scholarly journals Changes in the Length of the Neuraminidase Stalk Region Impact H7N9 Virulence in Mice

2015 ◽  
Vol 90 (4) ◽  
pp. 2142-2149 ◽  
Author(s):  
Yuhai Bi ◽  
Haixia Xiao ◽  
Quanjiao Chen ◽  
Yan Wu ◽  
Lifeng Fu ◽  
...  
Keyword(s):  
Influenza Virus ◽  
Amino Acid ◽  
Full Length ◽  
Virus Infectivity ◽  
Biological Functions ◽  
Amino Acid Deletion ◽  
H7n9 Influenza ◽  
Stalk Length

The neuraminidase stalk of the newly emerged H7N9 influenza virus possesses a 5-amino-acid deletion. This study focuses on characterizing the biological functions of H7N9 with varied neuraminidase stalk lengths. Results indicate that the 5-amino-acid deletion had no impact on virus infectivity or replicationin vitroorin vivocompared to that of a virus with a full-length stalk, but enhanced virulence in mice was observed for H7N9 encoding a 19- to 20-amino-acid deletion, suggesting that N9 stalk length impacts virulence in mammals, as N1 stalk length does.

scholarly journals Antiviral activity of plant juices and green tea against SARS-CoV-2 and influenza virus in vitro

2020 ◽  
Author(s):  
Bruno Frank ◽  
Carina Conzelmann ◽  
Tatjana Weil ◽  
Rüdiger Groß ◽  
Peggy Jungke ◽  
...  
Keyword(s):  
Influenza Virus ◽  
Antiviral Activity ◽  
Green Tea ◽  
Human Adenovirus ◽  
Virus Infectivity ◽  
Virucidal Activity ◽  
Black Chokeberry ◽  
Chokeberry Juice

AbstractMany plant juices, extracts and teas have been shown to possess antiviral activity. We here analyzed the virucidal activity of black chokeberry (Aronia melanocarpa), pomegranate (Punica granatum), and elderberry (Sambucus nigra) juice, as well as green tea (Camellia sinensis) against different respiratory viruses. We found that all tested plant derived products effectively inactivated influenza virus, whereas only chokeberry juice diminished SARS-CoV-2 and vaccinia virus infectivity. None of the products inactivated non-enveloped human adenovirus type 5. Thus, black chokeberry juice exerts virucidal activity against different enveloped viral pathogens under in vitro conditions. Whether application of virucidal juices or green tea as oral rinses may lower viral loads in the oral cavity in vivo remains to be evaluated.

scholarly journals The LINC00961 transcript and its encoded micropeptide, small regulatory polypeptide of amino acid response, regulate endothelial cell function

2020 ◽  
Vol 116 (12) ◽  
pp. 1981-1994 ◽  
Author(s):  
Helen L Spencer ◽  
Rachel Sanders ◽  
Mounia Boulberdaa ◽  
Marco Meloni ◽  
Amy Cochrane ◽  
...  
Keyword(s):  
Endothelial Cell ◽  
Amino Acid ◽  
Full Length ◽  
Binding Partners ◽  
Full Length Transcript ◽  
Opposing Effects ◽  
Amino Acid Response

