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At least 775 records · Page 43Linked to original sources
PubMed1998

Sequence analysis of Mus dunni endogenous virus reveals a hybrid VL30/gibbon ape leukemia virus-like structure and a distinct envelope.

Mus dunni endogenous virus (MDEV) can be activated from M. dunni cells by exposing the cells to hydrocortisone or 5-iodo-2'-deoxyuridine. Interference analysis has revealed that MDEV uses a receptor for cell entry that is different from those used by other murine retroviruses. The entire genome has now been sequenced, revealing a long terminal repeat (LTR)-gag-pol-env-LTR structure typical of simple retroviruses of the murine leukemia virus genus, with no additional open reading frames between env and the 3' LTR. The LTRs and other noncoding regions of MDEV are most closely related to those of VL30 elements, while the majority of the coding sequences are most closely related to those of gibbon ape leukemia virus. MDEV represents the first example of a naturally occurring, replication-competent virus with sequences closely related to VL30 elements. The U3 region of MDEV contains six nearly perfect 80-bp repeats and the beginning of a seventh, and the region expected to contain the packaging sequence contains approximately four imperfect 33-bp repeats. The receptor specificity domains of the envelope are unique among retroviruses and show no apparent similarity to regions of known proteins.

Animals↗
PubMed1992

Sequence analysis of the 5' noncoding region of hepatitis C virus.

We have determined the nucleotide sequence of the 5' noncoding (NC) region of the hepatitis C virus (HCV) genome in 44 isolates from around the world. We have identified several HCV isolates with significantly greater sequence heterogeneity than reported previously within the 5' NC region. The most distantly related isolates were only 90.1% identical. Nucleotide insertions were seen in three isolates. Analysis of the nucleotide sequence from 44 HCV isolates in this study combined with that of 37 isolates reported in the literature reveals that the 5' NC region of HCV consists of highly conserved domains interspersed with variable domains. The consensus sequence was identical to the prototype HCV sequence. Nucleotide variations were found in 45 (16%) of the 282 nucleotide positions analyzed and were primarily located in three domains of significant heterogeneity (positions -239 to -222, -167 to -118, and -100 to -72). Conversely, there were three highly conserved domains consisting of 18, 22, and 63 completely invariant nucleotides (positions -263 to -246, -199 to -178, and -65 to -3, respectively). Two nucleotide domains within the 5' NC region, conserved among all HCV isolates studied to date, shared statistically significant similarity with pestivirus 5' NC sequences, providing further evidence for a close evolutionary relationship between these two groups of viruses. Additional analysis revealed the presence of short open reading frames in all HCV isolates. Our sequence analysis of the 5' NC region of the HCV genome provides additional information about conserved elements within this region and suggests a possible functional role for the region in viral replication or gene expression. These data also have implications for selection of optimal primer sequences for the detection of HCV RNA by the PCR assay.

Base Sequence↗
PubMed1995

Detection of enteroviruses and rhinoviruses in clinical specimens by PCR and liquid-phase hybridization.

A sensitive method based on PCR followed by liquid-phase hybridization for detection of enterovirus and rhinovirus RNAs in clinical specimens and cell culture supernatants is described. RNA was extracted from stool samples, throat swabs, nasopharyngeal aspirates, cerebrospinal fluid, urine, and plasma with a commercial phenol-guanidinium-chloroform reagent and purified on a polysulfone membrane, on which the reverse transcriptase reaction was also done. Two sets of oligonucleotide primers from the 5' noncoding region of picornaviruses were selected for DNA amplification of 153-bp (enterovirus) and 120-bp (rhinovirus) regions. Double-stranded amplicons were digested into single strands with T7 gene 6 exonuclease and quantitated by an assay using a europium-labeled probe, streptavidin- and biotinylated probe-coated microtitration wells, and time-resolved fluorometry. The sensitivity of the assay was about one template molecule when purified coxsackievirus A9 RNA was used. All enterovirus prototype strains, except echoviruses 22 and 23, and clinical isolates grown in cell culture or suckling mice were strongly positive by the enterovirus PCR-hybridization, as were selected prototype strains and untyped isolates of rhinoviruses by the rhinovirus PCR-hybridization. In a series of 100 clinical specimens tested, the results for 92 agreed with virus culture results. The detection method described will be useful in etiopathogenic studies on enteroviruses and rhinoviruses.

