Top PDF Viral Genome-Linked Protein (VPg) Is Essential for Translation Initiation of Rabbit Hemorrhagic Disease Virus (RHDV).

Viral Genome-Linked Protein (VPg) Is Essential for Translation Initiation of Rabbit Hemorrhagic Disease Virus (RHDV).

Viral Genome-Linked Protein (VPg) Is Essential for Translation Initiation of Rabbit Hemorrhagic Disease Virus (RHDV).

Rabbit hemorrhagic disease virus (RHDV), the causative agent of rabbit hemorrhagic dis- ease, is an important member of the caliciviridae family. Currently, no suitable tissue culture system is available for proliferating RHDV, limiting the study of the pathogenesis of RHDV. In addition, the mechanisms underlying RHDV translation and replication are largely unknown compared with other caliciviridae viruses. The RHDV replicon recently con- structed in our laboratory provides an appropriate model to study the pathogenesis of RHDV without in vitro RHDV propagation and culture. Using this RHDV replicon, we dem- onstrated that the viral genome-linked protein (VPg) is essential for RHDV translation in RK-13 cells for the first time. In addition, we showed that VPg interacts with eukaryotic initia- tion factor 4E (eIF4E) in vivo and in vitro and that eIF4E silencing inhibits RHDV translation, suggesting the interaction between VPg and eIF4E is involved in RHDV translation. Our results support the hypothesis that VPg serves as a novel cap substitute during the initiation of RHDV translation.
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Análise comparativa de parâmetros bioquímicos e fisiológicos de um genótipo de feijãodecorda (Vigna unguiculata L. Walp.) suscetível e seu mutante derivado, resistente, infectados com o vírus do mosaico seo do caupi (CPSMV)

Análise comparativa de parâmetros bioquímicos e fisiológicos de um genótipo de feijãodecorda (Vigna unguiculata L. Walp.) suscetível e seu mutante derivado, resistente, infectados com o vírus do mosaico seo do caupi (CPSMV)

suggested that PTI is important for Arabidopsis resistance to viruses because they found evidence that viral MAMPs are directly or indirectly recognized by PRR which initiates a signaling cascade that culminates with PTI [42]. A third possibility, is that the mechanism of plant resistance results from the lack of interactions between plant and viral factors required for triggering translation of viral proteins. The replication of viral nucleic acid due to the absence or mutation of the proper host factor that leads to an incompatible relationship in which no or very limited virus replication occurs in the plant that shows no symptoms or extremely limited necrosis [43]. For example, various studies have shown that effective recessive resistance of various plants to Potyvirus resides in the inability of VPg (Viral Genome-linked Protein) to interact with the eukaryotic translation initiation factor eIF4E or eIF(iso)4E of the plant due to mutations in the genes that code for these proteins [44,45]. This incapacity of interaction occurs as the result of any replacement of single or multiple amino acid residues exposed on the surface of the eukaryotic translation initiation factor proteins regardless whether in the domain of interaction with 5' 7-methylguanosine cap (7- methyl-GTP cap) or not [46]. In most cases, the result is the inhibition of multiplication of viral material. Some studies showed that replacement of only one amino acid residue in eIF4E is sufficient to confer resistance, as in the case of replacing alanine with proline residue in lettuce, or valine with glutamine residue in pepper [47,48]. In other cases, the mechanism of the resistance breaking resulted from the change in more than one amino acid residues, as in common bean (Phaseolus vulgaris) against the Potyvirus BCMV (Bean Common Mosaic Virus genus Potyvirus) [49]. Although, the majority of cases of recessive resistance is conferred by mutations of genes coding for the translation initiation factors eIF4E and eIF(iso)4E [17,45,50], as well as in both affecting their interactions with VPg, mutations in other components of the family eIF4F [eIF4G and eIF(iso)4G], also affect the replication in some species of virus [51-54].
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The X-linked tumor suppressor TSPX interacts and promotes degradation of the hepatitis B viral protein HBx via the proteasome pathway.

