bluuebunny/crossref_metadata_2025_split huggingface dataset:
Measles Virus Spread between Neurons Requires Cell Contact but Not CD46 Expression, Syncytium Formation, or Extracellular Virus Production
Authors: Lawrence Diane M. P. , Patterson Catherine E. , Gales Tracy L. , D'Orazio Joseph L. , Vaughn Melinda M. , Rall Glenn F. 
Published: July 2002

ABSTRACT

In patients with subacute sclerosing panencephalitis (SSPE), which is associated with persistent measles virus (MV) infection in the brain, little infectious virus can be recovered despite the presence of viral RNA and protein. Based on studies of brain tissue from SSPE patients and our work with MV-infected NSE-CD46
+
mice, which express the measles receptor CD46 on neurons, several lines of evidence suggest that the mechanism of viral spread in the central nervous system differs from that in nonneuronal cells. To examine this alternate mechanism of viral spread, as well as the basis for the loss of normal transmission mechanisms, infection and spread of MV Edmonston was evaluated in primary CD46
+
neurons from transgenic mice and differentiated human NT2 neurons. As expected, unlike that between fibroblasts, viral spread between neurons occurred in the absence of syncytium formation and with minimal extracellular virus. Electron microscopy analysis showed that viral budding did not occur from the neuronal surface, although nucleocapsids were present in the cytoplasm and aligned at the cell membrane. We observed many examples of nucleocapsids present in the neuronal processes and aligned at presynaptic neuronal membranes. Cocultures of CD46
+
and CD46
−
neurons showed that cell contact but not CD46 expression is required for MV spread between neurons. Collectively, these results suggest that the neuronal environment prevents the normal mechanisms of MV spread between neurons at the level of viral assembly but allows an alternate, CD46-independent mechanism of viral transmission, possibly through the synapse.
Interaction of the Cauliflower Mosaic Virus Coat Protein with the Pregenomic RNA Leader
Authors: Guerra-Peraza Orlene , de Tapia Marc , Hohn Thomas , Hemmings-Mieszczak Maja 
Published: July 2002

ABSTRACTUsing the yeast three-hybrid system, the interaction of theCauliflower mosaic virus(CaMV) pregenomic 35S RNA (pgRNA) leader with the viral coat protein, its precursor, and a series of derivatives was studied. The purine-rich domain in the center of the pgRNA leader was found to specifically interact with the coat protein. The zinc finger motif of the coat protein and the preceding basic domain were essential for this interaction. Removal of the N-terminal portion of the basic domain led to loss of specificity but did not affect the strength of the interaction. Mutations of the zinc finger motif abolished not only the interaction with the RNA but also viral infectivity. In the presence of the very acidic C-terminal domain, which is part of the preprotein but is not present in the mature CP, the interaction with the RNA was undetectable.
Genetic Evidence for an Interaction between Human Immunodeficiency Virus Type 1 Matrix and α-Helix 2 of the gp41 Cytoplasmic Tail
Authors: Murakami Tsutomu , Freed Eric O. 
Published: July 2002

ABSTRACTThe incorporation of envelope (Env) glycoproteins into virions is an essential step in the retroviral replication cycle. Lentiviruses, including human immunodeficiency virus type 1 (HIV-1), encode Env glycoproteins with unusually long cytoplasmic tails, the functions of which have not been fully elucidated. In this study, we examine the effects on virus replication of a number of mutations in a helical motif (α-helix 2) located near the center of the HIV-1 gp41 cytoplasmic tail. We find that, in T-cell lines, small deletions in this domain disrupt the incorporation of Env glycoproteins into virions and markedly impair virus infectivity. Through the analysis of viral revertants, we demonstrate that a single amino acid change (34VI) in the matrix domain of Gag reverses the Env incorporation and infectivity defect imposed by a small deletion near the C terminus of α-helix 2. These results provide genetic evidence, in the context of infected T cells, for an interaction between HIV-1 matrix and the gp41 cytoplasmic tail and identify domains of both proteins involved in this putative interaction.
Interaction between the Open Reading Frame III Product and the Coat Protein Is Required for Transmission of Cauliflower Mosaic Virus by Aphids
Authors: Leh Véronique , Jacquot Emmanuel , Geldreich Angèle , Haas Muriel , Blanc Stéphane , Keller Mario , Yot Pierre 
Published: July 2002