Animals↗
PubMed2000

Coding changes in the poliovirus protease 2A compensate for 5'NCR domain V disruptions in a cell-specific manner.

Polioviruses are single-stranded RNA viruses with an unusually long noncoding region (NCR) at the 5' end predicted to have an elaborate secondary structure made up of six domains. Mutations in domain V of the poliovirus 5'NCR that disrupt secondary structure are responsible for attenuation of the virus and a temperature-sensitive (ts) phenotype in vitro. In addition to direct back mutation or compensatory second site mutation in the 5'NCR as previously documented, the ts phenotype was found to be compensated for in monkey kidney cells in vitro by a coding change in the protease 2A. These coding changes were found throughout the protease with no obvious pattern or trend. They were not all found to be equivalent and limited in ability to compensate for the severest domain V disruption. The compensatory effect of the 2A changes was found to be cell specific, having no effect on monkey neurovirulence and in a mouse cell line but a significant effect in two monkey cell lines and a human epithelial line.

Amino Acid Sequence↗
PubMed1984

RNA splicing as an error-screening mechanism.

Eukaryote nuclear genes are generally split into coding (exon) and noncoding (intron) regions. During the formation of messenger RNA the introns are precisely excised and the exons religated. A widely accepted explanation for the split structure of eukaryotic genes is that proposed by Gilbert who hypothesized that the division of coding information into small units speeded up the rate of protein evolution by allowing for the recombination of the independent peptide domains encoded by these units ("exon shuffling"). However it has recently become clear that the exon:intron structure of genes most likely preceded the uninterrupted form. This makes it difficult to accept Gilbert's argument as it applies to the origin(s) of split genes since early genes were very inaccurately copied and a highly error-prone system needs less variation not more. Here I propose that split genes and the concomitant process of RNA splicing arose as a mechanism for maintaining the stability of the genetic information in the face of a high level of noise in the gene copier mechanism.

Base Sequence↗
PubMed2002

Detection of HCV RNA in gastric mucosa-associated lymphoid tissue by in situ hybridization: evidence of a new extrahepatic localization of HCV with increased risk of gastric malt lymphoma.

OBJECTIVES: Mucosa-associated lymphoid tissue (MALT) is absent in the normal gastric mucosa but it can develop in several conditions, such as Helicobacter pylori infection. A certain correlation between hepatitis C virus (HCV) infection and low grade MALT lymphomas has recently been reported. The aim of this study was to investigate the presence of HCV RNA in acquired gastric MALT of HCV-infected patients using the in situ hybridization technique. METHODS: Twenty-five patients (16 male and nine female, average age = 56.6 yr [range = 33-75]) affected by chronic HCV hepatitis and with gastric MALT were studied. Giemsa stain and the rapid urease test were also used to evaluate the presence of H. pylori. A polymerase chain reaction product corresponding to the complete 5' noncoding region of the HCV genome was cloned directly in the pCR 1000 vector on gastric biopsies with acquired MALT. RESULTS: Twenty patients showed grade 2 gastric MALT and five showed grade 3, and H. pylori's presence was detected in 18 of 25 patients (72%). Using in situ hybridization, we detected HCV RNA in gastric acquired MALT of seven of 25 patients (28%): five showed grade 2 gastric MALT (two of these were H. pylori negative and the other three were positive), whereas two patients showed grade 3 gastric MALT (without H. pylori infection). CONCLUSIONS: This study shows that HCV not only may colonize gastric MALT but also may permit the development of a grade of acquired MALT, which may represent the first step toward a MALT lymphoma. However, further studies are needed to demonstrate the antigenic role of HCV in the progression of acquired MALT into MALT lymphoma.

Adult↗
PubMed1986

Matrix genes of measles virus and canine distemper virus: cloning, nucleotide sequences, and deduced amino acid sequences.

