bluuebunny/crossref_metadata_2025_split huggingface dataset:
The Herpes Simplex Virus Triplex Protein, VP23, Exists as a Molten Globule
Authors: Kirkitadze Marina D. , Barlow Paul N. , Price Nicholas C. , Kelly Sharon M. , Boutell Christopher J. , Rixon Frazer J. , McClelland David A. 
Published: December 2019

ABSTRACT

Two proteins, VP19C (50,260 Da) and VP23 (34,268 Da), make up the triplexes which connect adjacent hexons and pentons in the herpes simplex virus type 1 capsid. VP23 was expressed in
Escherichia coli
and purified to homogeneity by Ni-agarose affinity chromatography. In vitro capsid assembly experiments demonstrated that the purified protein was functionally active. Its physical status was examined by differential scanning calorimetry, ultracentrifugation, size exclusion chromatography, circular dichroism, fluorescence spectroscopy, and 8-anilino-1-naphthalene sulfonate binding studies. These studies established that the bacterially expressed VP23 exhibits properties consistent with its being in a partially folded, molten globule state. We propose that the molten globule represents a functionally relevant intermediate which is necessary to allow VP23 to undergo interaction with VP19C in the process of capsid assembly.
Thymic Dendritic Cells Are Primary Targets for the Oncogenic Virus SL3-3
Authors: Uittenbogaart Christel H. , Law Wendy , Leenen Pieter J. M. , Bristol Gregory , van Ewijk Willem , Hays Esther F. 
Published: December 2019

ABSTRACTThe murine retrovirus SL3-3 causes malignant transformation of thymocytes and thymic lymphoma in mice of the AKR and NFS strains when they are inoculated neonatally. The objective of the present study was to identify the primary target cells for the virus in the thymuses of these mice. Immunohistochemical studies of the thymus after neonatal inoculation of the SL3-3 virus showed that cells expressing the viral envelope glycoprotein (gp70+cells) were first seen at 2 weeks of age. These virus-expressing cells were found in the cortex and at the corticomedullary junction in both mouse strains. The gp70+cells had the morphology and immunophenotype of dendritic cells. They lacked macrophage-specific antigens. Cell separation studies showed that bright gp70+cells were detected in a fraction enriched for dendritic cells. At 3 weeks of age, macrophages also expressed gp70. At that time, both gp70+dendritic cells and macrophages were found at the corticomedullary junction and in foci in the thymic cortex. At no time during this 3-week period was the virus expressed in cortical and medullary epithelial cells or in thymic lymphoid cells. Infectious cell center assays indicated that cells expressing infectious virus were present in small numbers at 2 weeks after inoculation but increased at 5 weeks of age by several orders of magnitude, indicating virus spread to the thymic lymphoid cells. Thus, at 2 weeks after neonatal inoculation of SL3-3, thymic dendritic cells are the first cells to express the virus. At 3 weeks of age, macrophages also express the virus. In subsequent weeks, the virus spreads to the thymocytes. This pathway of virus expression in the thymus allows the inevitable provirus integration in a thymocyte that results in a clonal lymphoma.
Retrovirus Packaging Cells Expressing the
Mus dunni
Endogenous Virus Envelope Facilitate Transduction of CHO and Primary Hematopoietic Cells
Authors: Wolgamot Greg , Rasko John E. J. , Miller A. Dusty 
Published: December 2019

ABSTRACT

Mus dunni
endogenous virus (MDEV) infects a wide variety of cell types from many different species. To take advantage of this broad host range, we have constructed packaging cells (PD223) that produce virions bearing the MDEV envelope. PD223 cells are able to package Moloney murine leukemia virus-based vectors at a titer of 4 × 10
5
infectious units per ml in the absence of contaminating replication-competent retrovirus. Vectors packaged by PD223 cells are able to transduce CHO cells, which are resistant to transduction by many retroviruses, at ≥25-fold higher efficiency than vectors having other pseudotypes. A vector packaged by PD223 was found to be among the most efficient for transducing primary baboon and human CD34
+
cells.
Vaccine Protection against a Heterologous, Non-Syncytium-Inducing, Primary Human Immunodeficiency Virus
Authors: Robert-Guroff Marjorie , Kaur Harvinder , Patterson L. Jean , Leno Michel , Conley Anthony J. , McKenna Philip M. , Markham Phillip D. , Richardson Ersell , Aldrich Kristine , Arora Kamalpreet , Murty Lalita , Carter Lucretia , Zolla-Pazner Susan , Sinangil Faruk 
Published: December 2019