ABSTRACT
Transmission of cauliflower mosaic virus (CaMV) by aphids requires two viral nonstructural proteins, the open reading frame (ORF) II and ORF III products (P2 and P3). An interaction between a C-terminal domain of P2 and an N-terminal domain of P3 is essential for transmission. Purified particles of CaMV are efficiently transmitted only if aphids, previously fed a P2-containing solution, are allowed to acquire a preincubated mixture of P3 and virions in a second feed, thus suggesting a direct interaction between P3 and coat protein. Herein we demonstrate that P3 directly interacts with purified viral particles and unassembled coat protein without the need for any other factor and that P3 mediates the association of P2 with purified virus particles. The interaction domain of P3 is located in its C-terminal half, downstream of the P3-P2 interaction domain but overlapping a region which binds nucleic acids. Mutagenesis of P3 which interferes with the interaction between P3 and virions is correlated with the loss of transmission by aphids. Taken together, our results demonstrate that P3 plays a crucial role in the formation of the CaMV transmissible complex by serving as a bridge between P2 and virus particles.
Alteration of Zinc-Binding Residues of Simian Immunodeficiency Virus p8
NC
Results in Subtle Differences in Gag Processing and Virion Maturation Associated with Degradative Loss of Mutant NC
Authors: Yovandich Jason L. , Chertova Elena N. , Kane Brad P. , Gagliardi Tracy D. , Bess Julian W. , Sowder Raymond C. , Henderson Louis E. , Gorelick Robert J. 
Published: July 2002

ABSTRACT

In all retroviruses analyzed to date (except for the spumaretroviruses), the Zn
2+
-coordinating residues of nucleocapsid (NC) perform or assist in crucial reactions necessary to complete the retrovirus life cycle. Six replication-defective mutations have been engineered in the two NC Zn
2+
fingers (ZFs) of simian immunodeficiency virus [SIV(Mne)] that change or delete specific Zn
2+
-interacting Cys residues and were studied by using electron microscopy, reversed-phase high-performance liquid chromatography, immunoblotting, and RNA quantification. We focused on phenotypes of produced particles, specifically morphology, Gag polyprotein processing, and genomic RNA packaging. Phenotypes were similar among viruses containing a point or deletion mutation involving the same ZF. Mutations in the proximal ZF (ZF1) resulted in near-normal Gag processing and full-length genomic RNA incorporation and were most similar to wild-type (WT) virions with electron-dense, conical cores. Mutation of the distal ZF, as well as point mutations in both ZFs, resulted in more unprocessed Gag proteins than a deletion or point mutation in ZF1, with an approximate 30% reduction in levels of full-length genomic RNA in virions. These mutant virions contained condensed cores; however, the cores typically appeared less electron dense and more rod shaped than WT virions. Surprisingly, deletion of both ZFs, including the basic linker region between the ZFs, resulted in the most efficient Gag processing. However, genomic RNA packaging was ∼10% of WT levels, and those particles produced were highly abnormal with respect to size and core morphology. Surprisingly, all NC mutations analyzed demonstrated a significant loss of processed NC in virus particles, suggesting that Zn
2+
-coordinated NC is protected from excessive proteolytic cleavage. Together, these results indicate that Zn
2+
coordination is important for correct Gag precursor processing and NC protein stability. Additionally, SIV particle morphology appears to be the result of proper and complete Gag processing and relies less on full-length genomic RNA incorporation, as dictated by the Zn
2+
coordination in the ZFs of the NC protein.
The Ability of Integrin αvβ3To Function as a Receptor for Foot-and-Mouth Disease Virus Is Not Dependent on the Presence of Complete Subunit Cytoplasmic Domains
Authors: Neff Sherry , Baxt Barry 
Published: July 2002

ABSTRACTThe integrin αvβ3has been shown to function as one of the integrin receptors on cultured cells for foot-and-mouth disease virus (FMDV), and high-efficiency utilization of the bovine homolog of this integrin is dependent on the cysteine-rich repeat region of the bovine β3subunit. In this study we have examined the role of the cytoplasmic domains of the αvand β3subunits in FMDV infection. We have found that truncations or extensions of these domains of either subunit, including deletions removing almost all of the cytoplasmic domains, had little or no effect on the ability of the integrin to function as a receptor for FMDV. The lysosomotropic agent monensin inhibited viral replication in cells transfected with either intact or cytoplasmic domain-truncated αvβ3. In addition, viral replication in transfected cells was inhibited by an αvβ3function-blocking antibody but not by function-blocking antibodies to three other RGD-directed integrins, suggesting that these integrins are not involved in the infectious process. These results indicate that alterations to the cytoplasmic domains of either subunit, which lead to the inability of the integrin receptor to function normally, do not abolish the ability of the integrin to bind and internalize this viral ligand.
Residue 627 of PB2 Is a Determinant of Cold Sensitivity in RNA Replication of Avian Influenza Viruses
Authors: Massin Pascale , van der Werf Sylvie , Naffakh Nadia 
Published: July 2002

