. 42; P. glauca, 2n = 42, 50, 56, 64, 65, 70, 75, and 78; P. glaucantha, 2n = 44; P. glaucifolia, 2n = 56, ca. 70, and ca. 100; P. interior, 2n = 28, 34, 42, and 56; P. juncifolia, 2n = ca. 60; P. leptocoma, 2n = 42; P. nervosa, 2n = 56, and ca. 74; P. palustris, 2n = 28, 30, and 32; P. pratensis, 2n = 28, 42, 44, 56, ca. 66, 67, 70, 74, ca. 78, ca. 82, 84, 86, 88, and ca. 124; P. saltuensis, 2n = 28; P. secunda, 2n = 42, 56, 70, ca. 78, 80. and ca. 90; P. stenantha, 2n = 42; and P. trivialis, 2n = 14.(2) Puccinellia ambigua, 2n = 56; P. americana, 2n = ca. 42 and 56; P. angustata, 2n = 42; P. borealis, 2n = 42; P. bruggemannii, 2n = 28; P. cusickii, 2n = 28; P. distans, 2n = 42; P. glabra, 2n = 42 and 56; P. hauptiana, 2n = 28; P. laurentiana, 2n = 42 and 56; P.longiglumis, 2n = 56; P. lucida, 2n = 42 and 56; P. nuttalliana, 2n = 42; P. × phryganodes, 2n = 21; P. vaginata, 2n = 56; and P. vahliana, 2n = 14.
THE SMUTS OF THE GENUS CINTRACTIA PARASITING KOBRESIA SPECIES
Authors: Kukkonen Ilkka
Published: August 2008

In this paper the modern concept of infection by species of Cintractia is briefly discussed. Three species are treated, two of which, C. elynae Sydow and C. kobresiae Mundkur, are redescribed, and the third is recognized as new, C. lindebergiae n. sp. The spore germination of C. elynae is described and illustrated. A short account of the host genus Kobresia is given with special reference to the species attacked by the above smuts.
STUDIES ON FUSARIUM WILT OF BANANAS: VII. FIELD CONTROL
Authors: Stover R. H. , Hildreth R. C. , Thornton N. C.
Published: August 2008

Flood fallow and silting were the only practices that consistently reduced incidence of banana wilt induced by F. oxysporum f. cubense (E. F. S.) Snyd. and Hans. The efficiency of flood fallow was greatly increased by interflood plowing, and in most locations by postflood fungicide drenches. Even these treatments, however, did not insure economic control in some areas. It was necessary, therefore, to select areas where soil factors tended to retard rapid disease development. Also, the absence of the banana host for a number of years previous to flood fallow appeared to influence treatment success.
SEED DEVELOPMENT IN ADHATODA VASICA NEES
Authors: Ram H. Y. Mohan
Published: August 2008

The events following fertilization up to the maturation of the seed in Adhatoda vasica have been described. The first two transverse divisions accompanying the division of the primary endosperm nucleus partition the embryo sac into (1) a micropylar chamber, (2) a chalazal chamber, and (3) a central endosperm chamber. The first two develop into binucleate haustorial structures, while the bulk of the endosperm is derived from the products of the central endosperm chamber. An unusual case of a micropylar haustorium containing four hypertrophied nuclei and four small nuclei has been observed. The central endosperm chamber is partly cellular and bears a tubular free nuclear basal apparatus. The latter, however, ultimately becomes cellular. The embryo development is of the Solanad type. The mature seed has only two layers of thin-walled endosperm cells and a wavy testa representing the persistent epidermis of the integument. The rest of the integument is consumed by the endosperm. The uneven surface of the testa results from unequal development of the integumentary parenchyma and not due to either unequal radial stretching of the epidermis or irregular growth of the endosperm as in Andrographis serpyllifolia.
Effects of osmolality, cytokinin, and organic acids on pollen callus formation in Triticale anthers
Authors: Chien Y. C. , Kao K. N.
Published: August 2008