ABSTRACT

Vaccine-induced protection of chimpanzees against laboratory-adapted and syncytium-inducing, multiply passaged primary human immunodeficiency virus type 1 (HIV-1) isolates, but not against non-syncytium-inducing, minimally passaged ones, has been demonstrated. Following challenge with such an isolate, HIV-1
5016
, we obtained complete protection in one of three chimpanzees previously protected against low- and high-dose HIV-1
SF2
exposures after immunization with an adenovirus-HIV-1
MN
gp160 priming–HIV-1
SF2
gp120 boosting regimen. At challenge, the protected chimpanzee exhibited broad humoral immunity, including neutralizing antibody activity. These results demonstrate the potential of this combination vaccine strategy and suggest that vaccine protection against an HIV isolate relevant to infection of people is feasible.
The African Swine Fever Virus Thymidine Kinase Gene Is Required for Efficient Replication in Swine Macrophages and for Virulence in Swine
Authors: Moore D. M. , Zsak L. , Neilan J. G. , Lu Z. , Rock D. L. 
Published: December 2019

ABSTRACT

African swine fever virus (ASFV) replicates in the cytoplasm of infected cells and contains genes encoding a number of enzymes needed for DNA synthesis, including a thymidine kinase (TK) gene. Recombinant TK gene deletion viruses were produced by using two highly pathogenic isolates of ASFV through homologous recombination with an ASFV p72 promoter–β-glucuronidase indicator cassette (p72GUS) flanked by ASFV sequences targeting the TK region. Attempts to isolate double-crossover TK gene deletion mutants on swine macrophages failed, suggesting a growth deficiency of TK
−
ASFV on macrophages. Two pathogenic ASFV isolates, ASFV Malawi and ASFV Haiti, partially adapted to Vero cells, were used successfully to construct TK deletion viruses on Vero cells. The selected viruses grew well on Vero cells, but both mutants exhibited a growth defect on swine macrophages at low multiplicities of infection (MOI), yielding 0.1 to 1.0% of wild-type levels. At high MOI, the macrophage growth defect was not apparent. The Malawi TK deletion mutant showed reduced virulence for swine, producing transient fevers, lower viremia titers, and reduced mortality. In contrast, 100% mortality was observed for swine inoculated with the TK
+
revertant virus. Swine surviving TK
−
ASFV infection remained free of clinical signs of African swine fever following subsequent challenge with the parental pathogenic ASFV. The data indicate that the TK gene of ASFV is important for growth in swine macrophages in vitro and is a virus virulence factor in swine.
Human Lymphoblastoid CD4
+
T Cells Become Permissive to Macrophage-Tropic Strains of Human Immunodeficiency Virus Type 1 after Passage into Severe Combined Immunodeficient Mice through In Vivo Upregulation of CCR5: In Vivo Dynamics of CD4
+
T-Cell Differentiation in Pathogenesis of AIDS
Authors: Lapenta Caterina , Parlato Stefania , Spada Massimo , Santini Stefano M. , Rizza Paola , Logozzi Mariantonia , Proietti Enrico , Belardelli Filippo , Fais Stefano 
Published: December 2019

ABSTRACT

In this article, we show that passage in SCID mice rendered a human CD4
+
T-cell line (CEM cells) highly susceptible to infection by macrophage-tropic (M-tropic) strains and primary clinical isolates of human immunodeficiency virus type 1 (HIV-1). This in vivo-acquired permissiveness of CEM cells was associated with the induction of a CD45RO
+
phenotype as well as of some β-chemokine receptors. Regulated upon activation, normal T-cell expressed and secreted chemokine entirely inhibited the ability of M-tropic HIV-1 strains to infect these cells. These findings may lead to new approaches in investigating in vivo the capacity of different HIV strains to exploit chemokine receptors in relation to the dynamics of the activation and/or differentiation state of human CD4
+
T cells.
Modulation of Immune System Function by Measles Virus Infection: Role of Soluble Factor and Direct Infection
Authors: Fujinami Robert S. , Sun Xinmin , Howell Joseph M. , Jenkin James C. , Burns James B. 
Published: December 2019

