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PubMed1989

Determination of the coding capacity of the M genome segment of nephropathia epidemica virus strain Hällnäs B1 by molecular cloning and nucleotide sequence analysis.

The M genome RNA segment of nephropathia epidemica virus (NEV) strain Hällnäs B1 was characterized by molecular cloning and DNA nucleotide sequencing of the corresponding cDNA clones. The size of the M RNA segment is 3682 nucleotides. The 3' and 5' terminal sequences are complementary for 21 bases and their predicted secondary structure is very stable. The viral complementary messenger RNA possesses a single long open reading frame with a coding capacity of 1148 amino acids (polypeptide of 126 kDa). A comparison of the NEV M segment to that of Hantaan virus strain 76-118 reveals 61% sequence homology at the nucleotide level and 53% at the deduced amino acid level. Four out of five potential asparagine-linked glycosylation sites of the encoded glycoproteins have been conserved between NEV and Hantaan M. The isoelectric points (IEP) are nearly identical. Furthermore it was found that 90% of all cysteine residues have been conserved. Putative NEV G1 and G2 are preceded by a short hydrophobic sequence as shown for G1 and G2 of Hantaan virus. Hydrophilicity profiles of the two segments are of striking similarity. These data indicate that NEV- and Hantaan virus M-encoded polypeptides seem to be very similar in structure and function despite the relatively low amino acid sequence homology.

Amino Acid Sequence↗

PubMed2004

DAGchainer: a tool for mining segmental genome duplications and synteny.

SUMMARY: Given the positions of protein-coding genes along genomic sequence and probability values for protein alignments between genes, DAGchainer identifies chains of gene pairs sharing conserved order between genomic regions, by identifying paths through a directed acyclic graph (DAG). These chains of collinear gene pairs can represent segmentally duplicated regions and genes within a single genome or syntenic regions between related genomes. Automated mining of the Arabidopsis genome for segmental duplications illustrates the use of DAGchainer.

Algorithms↗

PubMed1995

Characterization of the small open reading frame on genome segment A of infectious pancreatic necrosis virus.

The genome of infectious pancreatic necrosis virus (IPNV) is composed of two segments of dsRNA. The larger segment contains a small ORF partly overlapping the 5' end of the polyprotein reading frame. Yet very little is known about this possible new gene, which presumably codes for a 17 kDa polypeptide (VP5). The region of the viral genome which encompasses the small ORF was reverse-transcribed and amplified by PCR before cloning and sequencing. Analysis of the sequences obtained from five different virus strains revealed that the small ORF is not found on one of them, and that it is truncated on two others. Moreover, the deduced amino acid sequences did not appear to be well conserved. Despite the large variations between IPNV strains at the genomic level, all predicted VP5 are arginine-rich basic polypeptides. To verify whether the small ORF is translated into protein in fish cells, the 17 kDa polypeptide of the VR-299 strain was expressed as fusion protein in a prokaryotic expression vector and used to produce a specific antiserum. This antiserum reacted with concentrated virus in an immunodot assay indicating that VP5 is synthesized in infected cells, but probably only in small quantities. When tested with 12 other IPNV strains, results were less conclusive than those obtained with strain VR-299. Nevertheless, three of the 12 viruses gave a clearly negative signal in the immunodot assay, suggesting that possibly more than one viral strain lacks the small ORF.

Amino Acid Sequence↗

PubMed1988

Virus-lymphocyte interactions. IV. Molecular characterization of LCMV Armstrong (CTL+) small genomic segment and that of its variant, Clone 13 (CTL-).

Immunocompetent adult mice mount a vigorous cytotoxic T lymphocyte (CTL) response against the Armstrong (ARM) 53b strain of LCMV after primary inoculation. In contrast, the Clone 13 variant of ARM 53b, originally isolated from the spleen of a persistently infected mouse, suppresses LCMV-specific CTL responses (R. Ahmed et al. (1984) J. Exp Med 60, 521). The induction and generation of CTL maps to the short (S) RNA segment and not the long (L) RNA segment of LCMV (Y. Riviere et al. (1986) J. Immunol. 136, 304). The CTL recognition epitope, expressed in virus-infected target cells, also maps to the S segment of the LCMV ARM genome, and is structurally and functionally intact in Clone 13-infected target cells. Here we report the S RNA sequences of both ARM 53b and its variant Clone 13. Comparison reveals a single amino acid difference. However, sequence divergence at this position also occurs among other strains of LCMV (Pasteur, Traub, WE) which do elicit CTL responses. Hence, (1) the amino acid difference is unrelated to the phenotypic divergence of Clone 13, (2) suppression of the CTL response by Clone 13 is not linked to the CTL recognition epitope, and (3) the structure or function responsible for CTL immunosuppression by Clone 13 most likely maps to the L RNA segment. Further, the availability of the complete S RNA sequence for LCMV ARM and ARM Clone 13 variant allows a detailed comparison with WE (V. Romanowski et al. (1985) Virus Res. 3, 110-114), the only other LCMV S RNA so far sequenced.