The X-linked tumor suppressor TSPX interacts and promotes degradation of the hepatitis B viral protein HBx via the proteasome pathway.

[23,40]. HBx has been shown to promote androgen receptor transactivation, while its expression is enhanced by androgen receptor via a couple of androgen response elements located within the enhancer region of HBV-genome [23]. Our study, however, shows that an X-linked tumor suppressor is capable of promoting HBx degradation. TSPX is ubiquitously expressed in normal tissues including liver [3], but its expression is markedly down regulated in primary tumors and human tumor cell lines [14]. Since male has one X chromosome, any loss-of-function mutation(s) and/or epigenetic dysregulation of X-linked tumor suppressors could render males without the corresponding oncogenic protection, thereby promoting carcinogenesis in male- preferential manner(s) [41]. Hence, genetic/epigenetic inactiva- tion of TSPX gene could closely associate with the HBV-mediated HCC development. Further, since pro-oncogenic TSPY expres- sion has been documented in selected HCC specimens [8], it could exert a male-specific effect(s) on the overall complex etiology for HBV-associated HCC in addition to inactivation mutation of its X-homologue, TSPX. Further studies on the roles of TSPX and TSPY in hepatocarcinogenesis could shed critical insights on the HBV-mediated pathologic process(es), and could lead to develop- ments of genderized strategies for the prevention, diagnosis and treatments of HBV-associated liver cancer.
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Bruna Prati, Bruna Marangoni, Enrique Boccardo

Bruna Prati, Bruna Marangoni, Enrique Boccardo

Many viruses, especially those with oncogenic properties, express proteins that affect the cell cycle and DNA damage repair regulatory pathways. Several studies have shown that during genome amplification, HPV proteins interact with different components of the cellular DNA repair machinery to activate or downregulate the expression or activity of factors of the ATM and ATR pathways. In 2009, Moody and coworkers showed that ATM activation is required for the productive genomic replication of HPV31 but not for episomal maintenance (20). On the other hand, it was obser- ved that CHK1 inhibition causes an important reduction in the number of HPV episomes in differentiated cells (21). In addition, the knockdown of DNA topoisomerase 2-binding protein 1 (TopBP1), a protein that acts upstream of ATR, sup- presses HPV31 replication (22). In line with these observa- tions, it has been reported that E2 protein from HPV16 interacts with TopBP1 and that this interaction improves E2- mediated viral transcription and replication (23). Moreover, ATM and ATR kinases are constitutively activated in HPV- positive keratinocytes, and ATR/CHK1 blockade is asso- ciated with the downregulation of HPV productive replica- tion and the reduced expression of late genes (22).
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Protein-RNA linkage and posttranslational modifications of feline calicivirus and murine norovirus VPg proteins

Protein-RNA linkage and posttranslational modifications of feline calicivirus and murine norovirus VPg proteins

Members of the Caliciviridae family of positive sense RNA viruses cause a wide range of diseases in both humans and animals. The detailed characterization of the calicivirus life cycle had been hampered due to the lack of robust cell culture systems and experimental tools for many of the members of the family. However, a number of caliciviruses replicate efficiently in cell culture and have robust reverse genetics systems available, most notably feline calicivirus (FCV) and murine norovirus (MNV). These are therefore widely used as representative members with which to examine the mechanistic details of calicivirus genome translation and replication. The replication of the calicivirus RNA genome occurs via a double-stranded RNA intermediate that is then used as a template for the production of new positive sense viral RNA, which is covalently linked to the virus-encoded protein VPg. The covalent linkage to VPg occurs during genome replication via the nucleotidylylation activity of the viral RNA-dependent RNA polymerase. Using FCV and MNV, we used mass spectrometry-based approach to identify the specific amino acid linked to the 5′ end of the viral nucleic acid. We observed that both VPg proteins are covalently linked to guanosine diphosphate (GDP) moieties via tyrosine positions 24 and 26 for FCV and MNV respectively. These data fit with previous observations indicating that mutations introduced into these specific amino acids are deleterious for viral replication and fail to produce infectious virus. In addition, we also detected serine phosphorylation sites within the FCV VPg protein with positions 80 and 107 found consistently phosphorylated on VPg-linked viral RNA isolated from infected cells. This work provides the first direct experimental characterization of the linkage of infectious calicivirus viral RNA to the VPg protein and highlights that post-translational modifications of VPg may also occur during the viral life cycle.
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REVIEW THE INFLUENCE OF THE HUMAN GENOME ON CHRONIC VIRAL HEPATITIS OUTCOME