ABSTRACT
Human influenza A viruses replicate in the upper respiratory tract at a temperature of about 33°C, whereas avian viruses replicate in the intestinal tract at a temperature close to 41°C. In the present study, we analyzed the influence of low temperature (33°C) on RNA replication of avian and human viruses in cultured cells. The kinetics of replication of the NP segment were similar at 33 and 37°C for the human A/Puerto-Rico/8/34 and A/Sydney/5/97 viruses, whereas replication was delayed at 33°C compared to 37°C for the avian A/FPV/Rostock/34 and A/Mallard/NY/6750/78 viruses. Making use of a genetic system for the in vivo reconstitution of functional ribonucleoproteins, we observed that the polymerase complexes derived from avian viruses but not human viruses exhibited cold sensitivity in mammalian cells, which was determined mostly by residue 627 of PB2. Our results suggest that a reduced ability of the polymerase complex of avian viruses to ensure replication of the viral genome at 33°C could contribute to their inability to grow efficiently in humans.
Genetic Diversity in RNA Virus Quasispecies Is Controlled by Host-Virus Interactions
Authors: Schneider William L. , Roossinck Marilyn J. 
Published: July 2002

ABSTRACT

Many RNA viruses have genetically diverse populations known as quasispecies. Important biological characteristics may be related to the levels of diversity in the quasispecies (quasispecies cloud size), including adaptability and host range. Previous work using
Tobacco mosaic virus
and
Cucumber mosaic virus
indicated that evolutionarily related viruses have very different levels of diversity in a common host. The quasispecies cloud size for these viruses remained constant throughout serial passages. Inoculation of these viruses on a number of hosts demonstrated that quasispecies cloud size is not constant for these viruses but appears to be dependent on the host. The quasispecies cloud size remained constant as long as the viruses were maintained on a given host. Shifting the virus between hosts resulted in a change in cloud size to levels associated with the new host. Quasispecies cloud size for these viruses is related to host-virus interactions, and understanding these interactions may facilitate the prediction and prevention of emerging viral diseases.
Evolutionary Relationships and Systematics of the Alphaviruses
Authors: Powers Ann M. , Brault Aaron C. , Shirako Yukio , Strauss Ellen G. , Kang WenLi , Strauss James H. , Weaver Scott C. 
Published: July 2002

ABSTRACT

Partial E1 envelope glycoprotein gene sequences and complete structural polyprotein sequences were used to compare divergence and construct phylogenetic trees for the genus
Alphavirus
. Tree topologies indicated that the mosquito-borne alphaviruses could have arisen in either the Old or the New World, with at least two transoceanic introductions to account for their current distribution. The time frame for alphavirus diversification could not be estimated because maximum-likelihood analyses indicated that the nucleotide substitution rate varies considerably across sites within the genome. While most trees showed evolutionary relationships consistent with current antigenic complexes and species, several changes to the current classification are proposed. The recently identified fish alphaviruses salmon pancreas disease virus and sleeping disease virus appear to be variants or subtypes of a new alphavirus species. Southern elephant seal virus is also a new alphavirus distantly related to all of the others analyzed. Tonate virus and Venezuelan equine encephalitis virus strain 78V3531 also appear to be distinct alphavirus species based on genetic, antigenic, and ecological criteria. Trocara virus, isolated from mosquitoes in Brazil and Peru, also represents a new species and probably a new alphavirus complex.
Interaction of Zyxin, a Focal Adhesion Protein, with the E6 Protein from Human Papillomavirus Type 6 Results in Its Nuclear Translocation
Authors: Degenhardt Yan Yan , Silverstein Saul 
Published: July 2002