Amino Acid Sequence↗

PubMed1993

Putative full-length clones of the genomic DNA segments of subterranean clover stunt virus and identification of the segment coding for the viral coat protein.

Subterranean clover stunt disease is an economically important aphid-borne virus disease affecting certain pasture and grain legumes in Australia. The virus associated with the disease, subterranean clover stunt virus (SCSV), was previously found to be representative of a new type of single-stranded DNA virus. Analysis of the virion DNA and restriction mapping of double-stranded cDNA synthesized from virion DNA suggested that SCSV has a segmented genome composed of 3 or 4 different species of circular ssDNA each of about 850-880 nucleotides. To further investigate the complexity of the SCSV genome, we have isolated the replicative form DNA from infected pea and from it prepared putative full-length clones representing the SCSV genome segments. Analysis of these clones by restriction mapping indicated that clones representing at least 4 distinct genomic segments were obtained. This method is thus suitable for generating an extensive genomic library of novel ssDNA viruses containing multiple genome segments such as SCSV and banana bunchy top virus. The N-terminal amino acid sequence and amino acid composition of the coat protein of SCSV were determined. Comparison of the amino acid sequence with partial DNA sequence data, and the distinctly different restriction maps obtained for the full-length clones suggested that only one of these clones contained the coat protein gene. The results confirmed that SCSV has a functionally divided genome composed of several distinct ssDNA circles each of about 1 kb.

Amino Acid Sequence↗

PubMed1991

Molecular characterization of the M genomic segment of the Seoul 80-39 virus; nucleotide and amino acid sequence comparisons with other hantaviruses reveal the evolutionary pathway.

The genomic M segment of Seoul 80-39 virus was characterized by cloning and nucleotide sequence analysis. The virion M RNA segment is 3651 nucleotides long with the 3' and 5' terminal sequences inversely complementary for 20 bases. A single open reading frame was detected in the viral complementary-sense RNA which can encode a polypeptide of 1133 amino acids. The Seoul 80-39 virus M segment was compared with the M segments of related viruses, Hantaan 76-118, Hallnas B1 and Sapporo Rat (SR-11) virus. Our results demonstrate a significant similarity between M RNA segments of the Seoul 80-39, Hantaan 76-118, Hallnas B1 and SR-11 viruses. The degree of conservation of both nucleic acid and protein sequences between these viruses reveals a close evolutionary relationship. Furthermore, it is evident that the serotypic profile of hantaviruses is determined by the rodent host species from which the virus was isolated and not by the geographical area.

Amino Acid Sequence↗

PubMed2003

Effects of a point mutation in the 3' end of the S genome segment of naturally occurring and engineered Bunyamwera viruses.

The genome of Bunyamwera virus (BUN) consists of three segments of single-stranded RNA of negative polarity. The smallest segment, S, encodes the N protein and a nonstructural protein called NSs. We recently described a mutant virus (BUNdelNSs) that does not express NSs but overexpresses N and grows to lower titres than wild-type (wt) BUN. Here we report a BUNdelNSs variant that expresses lower levels of N protein and grows to higher titres. Sequencing of the 3' and 5' termini of the BUNdelNSs S RNA segment and analysis using a minireplicon system show that the N overexpressing phenotype results from a single nucleotide substitution at position 16 in the 3' terminus. This mutation could also be detected in wtBUN populations, and was isolated by plaquing a 'wt' variant carrying the mutation. This variant was found to express increased N and NSs levels, and grew to lower titres than wtBUN.

3' Untranslated Regions↗

PubMed2004

Genomic segmental polymorphisms in inbred mouse strains.

By analyzing genomic copy-number differences using high-resolution mouse whole-genome BAC arrays, we uncover substantial differences in regional DNA content between inbred strains of mice. The identification of these apparently common segmental polymorphisms suggests that these differences can contribute to genetic variability and pathologic susceptibility.

Animals↗

PubMed1988

Rearrangement of genomic segment 11 in two swine rotavirus strains.