The nucleotide sequences encoding the matrix (M) proteins of measles virus (MV) and canine distemper virus (CDV) were determined from cDNA clones containing these genes in their entirety. In both cases, single open reading frames specifying basic proteins of 335 amino acid residues were predicted from the nucleotide sequences. Both viral messages were composed of approximately 1,450 nucleotides and contained 400 nucleotides of presumptive noncoding sequences at their respective 3' ends. MV and CDV M-protein-coding regions were 67% homologous at the nucleotide level and 76% homologous at the amino acid level. Only chance homology was observed in the 400-nucleotide trailer sequences. Comparisons of the M protein sequences of MV and CDV with the sequence reported for Sendai virus (B. M. Blumberg, K. Rose, M. G. Simona, L. Roux, C. Giorgi, and D. Kolakofsky, J. Virol. 52:656-663; Y. Hidaka, T. Kanda, K. Iwasaki, A. Nomoto, T. Shioda, and H. Shibuta, Nucleic Acids Res. 12:7965-7973) indicated the greatest homology among these M proteins in the carboxyterminal third of the molecule. Secondary-structure analyses of this shared region indicated a structurally conserved, hydrophobic sequence which possibly interacted with the lipid bilayer.

Amino Acid Sequence↗
PubMed1991

Bluetongue virus evolution: sequence analyses of the genomic S1 segments and major core protein VP7.

The S1 segments, encoding the group-specific antigen, VP7, from the five United States prototype BTV serotypes were cloned as full-length entities. The nucleotide and deduced amino acid sequences of segment S1 of BTV-2 were determined and compared with BTV-10, -11, -13, and -17, completing the sequencing of this cognate gene segment from all five US BTV serotypes. Each segment is 1156 bp long and contains an open reading frame encoding the 349-amino acid VP7 protein. Most (greater than 94%) of the amino acids of VP7 among the serotypes are conserved, including the location (position 255) of a single lysine residue. Secondary structure analyses of VP7 predict a putative eight-stranded beta-barrel between amino acid positions 150 and 250, a structure similar to that observed in ssRNA viruses. The S1 genes are flanked by conserved 5' and 3' noncoding regions. Stem-loop structures are predicted at the 3' end of each gene (nucleotide positions 1058-1097). The S1 segments of BTV-2, -10, -11, and -17 have greater than 93% of the nucleotides conserved, while less than 80% of their bases are identical with BTV-13. Analyses of nucleotide mismatches in each codon position of the VP7 open reading frame, transition frequencies, and evolutionary distances show that of the five, BTV-13 is the most distantly related and that BTV-10 and -17 are the most closely related serotypes. Evolutionary distance calculations of segment L2 from BTV-10, -11, and -17 concur with these observations. Comparison of this relationship with hybridization data of segment M3, which codes for VP5, suggests that BTV-17 has evolved by a combination of genetic drift and genomic reassortment. The data also indicate that the five US BTV serotypes are derived from two distinct gene pools. Evolution distances were used to estimate an evolution rate of 2.2 x 10(-3) nucleotide substitution/site/year for BTV segment S1. This rate is similar to the genes of retroviruses and implies an absence of RNA polymerase proofreading activity for dsRNA viruses.

Amino Acid Sequence↗
PubMed2001

Analysis of the noncoding regions of measles virus strains in the Edmonston vaccine lineage.

The noncoding sequence of five Edmonston vaccine viruses (AIK-C, Moraten, Rubeovax, Schwarz, and Zagreb) and those of a low-passage Edmonston wild-type (wt) measles virus have been determined and compared. Twenty-one nucleotide positions were identified at which Edmonston wt and one or more vaccine strains differed. The location of some of these nucleotide substitutions suggests that they may influence the efficiency of mRNA synthesis, processing, and translation, as well as genome replication and encapsidation. Five nucleotide substitutions were conserved in all of the vaccine strains. Two of these were in the genomic 3'-terminal transcriptional control region and could affect RNA synthesis or encapsidation. Three were found within the 5'-untranslated region of the F mRNA, potentially altering translation control sequences. The remaining vaccine virus base changes were found in one to four vaccine strains. Their genomic localization suggests that some may modify cis-acting regulatory domains, including the Kozak consensus element of the P and M genes, the F gene-end signal, and the F mRNA 5'-untranslated sequence.