Abstract Aims Long non-coding RNAs (lncRNAs) play functional roles in physiology and disease, yet understanding of their contribution to endothelial cell (EC) function is incomplete. We identified lncRNAs regulated during EC differentiation and investigated the role of LINC00961 and its encoded micropeptide, small regulatory polypeptide of amino acid response (SPAAR), in EC function. Methods and results Deep sequencing of human embryonic stem cell differentiation to ECs was combined with Encyclopedia of DNA Elements (ENCODE) RNA-seq data from vascular cells, identifying 278 endothelial enriched genes, including 6 lncRNAs. Expression of LINC00961, first annotated as an lncRNA but reassigned as a protein-coding gene for the SPAAR micropeptide, was increased during the differentiation and was EC enriched. LINC00961 transcript depletion significantly reduced EC adhesion, tube formation, migration, proliferation, and barrier integrity in primary ECs. Overexpression of the SPAAR open reading frame increased tubule formation; however, overexpression of the full-length transcript did not, despite production of SPAAR. Furthermore, overexpression of an ATG mutant of the full-length transcript reduced network formation, suggesting a bona fide non-coding RNA function of the transcript with opposing effects to SPAAR. As the LINC00961 locus is conserved in mouse, we generated an LINC00961 locus knockout (KO) mouse that underwent hind limb ischaemia (HLI) to investigate the angiogenic role of this locus in vivo. In agreement with in vitro data, KO animals had a reduced capillary density in the ischaemic adductor muscle after 7 days. Finally, to characterize LINC00961 and SPAAR independent functions in ECs, we performed pull-downs of both molecules and identified protein-binding partners. LINC00961 RNA binds the G-actin sequestering protein thymosin beta-4x (Tβ4) and Tβ4 depletion phenocopied the overexpression of the ATG mutant. SPAAR binding partners included the actin-binding protein, SYNE1. Conclusion The LINC00961 locus regulates EC function in vitro and in vivo. The gene produces two molecules with opposing effects on angiogenesis: SPAAR and LINC00961.

scholarly journals Cross-Protection of Chicken Immunoglobulin Y Antibodies against H5N1 and H1N1 Viruses Passively Administered in Mice

2011 ◽  
Vol 18 (7) ◽  
pp. 1083-1090 ◽  
Author(s):  
Michael G. Wallach ◽  
Richard J. Webby ◽  
Fakhrul Islam ◽  
Stephen Walkden-Brown ◽  
Eva Emmoth ◽  
...  
Keyword(s):  
Influenza Virus ◽  
Influenza Pandemic ◽  
Influenza Virus Infection ◽  
Influenza Viruses ◽  
Antigenic Drift ◽  
Virus Infectivity ◽  
Lethal Challenge ◽  
H5n1 Influenza

ABSTRACTInfluenza viruses remain a major threat to global health due to their ability to undergo change through antigenic drift and antigenic shift. We postulated that avian IgY antibodies represent a low-cost, effective, and well-tolerated approach that can easily be scaled up to produce enormous quantities of protective antibodies. These IgY antibodies can be administered passively in humans (orally and intranasally) and can be used quickly and safely to help in the fight against an influenza pandemic. In this study, we raised IgY antibodies against H1N1, H3N2, and H5N1 influenza viruses. We demonstrated that, using whole inactivated viruses alone and in combination to immunize hens, we were able to induce a high level of anti-influenza virus IgY in the sera and eggs, which lasted for at least 2 months after two immunizations. Furthermore, we found that by use ofin vitroassays to test for the ability of IgY to inhibit hemagglutination (HI test) and virus infectivity (serum neutralization test), IgYs inhibited the homologous as well as in some cases heterologous clades and strains of viruses. Using anin vivomouse model system, we found that, when administered intranasally 1 h prior to infection, IgY to H5N1 protected 100% of the mice against lethal challenge with H5N1. Of particular interest was the finding that IgY to H5N1 cross-protected against A/Puerto Rico/8/34 (H1N1) bothin vitroandin vivo. Based on our results, we conclude that anti-influenza virus IgY can be used to help prevent influenza virus infection.

A 113-amino-acid truncation at the NS1 C-terminus is a determinant for viral replication of H5N6 avian influenza virus in vitro and in vivo

2018 ◽  
Vol 225 ◽  
pp. 6-16 ◽  
Author(s):  
Xiaole Cui ◽  
Yanhong Ji ◽  
Zhengxiang Wang ◽  
Yingying Du ◽  
Haoran Guo ◽  
...  
Keyword(s):  
Influenza Virus ◽  
Amino Acid ◽  
Avian Influenza ◽  
Viral Replication ◽  
Avian Influenza Virus ◽  
C Terminus

scholarly journals Influence of N-Linked Glycosylation of Porcine Reproductive and Respiratory Syndrome Virus GP5 on Virus Infectivity, Antigenicity, and Ability To Induce Neutralizing Antibodies