We have recently reported the isolation of two group A swine rotaviruses each lacking normal genomic RNA segment 11 and showing instead one extra segment that migrated abnormally on gel electrophoresis. Hybridization studies performed with segment-specific probes and with a purified abnormal RNA segment showed that the extra bands had sequence homology to normal segment 11. Analysis of protein profiles of normal and rearranged strains showed that the gene product of segment 11 had no apparent change in its relative electrophoretic migration, suggesting that the rearranged genes remained functional.

Animals↗

PubMed2005

A base-specific recognition signal in the 5' consensus sequence of rotavirus plus-strand RNAs promotes replication of the double-stranded RNA genome segments.

Replication of the segmented double-stranded (ds)RNA genome of rotavirus requires the viral RNA-dependent RNA polymerase (RdRP) to use 11 different (+)RNAs as templates for (-) strand synthesis. Complementary sequences proximal to the 5' and 3' termini are predicted to direct cyclization of the (+)RNAs by forming panhandle structures from which short highly conserved terminal sequences protrude as single-stranded tails. Cell-free replication assays indicate that such structural organization of the 5'- and 3'-ends is required for efficient dsRNA synthesis. Multiple specifically recognized elements exist at the 3'-end that promote dsRNA synthesis including RdRP-recruitment signals and a (-) strand initiation sequence. In contrast to the 3'-end, the role of the 5'-end has been less well defined. In this study, we determined that the 5'-end contains a base-specific recognition signal that plays an important role in the assembly of the RdRP and cofactors into a stable initiation complex for (-) strand synthesis. The 5' recognition signal is associated with the G2 residue of the 5'-consensus sequence, a residue that shows absolute conservation among all rotavirus groups (A, B, and C) examined to date. From our results, we suggest that rotavirus (+)RNA cyclization, although likely initiated by 5'- 3' nucleotide complementarity, may be stabilized by RdRP-dependent bridging. Given that synthesis of the (-) strand on the (+)RNA template will disrupt 5'-3' nucleotide interactions, RdRP-dependent bridging may be the sole mechanism by which the dsRNA product can be held in the necessary cyclized conformation required for efficient multiple rounds of transcription.

Base Sequence↗

PubMed2004

Packaging, replication and recombination of the segmented genome of bacteriophage Phi6 and its relatives.

The genomes of bacteriophage Phi6 and its relatives are packaged through a mechanism that involves the recognition and translocation of the three different plus strand transcripts of the segmented dsRNA genomes into preformed polyhedral structures called procapsids or inner cores. The packaging requires hydrolysis of NTPs and takes place in the order S:M:L. Minus strand synthesis begins after the completion of the plus strand packaging. The packaging and replication reactions can be studied in vitro with purified components. A model has been presented that proposes that the program of serially dependent packaging is determined by the conformational changes at the surface of the procapsid due to the amount of RNA packaged at each step. The in vitro packaging and replication system has facilitated the application of reverse genetics and the study of recombination in the family of Cystoviridae.

Bacteriophage phi 6↗

PubMed1982

Assignment of reovirus mutant group 'G' to its genomic segment.

Temperature sensitive (ts+) recombinants between the UV-inactivated reovirus type 1 and the group G ts mutant of reovirus type 3 were obtained using a cross-reactivation technique. Analysis of the dsRNA of recombinant clones in PAGE allowed a preliminary mapping of the ts mutation into its genomic S3 segment. This was confirmed by the hybridization test.

Genes, Viral↗

PubMed1995

Coding strategy of the S and M genomic segments of a hantavirus representing a new subtype of the Puumala serotype.

The hantavirus strain Vranica was previously reported to have been isolated from a bank vole in Bosnia-Hercegovina and associated with the occurrence of hemorrhagic fever with renal syndrome (HRFS) in humans. The complete cDNA nucleotide sequences of the small (S) and medium (M) genomic RNA segments of this virus were determined. Major open reading frames were found in the S and M segment between nucleotide positions 43 and 1341 coding for a polypeptide of 433 amino acid residues and between nucleotide positions 41 and 3,484 coding for 1,148 amino acid residues, respectively. The analysis and the alignment of the nucleotide and the derived amino acid sequences with known sequences of other hantavirus strains demonstrate that Vranica resembles Swedish strains and represents a new virus subtype of the Puumala serotype distinct from the subtypes represented by virus strains CG18-20 and Sotkamo.

Amino Acid Sequence↗

PubMed2002

Genome segmentation using piecewise constant intensity models and reversible jump MCMC.