5' Untranslated Regions↗
PubMed1993

Relationship between alcoholic liver disease and HCV infection.

The high prevalence of hepatitis C virus (HCV) markers in alcoholic liver cirrhosis (AL-LC) and hepatocellular carcinoma (HCC) suggests a close aetiopathogenic relationship between alcoholic liver disease (ALD) and HCV infection. In the present study, HCV markers in ALD were measured by the highly sensitive methods, and the changes of sequential HCV markers after abstinence in ALD patients were analysed in order to elucidate the effect of alcohol on HCV. Antibodies to HCV-related antigen were determined using the first or second generation test kit. HCV-RNA genomes encoding the NS-5 region were detected using the RT-PCR method. In the HCV-NS5 negative serum, HCV genomes of the 5'-noncoding region were detected using the two-stage PCR method. Titres of HCV-RNA were measured by multiple cyclic PCR and cDNA dot blotting. Typing of HCV genomes was carried out on the PCR product from the NS-5 region by slot blot hybridization using type-specific cDNA probes, or by restriction fragment length polymorphisms analysis. In alcoholic fibrosis and alcoholic hepatitis, the prevalence of HCV markers was low, suggesting that the main aetiological factor is alcohol but not HCV in these types of ALD. HCV markers were positive in the half of the patients with AL-LC, and in more than 80% of patients with AL-CH and AL-HCC, indicating that HCV infection closely relates to these types of ALD. The ratio of the K1 type to the K2 type of HCV genomes was 4:1 in all types of NANB liver disease.(ABSTRACT TRUNCATED AT 250 WORDS)

Adult↗
PubMed1992

Neutralizing epitopes on herpes simplex virus-1-expressed rotavirus VP7 are dependent on coexpression of other rotavirus proteins.

We constructed a recombinant thymidine kinase-negative herpes simplex virus type 1 (HSV-1) that expressed the rotavirus major outer capsid glycoprotein, VP7. In the recombinant HSV-1, a promoter from the 5' noncoding region of the HSV-1 glycoprotein B locus regulated the expression of VP7 as a HSV-1 gamma 1 gene product. HSV-1-expressed VP7 resembled rotavirus-expressed VP7 in its SDS-PAGE mobility, high mannose-type glycosylation, disulfide bonding, perinuclear to cytoplasmic localization, intracellular retention, and reactivity with polyclonal antisera and nonneutralizing antibodies. Unlike rotavirus-expressed VP7, HSV-1-expressed VP7 lacked several neutralizing epitopes by immuno-histochemical staining and by ELISA. One neutralizing epitope identified on HSV-1-expressed VP7 by ELISA was masked by paraformaldehyde fixation of recombinant HSV-1- but not rotavirus-infected cells. Neutralizing epitopes were restored to HSV-1-expressed VP7 by coinfection of cells with the HSV-1 recombinant and a heterologous rotavirus that lack the neutralizing epitopes. The recovered neutralizing epitopes were detected on double-shelled rotavirus particles produced in the coinfected cells. This study indicates that the formation of several neutralizing epitopes on rotavirus VP7 requires interaction of VP7 with other rotavirus proteins. In addition, HSV-1 was a useful vector for studying the localization, processing, and antigenicity of an RNA virus glycoprotein.

Animals↗
PubMed2002

High level of sequence variation in the 3' noncoding region of Japanese encephalitis viruses isolated in Korea.

The 3' noncoding region (NCR) of Japanese encephalitis (JE) viruses isolated in Korea and Nakayama-NIH strain have been sequenced and compared with the 3' NCR sequences of other JE isolates reported previously. Sequence alignment of about 60 nucleotides (based on consensus sequence number) immediately downstream of the open reading frame (ORF) stop codon in the 3' NCR of the Korean isolates showed high degree of sequence variation and deletion; thus, this region was termed as the variable region. However, in the predicted RNA secondary structures, a similar type loop exists at the 5'-terminus of the 3' NCR of JE viruses, despite low level of sequence homology (22%) and deletion in the variable region. The phylogenetic tree based on the 3' NCR sequences of JE viruses including the variable region showed a similar pattern to that based on envelope genes; in that, there are two genetically different types of JE viruses in Korea. Therefore, the variable region would be a useful genetic marker for JE viruses.