REVIEW THE INFLUENCE OF THE HUMAN GENOME ON CHRONIC VIRAL HEPATITIS OUTCOME

Progressive hepatic fibrosis and cirrhosis develop in 20% to 30% of patients with chronic hepatitis C. Although this phenomenon is linked to HCV presence, it does not occur through HCV influence alone but also depends on collagen production by Ito cells or hepatic stellate cells (HSCs). These HSCs are situated between the endothelial lining and the hepatocytes (Disse’s space), a site consistent with their role in the recruitment of leukocytes and their targeting of damaged hepatocytes. During the inflammatory phenomenon that occurs in hepatitis C, Kupffer cells produce large quantities of secreted cytokines such as TNF-α and Interferon-gamma. Cytokine secretion into the hepatic microenvironment contributes to the activation of HSCs. The HSCs thus abandon their quiescent state and begin storing vitamin D, as well as reaching an actively proliferating myofibroblast-like cell state. Furthermore, HSC activation is characterized by differential gene expression of connective tissue components, matrix degrading enzymes, and their inhibitors, resulting in matrix accumulation. Activated HSC produces several collagen types. Anti-inflammatory cytokines, such as interleukin-10 (IL-10), which presents prominent antifibrotic activity through downregulation of collagen 1 expression, are also produced by HSCs 80 .
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Viral proteins acquired from a host converge to simplified domain architectures.

Viral proteins acquired from a host converge to simplified domain architectures.

The infection cycle of viruses creates many opportunities for the exchange of genetic material with the host. Many viruses integrate their sequences into the genome of their host for replication. These processes may lead to the virus acquisition of host sequences. Such sequences are prone to accumulation of mutations and deletions. However, in rare instances, sequences acquired from a host become beneficial for the virus. We searched for unexpected sequence similarity among the 900,000 viral proteins and all proteins from cellular organisms. Here, we focus on viruses that infect metazoa. The high- conservation analysis yielded 187 instances of highly similar viral-host sequences. Only a small number of them represent viruses that hijacked host sequences. The low-conservation sequence analysis utilizes the Pfam family collection. About 5% of the 12,000 statistical models archived in Pfam are composed of viral-metazoan proteins. In about half of Pfam families, we provide indirect support for the directionality from the host to the virus. The other families are either wrongly annotated or reflect an extensive sequence exchange between the viruses and their hosts. In about 75% of cross-taxa Pfam families, the viral proteins are significantly shorter than their metazoan counterparts. The tendency for shorter viral proteins relative to their related host proteins accounts for the acquisition of only a fragment of the host gene, the elimination of an internal domain and shortening of the linkers between domains. We conclude that, along viral evolution, the host-originated sequences accommodate simplified domain compositions. We postulate that the trimmed proteins act by interfering with the fundamental function of the host including intracellular signaling, post-translational modification, protein-protein interaction networks and cellular trafficking. We compiled a collection of hijacked protein sequences. These sequences are attractive targets for manipulation of viral infection.
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A search for RNA insertions and NS3 gene duplication in the genome of cytopathic isolates of bovine viral diarrhea virus

A search for RNA insertions and NS3 gene duplication in the genome of cytopathic isolates of bovine viral diarrhea virus