2006 ◽  
Vol 80 (8) ◽  
pp. 3994-4004 ◽  
Author(s):  
Israrul H. Ansari ◽  
Byungjoon Kwon ◽  
Fernando A. Osorio ◽  
Asit K. Pattnaik
Keyword(s):  
Amino Acid ◽  
Neutralizing Antibodies ◽  
Protective Efficacy ◽  
Neutralizing Antibody ◽  
Respiratory Syndrome Virus ◽  
Virus Infectivity ◽  
Neutralization Epitope ◽  
Enhanced Sensitivity

ABSTRACT Porcine reproductive and respiratory syndrome virus (PRRSV) glycoprotein 5 (GP5) is the most abundant envelope glycoprotein and a major inducer of neutralizing antibodies in vivo. Three putative N-linked glycosylation sites (N34, N44, and N51) are located on the GP5 ectodomain, where a major neutralization epitope also exists. To determine which of these putative sites are used for glycosylation and the role of the glycan moieties in the neutralizing antibody response, we generated a panel of GP5 mutants containing amino acid substitutions at these sites. Biochemical studies with expressed wild-type (wt) and mutant proteins revealed that the mature GP5 contains high-mannose-type sugar moieties at all three sites. These mutations were subsequently incorporated into a full-length cDNA clone. Our data demonstrate that mutations involving residue N44 did not result in infectious progeny production, indicating that N44 is the most critical amino acid residue for infectivity. Viruses carrying mutations at N34, N51, and N34/51 grew to lower titers than the wt PRRSV. In serum neutralization assays, the mutant viruses exhibited enhanced sensitivity to neutralization by wt PRRSV-specific antibodies. Furthermore, inoculation of pigs with the mutant viruses induced significantly higher levels of neutralizing antibodies against the mutant as well as the wt PRRSV, suggesting that the loss of glycan residues in the ectodomain of GP5 enhances both the sensitivity of these viruses to in vitro neutralization and the immunogenicity of the nearby neutralization epitope. These results should have great significance for development of PRRSV vaccines of enhanced protective efficacy.

scholarly journals Identification of Amino Acid Residues of Influenza Virus Nucleoprotein Essential for RNA Binding

1999 ◽  
Vol 73 (9) ◽  
pp. 7357-7367 ◽  
Author(s):  
Debra Elton ◽  
Liz Medcalf ◽  
Konrad Bishop ◽  
Deborah Harrison ◽  
Paul Digard
Keyword(s):  
Influenza Virus ◽  
Amino Acid ◽  
Electrostatic Interactions ◽  
Rna Binding ◽  
Mutational Analysis ◽  
High Affinity ◽  
Arginine Residues ◽  
Rna Interaction

ABSTRACT The influenza virus nucleoprotein (NP) is a single-strand-RNA-binding protein associated with genome and antigenome RNA and is one of the four virus proteins necessary for transcription and replication of viral RNA. To better characterize the mechanism by which NP binds RNA, we undertook a physical and mutational analysis of the polypeptide, with the strategy of identifying first the regions in direct contact with RNA, then the classes of amino acids involved, and finally the crucial residues by mutagenesis. Chemical fragmentation and amino acid sequencing of NP that had been UV cross linked to radiolabelled RNA showed that protein-RNA contacts occur throughout the length of the polypeptide. Chemical modification experiments implicated arginine but not lysine residues as important for RNA binding, while RNA-dependent changes in the intrinsic fluorescence spectrum of NP suggested the involvement of tryptophan residues. Supporting these observations, single-codon mutagenesis identified five tryptophan, one phenylalanine, and two arginine residues as essential for high-affinity RNA binding at physiological temperature. In addition, mutants unable to bind RNA in vitro were unable to support virus gene expression in vivo. The mutationally sensitive residues are not localized to any particular region of NP but instead are distributed throughout the protein. Overall, these data are inconsistent with previous models suggesting that the NP-RNA interaction is mediated by a discrete N-terminal domain. Instead, we propose that high-affinity binding of RNA by NP requires the concerted interaction of multiple regions of the protein with RNA and that the individual protein-RNA contacts are mediated by a combination of electrostatic interactions between positively charged residues and the phosphate backbone and planar interactions between aromatic side chains and bases.