3' Flanking Region↗
PubMedDate not supplied

Conjugation of an antisense oligodeoxynucleotide to ribonuclease h results in sequence-specific cleavage and intracellular inhibition of HCV gene expression.

A recombinant E. coli ribonuclease H (RNase H) was chemically coupled to an antisense oligodeoxynucleotide (ODN) against the 5'-noncoding region (5'-NCR) of the hepatitis C virus. Purity of the conjugates was confirmed by sodium deodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) as a band corresponding to approximately 23 kDa. Conjugate function was tested by the cleavage of a HCV RNA transcript including the 5'-NCR and core region and showed HCV sequence-specific cleavage by the appearance of an expected approximately 1000 nt fragment of RNA. Cleavage was not seen by RNase H alone, or ODN alone. Delivery studies using (32)P- and (125)I-labeling showed that while RNAse H failed to enter cells, the conjugate was efficiently taken into the cells. To assess intracellular effects, a cell line, Huh-7/CMV-NCRCDeltaluc, which expresses HCV mRNA (nt 1-585) fused to a marker gene, was transfected with the conjugate. Reporter gene expression was suppressed by 51.2% with the conjugate compared to only 39.7% by ODN alone, 35.8% by a mixture of RNase H plus ODN, and not at all by RNase H alone. In conclusion, the RNase H-ODN conjugate effectively cleaved an HCV transcript in vitro and inhibited expression of an HCV-marker fusion construct in a liver-derived cell line.

Cell Line↗
PubMed2001

Ribosomal protein S5 interacts with the internal ribosomal entry site of hepatitis C virus.

Translational initiation of hepatitis C virus (HCV) genome RNA occurs via its highly structured 5' noncoding region called the internal ribosome entry site (IRES). Recent studies indicate that HCV IRES and 40 S ribosomal subunit form a stable binary complex that is believed to be important for the subsequent assembly of the 48 S initiation complex. Ribosomal protein (rp) S9 has been suggested as the prime candidate protein for binding of the HCV IRES to the 40 S subunit. RpS9 has a molecular mass of approximately 25 kDa in UV cross-linking experiments. In the present study, we examined the approximately 25-kDa proteins of the 40 S ribosome that form complexes with the HCV IRES upon UV cross-linking. Immunoprecipitation with specific antibodies against two 25-kDa 40 S proteins, rpS5 and rpS9, clearly identified rpS5 as the protein bound to the IRES. Thus, our results support rpS5 as the critical element in positioning the HCV RNA on the 40 S ribosomal subunit during translation initiation.

5' Untranslated Regions↗
PubMed1986

Scrapie and cellular PrP isoforms are encoded by the same chromosomal gene.

PrP 27-30 is the major protein in purified preparations of scrapie agent. An almost complete PrP cDNA was used to select PrP-related genomic clones from normal hamster DNA. The gene contains a noncoding exon of 56 to 82 bp and a 2 kb coding exon, separated by a 10 kb intron. Transcription initiates at the same multiple sites in vivo and in vitro. The promoter lacks a TATA box and contains three repeats of the sequence GCCCCGCCC, which resembles the Sp1 binding site found in "housekeeping" genes. The PrP coding sequence encodes a presumptive amino-terminal signal peptide. The primary structure of PrP encoded by the gene of a healthy animal does not differ from that encoded by a cDNA from a scrapie-infected animal, suggesting that the different properties of PrP from normal and scrapie-infected brains are due to post-translational events.

Animals↗
PubMed1997

Impact of dengue virus infection and its control.