Calves born persistently infected with non-cytopathic bovine viral diarrhea virus (ncpBVDV) frequently develop a fatal gastroenteric illness called mucosal disease. Both the original virus (ncpBVDV) and an antigenically identical but cytopathic virus (cpBVDV) can be isolated from animals affected by mucosal disease. Cytopathic BVDVs originate from their ncp counterparts by diverse genetic mechanisms, all leading to the expression of the non-structural polypeptide NS3 as a discrete protein. In contrast, ncpBVDVs express only the large precursor polypeptide, NS2-3, which contains the NS3 sequence within its carboxy-terminal half. We report here the investigation of the mechanism leading to NS3 expression in 41 cpBVDV isolates. An RT-PCR strategy was employed to detect RNA insertions within the NS2-3 gene and/or duplication of the NS3 gene, two common mechan- isms of NS3 expression. RT-PCR amplification revealed insertions in the NS2-3 gene of three cp isolates, with the inserts being similar in size to that present in the cpBVDV NADL strain. Sequencing of one such insert revealed a 296-nucleotide sequence with a central core of 270 nucleotides coding for an amino acid sequence highly homolo- gous (98%) to the NADL insert, a sequence corresponding to part of the cellular J-Domain gene. One cpBVDV isolate contained a duplica- tion of the NS3 gene downstream from the original locus. In contrast, no detectable NS2-3 insertions or NS3 gene duplications were ob- served in the genome of 37 cp isolates. These results demonstrate that processing of NS2-3 without bulk mRNA insertions or NS3 gene duplications seems to be a frequent mechanism leading to NS3 expression and BVDV cytopathology.
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Determining host metabolic limitations on viral replication via integrated modeling and experimental perturbation.

Determining host metabolic limitations on viral replication via integrated modeling and experimental perturbation.

Figure S3 Infected host fluxes on succinate M9 minimal media. (A) Flux dynamics are displayed for a subset of the metabolic network map. Arrows representing reactions and the subplots of flux through those reactions are colored according to clustering of flux dynamics. Positive flux values correspond to the reaction direction indicated by the colored arrowhead, negative flux direction is depicted with light grey barbs. Asterisks (*) represent an abbrevi- ation of the arrow for uptake from media. Metabolite abbreviations are consistent with FBA model definition. For clustering, fluxes were treated as vectors with (1-correlation) as distance, and clustered using average hierarchical grouping with a cutoff height of 0.25. clusters with fewer than ten members appear in black, and clusters with constant dynamics are highlighted in grey. All nonzero fluxes in any media (tryptone, glucose, succinate, and acetate) were included in the flux clustering so that cluster designation and color coding is consistent across media and figures. (B) Select flux dynamics expanded for clarity ordered to exemplify host flux changes driven by viral dynamics: (i) host amino acid synthesis, (ii) major viral capsid protein synthesis, (iii) host nucleotide phosphor- ylation, (iv) viral digestion of host genome to dNMPs, (v) purine biosynthesis, (vi) viral mRNA synthesis, (vii) viral genome synthesis, (viii) host cell envelope biosynthesis, (ix) host biomass accumulation. (EPS)
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A bacteriophage-related chimeric marine virus infecting abalone.

A bacteriophage-related chimeric marine virus infecting abalone.

Marine viruses are unarguably abundant and diverse members of microbial communities in the marine biosphere [1,2]. They have major roles in the oceanic carbon and energy cycles and cause diseases in a wide range of organisms [3–5]. In late 2005, an outbreak of abalone fatal disease, referred to as abalone shriveling syndrome (AbSS), spread to several fisheries in Fuzhou, China. Abalones in all life-stages can suffer from the disease. The infection was characterized by pedal muscle atrophy and lesion of mantle tissue with nigrescence (Figure S1). Accompanied by a reduction in feeding, most infected abalones fall from the reef and die. These signs were similar to withering syndrome in the black abalone, previously reported to be associated with Rickettsiales bacteria [6]. Herein, we prove from experimental infection of healthy abalone that an unclassified, novel virus can cause AbSS, named as AbSS- associated Virus (AbSV). In infected abalone, the cytopathic changes (e.g. necrosis etc.) were observed and accompanied with the modifications of host macromolecules (i.e. hemocyanins and ferritins). The complete viral genomic sequence reveals that putative genes in the virus are linked to bacteriophages, eukaryotic viruses, bacteria and endosymbionts. In addition, the virus’s mosaic genome presents a gene organization similar to the majority of tailed phages.
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Host immune responses to a viral immune modulating protein: immunogenicity of viral interleukin-10 in rhesus cytomegalovirus-infected rhesus macaques.