ABSTRACTMeasles virus infection can result in a variety of immunologic defects. We have begun studies to determine the basis for the lack of immune responsiveness to antigen and mitogen following infection. Here we present data showing that Epstein-Barr virus-transformed B-cell lines infected with measles virus produce a soluble factor that can inhibit antigen-specific T-cell proliferation and inhibit the proliferation of uninfected B cells. The soluble factor was neither interleukin-10, transforming growth factor β, nor alpha/beta interferon. B cells infected with measles virus or treated with the soluble factor were unable to present antigen to T cells in a manner that supported antigen-specific proliferation. This could represent one mechanism of how measles virus limits T-cell expansion. However, we found that once CD4+or CD8+T cells were activated, their cytolytic activity was intact whether infected with measles virus or treated with soluble factor. Thus, while slow to be generated these cytoxic cells could participate in viral clearance.
Mouse Mammary Tumor Virus Sequences Responsible for Activating Cellular Oncogenes
Authors: Grimm Sandra L. , Nordeen Steven K. 
Published: December 2019

ABSTRACT

Integration of mouse mammary tumor virus (MMTV) near the
int
genes results in the inappropriate expression of these proto-oncogenes and initiates events that lead to the formation of mammary adenocarcinomas. In most cases, the MMTV provirus integrates in a transcriptional orientation opposite that of the
int
genes. We have used a novel, vector-based system designed to recapitulate the integration of MMTV upstream of the
int-2
promoter. Compared to a cellular promoter or another retroviral promoter, the MMTV long terminal repeat (LTR) in this configuration is particularly efficacious at activating the
int-2
promoter. The sequences responsible for enhancing the activity of the
int-2
promoter map to two domains in the 5′ end of the MMTV LTR. One domain is a previously defined element; the second is an element delineated by these studies that acts synergistically with the first. Both of these elements display mammary cell-specific activity. Thus, even though the MMTV promoter itself is weak without hormonal stimulation, viral integration can position the 5′ LTR elements to efficiently activate transcription from cellular proto-oncogenes. Other functional elements in the LTR have little effect on the activation of the
int-2
promoter. Even stimulation of the MMTV promoter with steroid hormones only modestly activates transcription from the
int-2
promoter, suggesting that the 5′ elements of the LTR are the predominant determinants of the tissue- and orientation-specific activation of cellular promoters by MMTV.
Cytokine Response in Multiple Lymphoid Tissues during the Primary Phase of Feline Immunodeficiency Virus Infection
Authors: Dean Gregg A. , Pedersen Niels C. 
Published: December 2019

ABSTRACTType 1 and 2 cytokine mRNA responses were measured at various time periods and in various lymphoid compartments during the acute stage (first 4 months) of feline immunodeficiency virus (FIV) infection in laboratory cats. Cytokine responses were correlated with virus replication. Virus was detected in plasma and tissue from day 14 postinfection (p.i.) onward, peaked at 56 to 70 days, and declined greatly by 70 days. Virus replication was highest in the thymus, followed by spleen, mesenteric lymph nodes, and cervical lymph nodes. Baseline cytokine levels were highest in the mesenteric lymph nodes and lowest in the cervical lymph nodes. Cytokine upregulation after FIV infection was most dramatic in the cervical lymph nodes, with the greatest increase in interleukin-10 (IL-10) and gamma interferon (IFN-γ). Cytokine transcription in the mesenteric lymph node increased above baseline by day 14 p.i. for IFN-γ, IL-12p40, IL-4, and IL-10, while elevations in the spleen were mainly for IFN-γ, IL-12p40 and IL-10. An increase in IFN-γ, IL-10, and IL-12p40 occurred in the thymus at day 56 p.i., concomitant with the onset of thymitis. In general, type 2 cytokines (IL-4 and IL-10) were increased greater than 1 log over baseline, while the elevations in type 1 cytokines were less than 1 log. In the tissues tested, CD4+cells were the primary source of IL-2, IL-4, and IL-10. Both CD4+and CD8+cells produced IFN-γ, while no cytokine mRNA was detected in B cells. These results demonstrate the presence of a heterogeneous cytokine response in lymphoid tissues during the primary stage of FIV infection. The nature and intensity of the response differed from one compartment to the other and, in the case of the thymus, also with inflammatory changes. Although limited in scope, the present study confirms the usefulness of the FIV infection model in studying early cytokine events that lead to the secondary subclinical carrier state typical of most lentivirus infections.
Control of Human Immunodeficiency Virus Type 1 RNA Metabolism: Role of Splice Sites and Intron Sequences in Unspliced Viral RNA Subcellular Distribution
Authors: Séguin Béatrice , Staffa Alfredo , Cochrane Alan 
Published: December 2019