Triticale anthers with pollen at middle to late uninucleated stages were cultured individually in Falcon micro test II tissue culture plates. The results indicate that when the anthers were cultured in the same growing conditions the differences in pollen callus formation among anthers from the first and second florets in the same spikelet were not statistically significant, whereas the differences in callus formation among anthers from different spikes (or plants) were statistically significant. These results suggest that comparisons of treatment effects should be made between samples consisting of anthers from the same spikelet only. Benzyl adenine (BA) and higher osmolality enhanced pollen callus induction, while a higher concentration of organic acids suppressed it. Under optimal conditions 31% of anthers formed callus, while the number of pollen calli per 100 seeded anthers was as high as 130. The pollen calli were able to develop into plants; however, the frequency was relatively low.
A study of the relationship between seed dormancy and pentose phosphate pathway activity in Avena fatua
Authors: Fuerst E. P. , Upadhyaya M. K. , Simpson G. M. , Naylor J. M. , Adkins S. W.
Published: August 2008

The hypothesis that loss of seed dormancy is associated with an increased activity of the pentose phosphate pathway (PPP) relative to glycolysis and the Krebs cycle was tested. The PPP activity was monitored by measuring the C6/C1 ratio in embryos excised from incubated caryopses of two genetically pure nondormant (ND) lines and in three dormant (D) lines of Avena fatua L., the wild oat. The C6/C1 ratios of all lines were similar at the commencement of incubation. In the two ND lines the ratio increased steadily prior to and during emergence of the radicle. In the three D lines the ratio increased during the first 24 h and then remained almost constant; there was no germination. When gibberellin treatment was used to overcome dormancy in the D lines, the C6/C1 ratio increased during the first 24 h in two of the lines and continued to increase parallel to germination in a manner similar to normal germination in ND lines. In the third D line, despite loss of dormancy from gibberellin treatment, the ratio did not increase after 24 h. Loss of dormancy during dry storage of seeds of a D-type pure line was accompanied by an increase in the C6/C1 ratio, as measured in freshly imbibed seeds. This indicates a decreased activity of the PPP relative to glycolysis and the Krebs cycle. These findings are contrary to Roberts's hypothesis that loss of dormancy in wild oats is associated with a relative decrease in the C6/C1 ratio.
The status of Stachys palustris (Labiatae) in North America
Authors: Mulligan Gerald A. , Munro Derek B. , McNeill John
Published: August 2008

The Stachys palustris L. complex in North America is composed of diploids, tetraploids, and hexaploids with the base number x = 17. Hexaploids are identical to S. palustris L. of Europe and are weedy plants introduced from that continent. Diploid and tetraploid plants of this complex are native to North America and should be referred to other species. The taxonomy of these native plants requires further study. Characteristics are presented that distinguish native diploids and tetraploids from the introduced weedy hexaploid S. palustris L.
Les nectaires extrafloraux pédicellés chez l’Ailanthus glandulosa
Authors: Clair-Maczulajtys Danielle , Bory Gérard
Published: August 2008

Extrafloral nectaries are present on the first leaves appearing when the buds open. Located at the base of the petiole, they are stalked and have an apical pore. They are initiated early on the margins of leaf primordia. These organs are interpreted as reduced stipules consisting of one axis with its vascular system with the extremity occupied by a nectary. A viscous and clear nectar is extruded from the pore. Without vascularization, there is never an apical gland. Stalked nectaries are also observed on the adaxial surface of the lamina. Described as teratological events, they correspond to the deviation of a vein perpendicular to the plane of the leaf. For aerial organs these nectaries show clearly the close connection between the vascular system and the glandular extremity. The tissue structure of these stalked nectaries is similar to that of the marginal nectaries. The morphological and physiological signification of these structures is discussed.
Flavonoids and affinities of some North American lupines
Authors: Nicholls K. W. , Bohm B. A.
Published: August 2008

Seventy-three taxa of the genus Lupinus in North America have been studied for their flavonoid constituents. In all, 56 compounds were isolated and either partially or totally characterized. Flavonoid types encountered were flavones, C-glycoflavones, isoflavones, flavonols, and, tentatively, flavanones. Further structural modifications seen were O-methylation, O-glycosylation at a variety of positions and acylation of several of the flavonoid glycosides. Principal-components analysis was performed on (i) presence–absence data of individual compounds with consideration given to relative concentrations and (ii) presence–absence of the types of flavonoid structures. Several groupings of taxa emerged from these analyses based upon the presence of unique compounds or combinations of compounds. In a number of these groups the chemical affinities paralleled morphological affinities closely. In all, 10 groupings were seen. Lupinus diffusus, a simple-leaved taxon from southeastern United States, stood apart from all groups.