Host immune responses to a viral immune modulating protein: immunogenicity of viral interleukin-10 in rhesus cytomegalovirus-infected rhesus macaques.

HCMV immune modulating proteins that disrupt host cell functions, including antigen presentation, signaling, trafficking, activation, and metabolism, have not been evaluated as vaccine candidates. A large percentage of the coding capacity of the HCMV genome is devoted to these proteins, and in vitro studies strongly suggest that HCMV immune modulating proteins collectively could skew host immunity during critical junctures of HCMV infection in vivo. Limited studies have observed cellular immune responses specific to presumptive HCMV immune modulating proteins, including those that disrupt MHC class I antigen presentation (US2, US3, US6, US11), and those that inhibit NK function (UL16, UL18) [8,9]. Antibody responses have also been detected against UL111A (cmvIL-10) in some HCMV- infected individuals [10]. One group profiling HCMV T cell responses noted that, based on the in vitro phenotype of the immune modulating proteins and their potential role in vivo, ‘‘Future investigations will need to assess the levels of protection afforded by responses directed toward these epitopes’’ [8].
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The central repeat domain 1 of Kaposi's sarcoma-associated herpesvirus (KSHV) latency associated-nuclear antigen 1 (LANA1) prevents cis MHC class I peptide presentation

The central repeat domain 1 of Kaposi's sarcoma-associated herpesvirus (KSHV) latency associated-nuclear antigen 1 (LANA1) prevents cis MHC class I peptide presentation

Here we show that the LANA1 CR1 domain is primarily responsible for in cis prevention of MHC I presentation of LANA1 peptides. Unlike EBV EBNA1 (Apcher et al., 2009), this effect can be physically separated from the translation retardation and proteasome inhibition domains, suggesting that neither is critical for LANA1 evasion of antigen presentation. Surprisingly, CR1's immune evasion mechanism appears to be more complex than simple inhibition of proteasomal processing. The CR2 domain expressed as an isolated fragment markedly retards cis proteasomal processing yet its contribution to inhibition of MHC I peptide presentation is minimal compared to CR1. Since MHC I presentation inhibition occurs in cis and yet proteasomal processing is not markedly inhibited by CR1, one possibility is that CR1 domain decouples proteasomal processing of LANA1 from the ER translocation machinery. Immunoproteasomal processing is tightly linked to the surface of the ER membrane so that peptides generated by processing are efficiently translocated by the transporter associated with antigen presentation 1 (TAP1) (Brooks et al., 2000). Herpes simplex virus ICP47 binds to TAP1 and blocks transport of viral peptides into the ER (Fruh et al., 1995; Hill et al., 1995; York et al., 1994). Cytomegalovirus also blocks peptide transport by producing a protein, US6, that blocks TAP1 (Hengel et al., 1997; Lehner et al., 1997). When we bypass this machinery using an ER signal peptide to translocate LANA1 directly into the ER, both LANA1 and LANA1 lacking a CR1 domain are efficiently presented to MHC I. LANA1 expressed into the lumen of the ER might be processed by ER aminopeptidase associated with antigen processing (ERAAP) (Blanchard et al., 2010) or by retrotranslocation and proteasomal processing of LANA1 protein, either of which may be TAP-dependent. This raises the possibility that CR1 delocalizes the active immuno- proteasome from the TAP machinery so that peptides generated in cis are not efficiently translocated into the ER. LANA1 does not affect expression of TAP1 or immunoproteasome components, LMP2 or LMP7 (data not shown), consistent with CR1 not disrupting the
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Hepatitis B virus polymerase blocks pattern recognition receptor signaling via interaction with DDX3: implications for immune evasion.

Hepatitis B virus polymerase blocks pattern recognition receptor signaling via interaction with DDX3: implications for immune evasion.