ABSTRACT

In the course of examining the various factors which affect the metabolism of human immunodeficiency virus type 1 (HIV-1) RNA, we examined the role of intron sequences and splice sites in determining the subcellular distribution of the RNA. Using in situ hybridization, we demonstrated that in the absence of Rev, unspliced RNA generated with an HIV-1
env
expression construct displayed discrete localization in the nucleus, coincident with the location of the gene and not associated with SC35-containing nuclear speckles. Expression of Rev resulted in a disperse signal for the unspliced RNA throughout both the nucleus and the cytoplasm. Subsequent fractionation of the nucleus revealed that the majority of unspliced viral RNA within the nucleus is associated with the nuclear matrix and that upon expression of Rev, a small proportion of the unspliced RNA is found within the nucleoplasm. Mutations which altered splice site utilization did not alter the sequestration of unspliced RNA into discrete nuclear regions. In contrast, a 2.2-kb deletion of intron sequence resulted in a shift from discrete regions within the nucleus to a disperse signal throughout the cell, indicating that intron sequences, and not just splice sites, are required for the observed nuclear sequestration of unspliced viral RNA.
Immunization with a Single Major Histocompatibility Complex Class I-Restricted Cytotoxic T-Lymphocyte Recognition Epitope of Herpes Simplex Virus Type 2 Confers Protective Immunity
Authors: Blaney Joseph E. , Nobusawa Eri , Brehm Michael A. , Bonneau Robert H. , Mylin Lawrence M. , Fu Tong-Ming , Kawaoka Yoshihiro , Tevethia Satvir S. 
Published: December 2019

ABSTRACTWe have evaluated the potential of conferring protective immunity to herpes simplex virus type 2 (HSV-2) by selectively inducing an HSV-specific CD8+cytotoxic T-lymphocyte (CTL) response directed against a single major histocompatibility complex class I-restricted CTL recognition epitope. We generated a recombinant vaccinia virus (rVV-ES-gB498-505) which expresses the H-2Kb-restricted, HSV-1/2-cross-reactive CTL recognition epitope, HSV glycoprotein B residues 498 to 505 (SSIEFARL) (gB498-505), fused to the adenovirus type 5 E3/19K endoplasmic reticulum insertion sequence (ES). Mucosal immunization of C57BL/6 mice with this recombinant vaccinia virus induced both a primary CTL response in the draining lymph nodes and a splenic memory CTL response directed against HSV gB498-505. To determine the ability of the gB498-505-specific memory CTL response to provide protection from HSV infection, immunized mice were challenged with a lethal dose of HSV-2 strain 186 by the intranasal (i.n.) route. Development of the gB498-505-specific CTL response conferred resistance in 60 to 75% of mice challenged with a lethal dose of HSV-2 and significantly reduced the levels of infectious virus in the brains and trigeminal ganglia of challenged mice. Finally, i.n. immunization of C57BL/6 mice with either a recombinant influenza virus or a recombinant vaccinia virus expressing HSV gB498-505 without the ES was also demonstrated to induce an HSV-specific CTL response and provide protection from HSV infection. This finding confirms that the induction of an HSV-specific CTL response directed against a single epitope is sufficient for conferring protective immunity to HSV. Our findings support the role of CD8+T cells in the control of HSV infection of the central nervous system and suggest the potential importance of eliciting HSV-specific mucosal CD8+CTL in HSV vaccine design.
A
cis
Repression Sequence Adjacent to the Transcription Start Site of the Human Cytomegalovirus US3 Gene Is Required To Down Regulate Gene Expression at Early and Late Times after Infection
Authors: Lashmit Philip E. , Stinski Mark F. , Murphy Eain A. , Bullock Grant C. 
Published: December 2019