ABSTRACT

Zyxin, a focal adhesion molecule, interacts specifically with the E6 protein from human papillomavirus (HPV) type 6 in a yeast two-hybrid screen of a cDNA library prepared from human keratinocytes. Zyxin does not interact significantly with E6 proteins from HPV types 11, 16, or 18. The interaction was confirmed by in vitro and in vivo analyses and it requires the LIM domains (Lin-11, Isl-1, and Mec-3 [G. Freyd, S. K. Kim, and H. R. Horvitz, Nature 344:876–879, 1990]) found at the carboxyl terminus of zyxin. Cotransfection of E6 from HPV (
6
E6) and zyxin results in the accumulation of zyxin in the nucleus where it can function as a transcriptional activator.
6
E6 can also mobilize endogenous zyxin to the nucleus.
Molecular Evolution of Puumala Hantavirus
Authors: Sironen Tarja , Vaheri Antti , Plyusnin Alexander 
Published: July 2002

ABSTRACT

Puumala virus (PUUV) is a negative-stranded RNA virus in the genus
Hantavirus
, family
Bunyaviridae.
In this study, detailed phylogenetic analysis was performed on 42 complete S segment sequences of PUUV originated from several European countries, Russia, and Japan, the largest set available thus far for hantaviruses. The results show that PUUV sequences form seven distinct and well-supported genetic lineages; within these lineages, geographical clustering of genetic variants is observed. The overall phylogeny of PUUV is star-like, suggesting an early split of genetic lineages. The individual PUUV lineages appear to be independent, with the only exception to this being the Finnish and the Russian lineages that are closely connected to each other. Two strains of PUUV-like virus from Japan form the most ancestral lineage diverging from PUUV. Recombination points within the S segment were searched for and evidence for intralineage recombination events was seen in the Finnish, Russian, Danish, and Belgian lineages of PUUV. Molecular clock analysis showed that PUUV is a stable virus, evolving slowly at a rate of 0.7 × 10
−7
to 2.2 × 10
−6
nt substitutions per site per year.
Nuclear Covalently Closed Circular Viral Genomic DNA in the Liver of Hepatocyte Nuclear Factor 1α-Null Hepatitis B Virus Transgenic Mice
Authors: Raney Anneke K. , Eggers Carrie M. , Kline Eric F. , Guidotti Luca G. , Pontoglio Marco , Yaniv Moshe , McLachlan Alan 
Published: July 2002

ABSTRACT
The role of hepatocyte nuclear factor 1α (HNF1α) in the regulation of hepatitis B virus (HBV) transcription and replication in vivo was investigated using a HNF1α-null HBV transgenic mouse model. HBV transcription was not measurably affected by the absence of the HNF1α transcription factor. However, intracellular viral replication intermediates were increased two- to fourfold in mice lacking functional HNF1α protein. The increase in encapsidated cytoplasmic replication intermediates in HNF1α-null HBV transgenic mice was associated with the appearance of nonencapsidated nuclear covalently closed circular (CCC) viral genomic DNA. Viral CCC DNA was not readily detected in HNF1α-expressing HBV transgenic mice. This indicates the synthesis of nuclear HBV CCC DNA, the proposed viral transcriptional template found in natural infection, is regulated either by subtle alterations in the levels of viral transcripts or by changes in the physiological state of the hepatocyte in this in vivo model of HBV replication.
Human Immunodeficiency Virus Type 1 Central DNA Flap: Dynamic Terminal Product of Plus-Strand Displacement DNA Synthesis Catalyzed by Reverse Transcriptase Assisted by Nucleocapsid Protein
Authors: Hameau Laurence , Jeusset Josette , Lafosse Sophie , Coulaud Dominique , Delain Etienne , Unge Torsten , Restle Tobias , Le Cam Eric , Mirambeau Gilles 
Published: July 2002