DEAD-box RNA helicases constitute a large family of proteins that comprises at least 38 members in human genome [28]. DEAD-box RNA helicases exhibit multiple roles in diverse aspects of RNA metabolism such as transcription, pre-mRNA splicing, RNA export, translation, and RNA decay [29]. Not surprisingly, DDX3 has also been implicated in multiple distinct cellular processes as well. First, DDX3 has been reported to act as a transcriptional factor in the nucleus [30]. Secondly, DDX3 was shown to bind to eIF4E, a translation initiation factor with cap- binding properties, effectively suppressing translation [31]. Finally, DDX3 has been implicated in various viral life cycles. For instance, DDX3 was shown to be essential for nuclear export of human immunodeficiency virus-1 (HIV-1) RNA through the Rev/ RRE pathway [32]. In addition, DDX3 supports viral replication of hepatitis C virus (HCV) genome via its interaction with the HCV core protein [33,34]. In contrast, DDX3 was shown to inhibit HBV genome replication via its interaction with HBV Pol [11]. Besides its roles in RNA metabolism, a novel function relevant to innate immunity was recently reported by two groups [9,10]. Although the augmentation of IRF signaling by DDX3 was found independently by two groups, discrepancies have been noted regarding the specific mechanism for DDX3-mediated IRF3 activation. Specifically, Bowie and colleagues [9] demonstrated that the IKKe-DDX3 interaction is significantly enhanced upon SenV infection with concomitant IRF3 phosphorylation, indicat- ing that DDX3 stimulates the protein kinase activity of IKKe. In contrast, Decker and colleagues [10] concluded that DDX3 acts as a transcription factor of the IFNb promoter, which is in agreement with the transcriptional role of DDX3 reported by another study [30,35]. Interestingly, we have demonstrated that HBV Pol disrupts the IKKe-DDX3 interaction (Fig. 6C), which is consistent with the conclusions of Bowie and colleagues (7). However, in- depth analyses are needed to clarify this mechanistic issue.
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The interface between methyltransferase and polymerase of NS5 is essential for flavivirus replication.

The interface between methyltransferase and polymerase of NS5 is essential for flavivirus replication.

We first performed a systematic mutagenesis analysis of the six key residues (P113, L115, W121, F467, F351 and P585) in hydrophobic network for its biological function study using an infectious clone of JEV (Fig. 1A). The substitutions of each of these six hydrophobic residues with Arginine (R), Aspartic acid (D) and Serine (S) were designed to alter or disrupt the hydrophobic network. D and S represent amino acids with charged and uncharged polar side chains, while the side chain of Figure 2. Functional analysis of hydrophobic network mutations in NS5 interface in a JEV infectious cDNA clone. (A) Immunofluorescence analysis and plaque morphology of JEV genome-length viral RNA replication containing hydrophobic network mutations in transfected BHK-21 cells at the indicated time points. Monoclonal antibody against SLEV envelope protein and Texas Red-conjugated goat anti- mouse IgG were used as primary and secondary antibodies, respectively. The supernatants collected at 120 hpt were assayed for plaque morphology analysis by double-layer plaque assay as described in Materials and Methods. (B) The supernatants were collected at each time point post transfection, and subjected to monolayer plaque assay for measurement of virus production. The visible plaques were used to calculate titers of JEV WT and mutants.
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Genome-wide association for mapping QTLs linked to protein and oil contents in soybean

Genome-wide association for mapping QTLs linked to protein and oil contents in soybean