Dengue virus infection has been counted among emerging and re-emerging diseases because of (1) the increasing number of patients, (2) the expansion of epidemic areas, and (3) the appearance of severe clinical manifestation of dengue hemorrhagic fever (DHF)/dengue shock syndrome (DSS), which is often fatal if not properly treated. In the meantime, there are no effective dengue control measures: a dengue vaccine is still under development and vector control does not provide a long-lasting effect. In order to obtain direct evidence for the virulent virus theory concerning the pathogenesis of DHF/DSS, type 2 dengue virus strains isolated from patients with different clinical severities in the same epidemic area in northeast Thailand, during the same season, were comparatively sequenced. The result revealed a DF strain specific amino acid substitution from I to R in the PrM, and a DSS strain specific amino acid substitution from D to G in the NS1 gene regions, which could significantly alter the nature of these proteins. Moreover, DF strain specific nucleotide substitutions in the 3' noncoding region were predicted to alter its secondary structure. These amino acid and nucleotide substitutions in other strains isolated in different epidemic areas during other seasons, together with their biological significance, remain to be confirmed. In order to innovate dengue vector control, field tests were carried out in dengue epidemic areas in Vietnam to examine the efficacy of Olyset Net screen, which is a wide-mesh net made of polyethylene thread impregnated with permethrin. The results show that Olyset Net (1) reduced the number of principal dengue vector species, Aedes aegypti, (2) interrupted the silent transmission of dengue viruses and (3) was highly appreciated by the local people as a convenient and comfortable vector control method. This encouraging evaluation of the Olyset Net screen should be confirmed further by other tests under different settings.

Aedes↗
PubMed1988

Presence of transcription regulatory elements within an intron of the virus-inducible murine TIMP gene.

The expression of the gene for the murine tissue inhibitor of metalloproteinases (TIMP) is induced in response to viruses, growth factors, and phorbol esters. In this report we show that the accumulation of TIMP mRNA after Newcastle disease virus induction is caused by transcriptional activation of the gene. Comparison of the sequences of cDNA and genomic clones along with RNase protection and primer extension analyses revealed that the murine TIMP gene possesses multiple cap sites and that the exon 1 consists exclusively of 5'-noncoding sequences. We observed that DNA regions analogous to those found upstream of the virus-inducible interferon genes are present within intron 1 of the TIMP gene. To investigate the possible role of TIMP intron 1 in gene expression, we used a functional assay based on the transfection of plasmids in which the DNA segment to be tested is placed in proximity to a marker gene driven by the heterologous herpes simplex virus thymidine kinase promoter. Our results indicate that TIMP intron 1 contains DNA sequence elements capable of modulating the activity of a heterologous promoter in two different ways: (i) by enhancing constitutive expression and (ii) by conferring virus inducibility. These results suggest that intron 1 may be involved in the transcriptional regulation of TIMP gene expression.

Animals↗
PubMedDate not supplied

Global eradication of poliomyelitis: reinvent the wheel or use existing options effectively?

The development of serum and nasopharyngeal antibody response, and the magnitude and temporal pattern of fecal shedding of vaccine and revertant polioviruses was examined in infants previously immunized with one or more doses of oral poliovirus vaccine (OPV), enhanced-potency inactivated poliovirus vaccine (EIPV), or both. The fecal samples were collected at different intervals after the last OPV dose. The nucleic acid sequences of the purified RNA obtained from all virus isolates were examined in the 5' noncoding region by dideoxysequencing to determine whether the viruses shed represent revertants (vaccine), nonrevertants, or both. The frequency and duration of vaccine virus-shedding appeared to be similar in both immunization schedules. Revertant virus shedding was not demonstrated 30 days after immunization with OPV alone. However, shedding of revertants was detected for as long as 60 days in some subjects previously immunized with EIPV. The duration of shedding of revertant virus differed with different serotypes and different immunization regimens. Prior immunization with one or more doses of OPV reduced the length of shedding of revertant virus. Significantly, however, prior immunization with one or more doses of EIPV was not associated with reduced shedding of revertant virus types. The nature of serum immune response as determined by ELISA or neutralizing antibody appeared to be similar after either immunization schedule, although the antibody titers were quantitatively higher after two doses of IPV than observed after a similar schedule with OPV. Highest antibody activity was detected in subjects immunized with a combination of IPV followed by OPV. ELISA antibody activity in the nasopharynx was regularly detected after either form of immunization.(ABSTRACT TRUNCATED AT 250 WORDS)

Antibodies, Viral↗