ABSTRACT
To terminate the reverse transcription of the human immunodeficiency virus type 1 (HIV-1) genome, a final step occurs within the center of the proviral DNA generating a 99-nucleotide DNA flap (6). This step, catalyzed by reverse transcriptase (RT), is defined as a discrete strand displacement (SD) synthesis between the first nucleotide after the central priming (cPPT) site and the final position of the central termination sequence (CTS) site. Using recombinant HIV-1 RT and a circular single-stranded DNA template harboring the cPPT-CTS sequence, we have developed an SD synthesis-directed in vitro termination assay. Elongation, strand displacement, and complete central flap behavior were analyzed using electrophoresis and electron microscopy approaches. Optimal conditions to obtain complete central flap, which ended at the CTS site, have been defined in using nucleocapsid protein (NCp), the main accessory protein of the reverse transcription complex. A full-length HIV-1 central DNA flap was then carried out in vitro. Its synthesis appears faster in the presence of the HIV-1 NCp or the T4-encoded SSB protein (gp32). Finally, a high frequency of strand transfer was shown during the SD synthesis along the cPPT-CTS site with RT alone. This reveals a local and efficient 3′-5′ branch migration which emphasizes some important structural fluctuations within the flap. These fluctuations may be stabilized by the NCp chaperone activity. The biological implications of the RT-directed NCp-assisted flap synthesis are discussed within the context of reverse transcription complexes, assembly of the preintegration complexes, and nuclear import of the HIV-1 proviral DNA to the nucleus toward their chromatin targets.
Frequencies of Virus-Specific CD4+and CD8+T Lymphocytes Secreting Gamma Interferon after Acute Natural Rotavirus Infection in Children and Adults
Authors: Jaimes María C. , Rojas Olga Lucía , González Ana María , Cajiao Isabela , Charpilienne Annie , Pothier Pierre , Kohli Evelyne , Greenberg Harry B. , Franco Manuel A. , Angel Juana 
Published: July 2002

ABSTRACTHuman rotavirus-specific CD4+and CD8+T-cell responses in peripheral blood lymphocytes were studied using a flow cytometric assay that detects the intracellular accumulation of cytokines after short-term in vitro antigen stimulation. The frequencies of virus-specific T cells that secrete gamma interferon and interleukin-13 (IL-13) were determined in adults and children during the acute or convalescent phase of rotavirus-induced diarrhea, in asymptomatically infected adults and laboratory workers who worked with human stool samples containing rotavirus, and in healthy adults. Significantly higher frequencies of rotavirus-specific interferon gamma-secreting CD8+and CD4+T cells, but not IL-13-secreting T cells, were detected in symptomatically infected adults and exposed laboratory workers than in healthy adults and children with acute rotavirus diarrhea. The levels of rotavirus-specific T cells returned to levels found in healthy adults by 32 days after the onset of rotavirus diarrhea in most adult subjects. Children with rotavirus diarrhea had undetectable or very low levels of CD4+and CD8+T cells that secrete gamma interferon. Adult cytomegalovirus-seropositive individuals had frequencies of cytomegalovirus-specific T cells that secrete gamma interferon that were approximately 20 times the level of rotavirus-specific T cells. This result suggests that rotavirus is a relatively poor inducer of circulating memory T cells that secrete gamma interferon. The frequencies of gamma interferon-secreting CD4+and CD8+T cells and the frequencies of IL-13-secreting CD4+T cells responding to the T-cell superantigen staphylococcal enterotoxin B (SEB) were lower in children than in adults. In both adults and children, the frequencies of CD4+cells secreting gamma interferon in response to SEB were higher than the frequencies of cells secreting IL-13.
Mutations in the Yellow Fever Virus Nonstructural Protein NS2A Selectively Block Production of Infectious Particles
Authors: Kümmerer Beate M. , Rice Charles M. 
Published: July 2002

ABSTRACT

Little is known about the function of flavivirus nonstructural protein NS2A. Two forms of NS2A are found in yellow fever virus-infected cells. Full-length NS2A (224 amino acids) is the product of cleavage at the NS1/2A and NS2A/2B sites. NS2Aα, a C-terminally truncated form of 190 amino acids, results from partial cleavage by the viral NS2B-3 serine protease at the sequence QK↓T within NS2A. Exchange of serine for lysine at this site (QKT→QST) blocks the production of both NS2Aα and infectious virus. The present study reveals that this defect is not at the level of RNA replication. Despite normal structural region processing, infectious particles containing genome RNA and capsid protein were not released from cells transfected with the mutant RNA. Nevertheless, production of subviral prM/M- and E-containing particles was unimpaired. The NS2A defect could be complemented in
trans
by providing NS1-2A or NS1-2Aα. However,
trans
complementation was not observed when the C-terminal lysine of NS1-2Aα was replaced with serine. In addition to true reversions, NS2Aα cleavage site mutations could be suppressed by two classes of second-site changes. The first class consisted of insertions at the NS2Aα cleavage site that restored its basic character and cleavability. A second class of suppressors occurred in the NS3 helicase domain, in which NS3 aspartate 343 was replaced with an uncharged residue (either valine, alanine, or glycine). These mutations in NS3 restored infectious-virus production in the absence of cleavage at the mutant NS2Aα site. Taken together, our results reveal an unexpected role for NS2A and NS3 in the assembly and/or release of infectious flavivirus particles.