Abstract – The objective of this work was to identify single-nucleotide polymorphism (SNP) markers linked with quantitative trait loci (QTLs) associated with increased contents of protein and oil in soybean. A total of 169 Brazilian soybean varieties, genotyped with 6,000 SNP markers, were evaluated. Protein and oil contents were obtained with the near-infrared reflectance method. Correlation and multiple linear regression analyses were used to identify linkage disequilibrium between SNP markers and the QTLs associated with the two characteristics. Seven QTLs were found to be associated with protein content, on six chromosomes (2, 6, 11, 12, 13, and 16), explaining 60.9% of the variation in this trait. For oil content, eight QTLs were identified on six chromosomes (1, 4, 5, 6, 17, and 19), explaining 78.3% of the variation in the trait. The correlation between the number of loci containing favorable alleles and the evaluated characteristics was 0.49 for protein content and 0.60 for oil content. The molecular markers identified are mapped in genomic regions containing QTLs previously mapped for both characteristics, which reinforces the association between these regions and the genetic control of oil and protein contents in soybean.
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An isoform of eukaryotic initiation factor 4E from Chrysanthemum morifolium interacts with Chrysanthemum virus B coat protein.

An isoform of eukaryotic initiation factor 4E from Chrysanthemum morifolium interacts with Chrysanthemum virus B coat protein.

The mRNAs of many plant RNA viruses lack a cap structure, a poly (A) tail or both, yet they efficiently compete with host mRNAs for the translational machinery. Several host proteins that interact with viral coat proteins are involved in the cell-to-cell movement or subcellular localization of viruses [30–32]. Furthermore, a report suggested that translation of Alfalfa mosaic virus (AMV) genomic RNA is enhanced by binding of several coat protein molecules to its 3 9 end, apparently mimicking the function of poly(A)-binding protein (PABP) [33]. GST pull-down revealed that AMV coat protein interacts with eIF4F and eIF(iso)4F from wheat germ [34]. In the genera Alfamovirus and Ilarvirus, initiation of infection by viruses requires the addition of coat protein (CP) to a mixture of the genomic RNAs [35–37]. Chrysanthemum is ranked among the top ten most important flower crops in the international cut-flower market [38]. Several viruses and viroids have been reported in chrysanthemum, and Chrysanthemum virus B (CVB, genus Carlavirus, family Flexiviridae) is one of the major pathogens of chrysanthemum [39]. The Carlavirus genome is a positive, single-strand RNA with a 5 9-cap and 39-poly(A) structure that contains six open reading frames (ORFs), of which ORF5 encodes the coat protein [40]. In this study, we report the cloning of a full-length cDNA encoding eIF(iso)4E from chrysanthemum. We used the cloned cDNA for expression profiling of the CmeIF(iso)4E gene. Finally, we demonstrated the interaction of CVBCP with CmeIF(iso)4E by yeast two-hybrid assay and BiFC (Bimolecular Fluorescence Complementation) in onion epidermal cells, and that CVBCP increased the RLU of Luc-CVB through Luminescence assay, suggesting CVBCP might participate in the translation of viral proteins.
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REVIEW MOLECULAR ASPECTS OF HEPATIC CARCINOGENESIS

REVIEW MOLECULAR ASPECTS OF HEPATIC CARCINOGENESIS

only HBV-derived regulatory sequences that express all of the viral gene products and replicate the virus in the hepatocyte. Therefore, they are considered to be excellent models for dissecting the human disease. One potential problem with this methodology is that the expression of HBV genes and the process of viral replication are not per se cytotoxic for the hepatocytes. On the other hand, the abnormal expression of two products of viral genome, the large envelope protein and the protein HVB X can lead to HCC development (Fig. 2). Transgenic mice expressing large Table 1
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Tracking Dengue Virus Intra-host Genetic Diversity during Human-to-Mosquito Transmission.

Tracking Dengue Virus Intra-host Genetic Diversity during Human-to-Mosquito Transmission.

Adaptability is especially important for mosquito-borne viruses, which encounter distinct selection pressures when cycling between vertebrate and invertebrate hosts. Even within a sin- gle host, intra-host variants generated through multiple rounds of virus replication are subject to evolutionary mechanisms such as bottlenecks, drift, and positive and negative selection pres- sures. In human-derived DENV populations, highly immunogenic E protein domains also dis- play higher levels of intra-host genetic diversity, suggesting that selection pressures on low frequency population variants operate even during acute infection [7]. In mosquitoes, RNA interference (RNAi), a key antiviral defense mechanism in insects, has been proposed to be a driver of viral intra-host genetic diversity in the Culex-West Nile virus (WNV, family Flaviviri- dae) system [8], where higher intra-host diversity levels have been reported in the mosquito than in vertebrate hosts [9,10].
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Anti-foot-and-mouth disease virus effects of Chinese herbal kombucha in vivo