scholarly journals Attenuation and Immunogenicity of a Live High Pathogenic PRRSV Vaccine Candidate with a 32-Amino Acid Deletion in the nsp2 Protein

2014 ◽  
Vol 2014 ◽  
pp. 1-11 ◽  
Author(s):  
Wenhui Lu ◽  
Baoli Sun ◽  
Jianyue Mo ◽  
Xiduo Zeng ◽  
Guanqun Zhang ◽  
...  
Keyword(s):  
Amino Acid ◽  
Vaccine Candidate ◽  
Clinical Signs ◽  
Respiratory Syndrome Virus ◽  
Amino Acid Deletion ◽  
Lethal Challenge ◽  
Nsp2 Gene ◽  
Amino Acid Mutations

A porcine reproductive and respiratory syndrome virus (PRRSV) QY1 was serially passed on Marc-145 cells. Virulence of different intermediate derivatives of QY1 (P5, P60, P80, and P100) were determined. The study found that QY1 had been gradually attenuated during the in vitro process. Pathogenicity study showed that pigs inoculated with QY1 P100 and P80 did not develop any significant PRRS clinic symptoms. However, mild-to-moderate clinical signs and acute HP-PRRSV symptoms of infection were observed in pigs inoculated with QY1 P60 and P5, respectively. Furthermore, we determined the whole genome sequences of these four intermediate viruses. The results showed that after 100 passages, compared to QY1 P5, a total of 32 amino acid mutations were found. Moreover, there were one nucleotide deletion and a unique 34-amino acid deletion found at 5′UTR and in nsp2 gene during the attenuation process, respectively. Such deletions were genetically stable in vivo. Following PRRSV experimental challenge, pigs inoculated with a single dose of QY1 P100 developed no significant clinic symptoms and well tolerated lethal challenge, while QY1 P80 group still developed mild fever in the clinic trial after challenge. Thus, we concluded that QY1 P100 was a promising and highly attenuated PRRSV vaccine candidate.

scholarly journals The Lck Binding Domain of Herpesvirus Saimiri Tip-484 Constitutively Activates Lck and STAT3 in T Cells

1999 ◽  
Vol 73 (2) ◽  
pp. 1689-1694 ◽  
Author(s):  
Troy C. Lund ◽  
Paul C. Prator ◽  
Maria M. Medveczky ◽  
Peter G. Medveczky
Keyword(s):  
T Cells ◽  
Amino Acid ◽  
Cell Transformation ◽  
Binding Domain ◽  
Full Length ◽  
Herpesvirus Saimiri ◽  
Dependent Manner ◽  
Interacting Protein

ABSTRACT Constitutive activation of signal transducers and activators of transcription (STATs) has been associated with oncogenesis. Previously, a protein required for T-cell transformation by the DNA tumor virus herpesvirus saimiri (HVS) strain 484, designated tyrosine kinase-interacting protein (Tip-484), was shown to interact with and dramatically upregulate the activity of the STATs in an Lck-dependent manner. The minimal region of Tip-484 responsible for binding Lck was defined as a 10-residue C-terminal Src-related kinase homology domain, an 18-amino-acid spacer, and a 10-residue potential SH3 binding domain. This region is termed the LBD (for Lck binding domain). The present data show that only the LBD of Tip-484 is needed to activate Lck in vitro and in vivo. Finally, the LBD was shown to form a complex with STAT3 in vitro, and expression of the LBD in T cells led to STAT3 activation equal to that of full-length Tip-484. These studies demonstrate that the 48-amino-acid LBD of Tip-484 can perform as effectively as the full-length protein in vitro and in vivo.

scholarly journals Functional characterization of the 180-kD ribosome receptor in vivo.