ABSTRACT

Human cytomegalovirus has two enhancer-containing immediate-early (IE) promoters with a
cis
repression sequence (CRS) positioned immediately upstream of the transcription start site, designated the major IE (MIE) promoter and the US3 promoter. The role of the CRS upstream of the US3 transcription start site in the context of the viral genome was determined by comparing the levels of transcription from these two enhancer-containing promoters in recombinant viruses with a wild-type or mutant CRS. Upstream of the CRS of the US3 promoter was either the endogenous enhancer (R
2
) or silencer (R
1
). The downstream US3 gene was replaced with the indicator gene chloramphenicol acetyltransferase (CAT). Infected permissive human fibroblast cells or nonpermissive, undifferentiated monocytic THP-1 cells were analyzed for expression from the US3 promoter containing either the wild-type or mutant CRS. With the wild-type CRS, the maximum level of transcription in permissive cells was detected within 4 to 6 h after infection and then declined. With the mutant CRS and the R
2
enhancer upstream, expression from the US3 promoter continued to increase throughout the viral replication cycle to levels 20- to 40-fold higher than for the wild type. In nonpermissive or permissive monocytic THP-1 cells, expression from the US3 promoter was also significantly higher when the CRS was mutated. Less expression was obtained when only the R
1
element was present, but expression was higher when the CRS was mutated. Thus, the CRS in the enhancer-containing US3 promoter appears to allow for a short burst of US3 gene expression followed by repression at early and late times after infection. Overexpression of US3 may be detrimental to viral replication, and its level of expression must be stringently controlled. The role of the CRS and the viral IE86 protein in controlling enhancer-containing promoters is discussed.
Targeting of a Short Peptide Derived from the Cytoplasmic Tail of the G1 Membrane Glycoprotein of Uukuniemi Virus (
Bunyaviridae
) to the Golgi Complex
Authors: Andersson Agneta M. , Pettersson Ralf F. 
Published: December 2019

ABSTRACT

Members of the
Bunyaviridae
family acquire an envelope by budding through the lipid bilayer of the Golgi complex. The budding compartment is thought to be determined by the accumulation of the two heterodimeric membrane glycoproteins G1 and G2 in the Golgi. We recently mapped the retention signal for Golgi localization in one
Bunyaviridae
member (Uukuniemi virus) to the cytoplasmic tail of G1. We now show that a myc-tagged 81-residue G1 tail peptide expressed in BHK21 cells is efficiently targeted to the Golgi complex and retained there during a 3-h chase. Green-fluorescence protein tagged at either end with this peptide or with a C-terminally truncated 60-residue G1 tail peptide was also efficiently targeted to the Golgi. The 81-residue peptide colocalized with mannosidase II (a medial Golgi marker) and partially with p58 (an intermediate compartment marker) and TGN38 (a
trans
-Golgi marker). In addition, the 81-residue tail peptide induced the formation of brefeldin A-resistant vacuoles that did not costain with markers for other membrane compartments. Removal of the first 10 N-terminal residues had no effect on the Golgi localization but abolished the vacuolar staining. The shortest peptide still able to become targeted to the Golgi encompassed residues 10 to 40. Subcellular fractionation showed that the 81-residue tail peptide was associated with microsomal membranes. Removal of the two palmitylation sites from the tail peptide did not affect Golgi localization and had only a minor effect on the association with microsomal membranes. Taken together, the results provide strong evidence that Golgi retention of the heterodimeric G1-G2 spike protein complex of Uukuniemi virus is mediated by a short region in the cytoplasmic tail of the G1 glycoprotein.
Interaction of Poly(rC) Binding Protein 2 with the 5′ Noncoding Region of Hepatitis A Virus RNA and Its Effects on Translation
Authors: Graff Judith , Cha John , Blyn Lawrence B. , Ehrenfeld Ellie 
Published: December 2019

ABSTRACT
Utilization of internal ribosome entry segment (IRES) structures in the 5′ noncoding region (5′NCR) of picornavirus RNAs for initiation of translation requires a number of host cell factors whose distribution may vary in different cells and whose requirement may vary for different picornaviruses. We have examined the requirement of the cellular protein poly(rC) binding protein 2 (PCBP2) for hepatitis A virus (HAV) RNA translation. PCBP2 has recently been identified as a factor required for translation and replication of poliovirus (PV) RNA. PCBP2 was shown to be present in FRhK-4 cells, which are permissive for growth of HAV, as it is in HeLa cells, which support translation of HAV RNA but which have not been reported to host replication of the virus. Competition RNA mobility shift assays showed that the 5′NCR of HAV RNA competed for binding of PCBP2 with a probe representing stem-loop IV of the PV 5′NCR. The binding site on HAV RNA was mapped to nucleotides 1 to 157, which includes a pyrimidine-rich sequence. HeLa cell extracts that had been depleted of PCBP2 by passage over a PV stem-loop IV RNA affinity column supported only low levels of HAV RNA translation. Translation activity was restored upon addition of recombinant PCBP2 to the depleted extract. Removal of the 5′-terminal 138 nucleotides of the HAV RNA, or removal of the entire IRES, eliminated the dependence of HAV RNA translation on PCBP2.