Anti-foot-and-mouth disease virus effects of Chinese herbal kombucha in vivo

To assess the level of viral inhibition, blood samples from swine in the experimental and control groups were collected and analyzed by qRT-PCR to determine the num- ber of FMDV genome copies they contained. None of the samples collected before the viral challenge contained any copies of the FMDV genome (Table 3). After the viral chal- lenge, samples from the intermediate dose group contained the highest levels of FMDV. Samples from the high dose group contained higher levels of FMDV than did those from the control group, and the lowest viral loads were en- countered in the low dose group. These results suggest that Chinese herbal kombucha has a dual effect on FMDV repli- cation, stimulating its reproduction at high doses but inhib- iting it at lower doses. At low doses, Chinese herbal kombucha efficiently inhibits FMDV replication in swine: samples from two animals in the low dose group contained no detectable viral genome copies. Treatment with Chinese herbal kombucha thus completely inhibited FMDV replica- tion in the three asymptomatic animals from the low dose group and the single asymptomatic animal from the inter- mediate dose group. These results are consistent with those presented in Table 3. As shown in Table 4, one-way ANOVA indicated that the numbers of RNA virus copies in blood samples collected from the low dosage group on the first day differed significantly from those for the positive control (C) group, further demonstrating that treatment
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Gene-targeted strategies to eliminate HIV latent cells through synthetic activators and suicide lentivectors

Gene-targeted strategies to eliminate HIV latent cells through synthetic activators and suicide lentivectors

Despite the success of antiretroviral therapy, a cure for HIV-1 infection remains elusive. The persistence of cellular reservoirs harboring transcriptionally silent (latent) HIV provirus is responsible for the viremia rebound observed following treatment withdrawal. Stimulation of latent viral expression is considered critical to target HIV reservoirs for elimination through a “shock and kill” approach. Pharmacological drugs have systematically proven ineffective to drastically reduce the reservoir size and may cause severe side effects owing to their indiscriminate mode of action. In the present thesis, gene-targeted strategies were explored to stimulate and eliminate HIV latent cells. To stimulate latent virus expression, we designed synthetic activators based on transcription activator-like effector (TALE) proteins that recognize conserved regions on HIV 5’LTR promoter. Four TALE activators strongly induced HIV transcription, acting in cooperation to specifically enhance viral expression from cell line models of HIV-1 latency. Moreover, we show that histone deacetylase inhibitors can further enhance the effect of TALE-mediated activation in highly repressed latent cells. To further potentiate the elimination of stimulated latent cells, we conjugated an HIV-responsive suicide lentivector to our TALE activator technology. For this purpose, we incorporated a modified 5’LTR promoter into the suicidal lentivector as a safety mechanism to dissociate TALE-driven activation, restricting the responsiveness of this plasmid to the HIV regulatory proteins. The therapeutic plasmid was capable of specifically eliminate latently infected cells stimulated by TALE activators through a Tat/Rev-dependent expression of the diphtheria toxin. Finally, we presented a “gene-free” approach to specifically activate latent HIV expression through protein delivery of cell-penetrating zinc-finger activators (CPP-ZFA). A single activator based on Cys2His2 zinc-finger domains proved effective at inducing viral expression from the primer binding site downstream of 5’LTR promoter. When conjugated with positively charged nuclear localization signal repeats, this synthetic activator efficiently translocated across cell membrane without the need of carriers. Short-term presence of CPP-ZFA following protein delivery was sufficient to stimulate gene expression in HIV-1 latent cells, offering a safer alternative to avoid off-target effects from prolonged exposure to these synthetic activators. In resume, this work provides proof-of-concept that synthetic activators and suicide lentivectors constitute promising candidates for the eradication of HIV-1 reservoirs through gene-targeted strategies.
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