The existence of whole genome sequences makes it possible to search for global structure in the genome. We consider modeling the occurrence frequencies of discrete patterns (such as starting points of ORFs or other interesting phenomena) along the genome. We use piecewise constant intensity models with varying number of pieces, and show how a reversible jump Markov Chain Monte Carlo (RJMCMC) method can be used to obtain a posteriori distribution on the intensity of the patterns along the genome. We apply the method to modeling the occurrence of ORFs in the human genome. The results show that the chromosomes consist of 5-35 clearly distinct segments, and that the posteriori number and length of the segments shows significant variation. On the other hand, for the yeast genome the intensity of ORFs is nearly constant.

Algorithms↗

PubMed1992

Nucleotide sequence comparison of the M1 genome segment of reovirus type 1 Lang and type 3 Dearing.

The mammalian reoviruses possess a genome composed of 10 double stranded RNA segments. The serotype 1 strain Lang M1 segment was sequenced and compared to the published type 3 sequence. Both segments were 2304 base-pairs long coding for the mu 2 protein predicted to be 736 amino acids long. The sequences were highly conserved with 97.2% conservation of nucleotide sequence and 98.6% conservation of amino acid sequence. The M1 segments of serotypes 1 and 3 have recently diverged as indicated by the distribution of variation with respect to codon positions. The conservation of amino acid sequence indicated that the mu 2 protein has a relatively high functional density.

Amino Acid Sequence↗

PubMed1988

Molecular epidemiology of human rotavirus infections based on genome segment variations in viral strains.

Molecular epidemiology of rotavirus infections in 621 hospitalized children was investigated by analysis of migration patterns of viral genomic ribonucleic acid (RNA) segments by electrophoresis in polyacrylamide gels. Based on migration patterns of RNA segments of 184 rotavirus strains, seven different electropherotypes were identified: 146 (79.3%) strains were "long," and 38 (20.7%) were "short" electropherotypes; 61% belonged to a single dominant "long" electropherotype, which persisted throughout the 15-month period of study, whereas six other cocirculating types appeared at varying intervals. Electrophoretic migration patterns of RNA from viral isolates of two patients suggested mixed infections with different rotaviruses. There was a lack of correlation between the electrophoretic migration of segments 10 and 11 and serologically defined subgroup specificity in three of the rotavirus strains. Rotavirus infections and different electropherotypes were observed throughout the year.

Child, Preschool↗

PubMed2000

Duplication and diversification of the apolipoprotein CI (APOCI) genomic segment in association with retroelements.

We have previously shown that several multicopy gene families within the major histocompatibility complex (MHC) arose from a process of segmental duplication. It has also been observed that retroelements play a role in generating diversity within these duplicated segments. The objective of this study was to compare the genomic organization of a gene duplication within another multicopy gene family outside the MHC. Using new continuous genomic sequence encompassing the APOE-CII gene cluster, we show that APOCI and its pseudogene, APOCI', are contained within large duplicated segments which include sequences from the hepatic control region (HCR). Flanking Alu sequences are observed at both ends of the duplicated unit, suggesting a possible role in the integration of these segments. As observed previously within the MHC, the major differences between the segments are the insertion of sequences (approximately 200-1000 bp in length), consisting predominantly of Alu sequences. Ancestral retroelements also contribute to the generation of sequence diversity between the segments, especially within the 3' poly(A) tract of Alu sequences. The exonic and regulatory sequences of the APOCI and HCR loci show limited sequence diversity, with exon 3 being an exception. Finally, the typing of pre- and postduplication Alus from both segments indicates an estimated time of duplication of approximately 37 million years ago (mya), some time prior to the separation of Old and New World monkeys.

Alu Elements↗

PubMed1982

Polymorphism of the migration of double-stranded RNA genome segments of avian reoviruses.

A number of field isolates of avian reovirus were characterized by analysis of the migration pattern of their genomic double-stranded RNA (dsRNA) segments upon polyacrylamide gel electrophoresis. Comparison of the various isolates has demonstrated (i) no relationship between serotype and migration of any individual dsRNA segment, (ii) marked polymorphism of migration patterns of all dsRNA segments among isolates of the same serotype as well as among different serotypes, (iii) no correlation between genotype and disease state, (iv) less marked variability in migration pattern from isolates within a restricted geographic locale compared to isolates from distant locales, (v) the presence of a single genotype in local outbreaks of disease, and (vi) the relative invariant migration of several dsRNA segments among the avian reoviruses, one of which (S1) may serve to distinguish the avian from the mammalian reoviruses.

Animals↗