1995 ◽  
Vol 130 (1) ◽  
pp. 29-39 ◽  
Author(s):  
E E Wanker ◽  
Y Sun ◽  
A J Savitz ◽  
D I Meyer
Keyword(s):  
Amino Acid ◽  
Binding Domain ◽  
Full Length ◽  
Cell Fractionation ◽  
Ribosome Binding ◽  
Basic Domain ◽  
Biochemical Analyses ◽  
Er Membrane

A cDNA encoding the 180-kD canine ribosome receptor (RRp) was cloned and sequenced. The deduced primary structure indicates three distinct domains: an NH2-terminal stretch of 28 uncharged amino acids representing the membrane anchor, a basic region (pI = 10.74) comprising the remainder of the NH2-terminal half and an acidic COOH-terminal half (pI = 4.99). The most striking feature of the amino acid sequence is a 10-amino acid consensus motif, NQGKKAEGAP, repeated 54 times in tandem without interruption in the NH2-terminal positively charged region. We postulate that this repeated sequence represents a ribosome binding domain which mediates the interaction between the ribosome and the ER membrane. To substantiate this hypothesis, recombinant full-length ribosome receptor and two truncated versions of this protein, one lacking the potential ribosome binding domain, and one lacking the COOH terminus, were expressed in Saccharomyces cerevisiae. Morphological and biochemical analyses showed all proteins were targeted to, and oriented correctly in the ER membrane. In vitro ribosome binding assays demonstrated that yeast microsomes containing the full-length canine receptor or one lacking the COOH-terminal domain were able to bind two to four times as many human ribosomes as control membranes lacking a recombinant protein or microsomes containing a receptor lacking the NH2-terminal basic domain. Electron micrographs of these cells revealed that the expression of all receptor constructs led to a proliferation of perinuclear ER membranes known as "karmellae." Strikingly, in those strains which expressed cDNAs encoding a receptor containing the putative ribosome binding domain, the induced ER membranes (examined in situ) were richly studded with ribosomes. In contrast, karmellae resulting from the expression of receptor cDNA lacking the putative ribosome binding domain were uniformly smooth and free of ribosomes. Cell fractionation and biochemical analyses corroborated the morphological characterization. Taken together these data provide further evidence that RRp functions as a ribosome receptor in vitro, provide new evidence indicating its functionality in vivo, and in both cases indicate that the NH2-terminal basic domain is essential for ribosome binding.

scholarly journals The C Terminus of the Histone Chaperone Asf1 Cross-Links to Histone H3 in Yeast and Promotes Interaction with Histones H3 and H4

2012 ◽  
Vol 33 (3) ◽  
pp. 605-621 ◽  
Author(s):  
Briana K. Dennehey ◽  
Seth Noone ◽  
Wallace H. Liu ◽  
Luke Smith ◽  
Mair E. A. Churchill ◽  
...  
Keyword(s):  
Amino Acid ◽  
Phosphorylation Site ◽  
Histone H3 ◽  
Unnatural Amino Acid ◽  
Full Length ◽  
Content Type ◽  
C Terminus ◽  
Cross Links

ABSTRACTThe central histone H3/H4 chaperone Asf1 comprises a highly conserved globular core and a divergent C-terminal tail. While the function and structure of the Asf1 core are well known, the function of the tail is less well understood. Here, we have explored the role of the yeast (yAsf1) and human (hAsf1a and hAsf1b) Asf1 tails inSaccharomyces cerevisiae. We show, using a photoreactive, unnatural amino acid, that Asf1 tail residue 210 cross-links to histone H3in vivoand, further, that loss of C-terminal tail residues 211 to 279 weakens yAsf1-histone binding affinityin vitronearly 200-fold. Via several yAsf1 C-terminal truncations and yeast-human chimeric proteins, we found that truncations at residue 210 increase transcriptional silencing and that the hAsf1a tail partially substitutes for full-length yAsf1 with respect to silencing but that full-length hAsf1b is a better overall substitute for full-length yAsf1. In addition, we show that the C-terminal tail of Asf1 is phosphorylated at T270 in yeast. Loss of this phosphorylation site does not prevent coimmunoprecipitation of yAsf1 and Rad53 from yeast extracts, whereas amino acid residue substitutions at the Asf1-histone H3/H4 interface do. Finally, we show that residue substitutions in yAsf1 near the CAF-1/HIRA interface also influence yAsf1's function in silencing.

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