Dugbe nairovirus M RNA: Nucleotide sequence and coding strategy

The coding assignments of the medium-sized (M) RNA segment of the Dugbe (DUG) virus (Nairovirus, Bunyaviridae) were investigated. The complete nucleotide sequence of 4888 nucleotides (nt) contained one long open reading frame in the viral complementary RNA, extending from an AUG start codon at nt 48-50 to a stop codon at nt 4701-4703 (numbered from the 5' terminus of vcRNA). Comparison of the terminal sequences with the ends of the DUG S segment revealed sequence identity between the first nine nucleotides of both segments. No sequence homologies were found with the M segments of other members of the Bunyaviridae, or with their polypeptide products. Expression of portions of the DUG M open reading frame in Escherichia coli demonstrated the carboxyl terminal region of the M open reading frame codes for the G1 structural glycoprotein, which is the target for neutralising antibodies. Confirmation of this assignment was obtained by sequencing the amino terminus of the G1 protein. Two nonstructural glycoproteins which share epitopes with G1 were identified in virus-infected cells, one of which (85 kDa) is processed over a period of several hours to produce G1. The G2 coding region was located upstream of the G1 sequence. The region between the carboxyl terminus of G2 and the 5' end of the long open reading frame apparently encodes a nonstructural protein of about 70 kDa, which is a precursor of the G2 protein.     

Nucleotide sequence of barley stripe mosaic virus RNAα: RNAα encodes a single polypeptide with homology to corresponding proteins from other viruses

The complete nucleotide sequence of RNAα from the Type strain of barley stripe mosaic virus has been determined. The RNA is 3768 nucleotides long and contains a single open reading frame which codes for a polypeptide of 1139 amino acids (mw 129,634). The open reading frame is flanked by a 5′-terminal sequence of 91 nucleotides and a 3′-nontranslated region composed of a short poly(A) tract followed by a 238-nucleotide tRNA-like structure. The amino acid sequence of the polypeptide (αa) encoded by the open reading frame has homology with the TMV 126K protein and with related polypeptides from other viruses. The carboxy-terminal portion of the as polypeptide also has limited homology with the 58K (βb) protein encoded by BSMV RNAβ and includes a consensus sequence found in mononucleotide-binding polypeptides.     

Nucleotide sequence of barley stripe mosaic virus RNA alpha: RNA alpha encodes a single polypeptide with homology to corresponding proteins from other viruses

The complete nucleotide sequence of RNA alpha from the Type strain of barley stripe mosaic virus has been determined. The RNA is 3768 nucleotides long and contains a single open reading frame which codes for a polypeptide of 1139 amino acids (mw 129,634). The open reading frame is flanked by a 5'-terminal sequence of 91 nucleotides and a 3'-nontranslated region composed of a short poly(A) tract followed by a 238-nucleotide tRNA-like structure. The amino acid sequence of the polypeptide (alpha a) encoded by the open reading frame has homology with the TMV 126K protein and with related polypeptides from other viruses. The carboxy-terminal portion of the alpha a polypeptide also has limited homology with the 58K (beta b) protein encoded by BSMV RNA beta and includes a consensus sequence found in mononucleotide-binding polypeptides.     

Structural homologies between RNA gene segments 10 and 11 from UK bovine, simian SA11, and human Wa rotaviruses

The nucleotide sequences of gene segments 10 and 11 from UK bovine rotavirus have been determined. Gene 10 is 751 nucleotides long and contains a single long open reading frame capable of coding for a protein of 175 amino acids. When compared with the published data for gene 10 of the simian rotavirus SA11 and human Wa strains it was found to be more closely related to the SA11 structure (92% nucleotide sequence homology; 97% amino acid sequence homology) than to the human Wa structure (84% nucleotide, 86% amino acid sequence homology). All three strains have two potential N-glycosylation sites in the hydrophobic N terminus of the gene 10 protein. Gene 11 from UK bovine rotavirus is 667 nucleotides long with a single long open reading frame capable of coding for a protein of 198 amino acids. When compared with the published sequence of gene 11 from the human rotavirus Wa, the UK bovine rotavirus gene 11 was found to be one nucleotide longer in the 5'-noncoding region and three nucleotides longer in the coding region. The nucleotide sequence homology was 86%. The predicted proteins coded by segment 11 in UK and Wa rotaviruses are both rich in serine and threonine (23%) and very hydrophilic, but differ appreciably in amino acid sequence (83% homology).     

Conservation of a potential metal binding motif despite extensive sequence diversity in the rotavirus nonstructural protein NS53

The nucleotide sequence for the simian rotavirus SA11 gene segment 5 has been determined. The gene is 1611 nucleotides in length and contains a single open reading frame of 1485 nucleotides. The segment codes for the nonstructural protein NS53 which is predicted to be a polypeptide of 495 amino acids with a molecular weight of 58,484. When compared to the sequence of bovine RF gene segment 5 there are homologies of only 49 and 36% at the nucleotide and amino acid levels, respectively. This is in marked contrast to the situation with other rotavirus nonstructural proteins which are highly conserved between isolates. Nevertheless, there is a conserved region between amino acids 37-81 which contains a generalized motif for a metal binding domain. All eight cysteine and two histidine residues in this short sequence are conserved between the simian and bovine NS53 proteins. The conservation of this domain despite extensive sequence diversity in the remainder of the protein suggests that this region is functionally important.     

Sequences of the four larger proteins of a porcine group C rotavirus and comparison with the equivalent group A rotavirus proteins.

The sequences of the four larger proteins of rotavirus group C (Cowden strain) are presented and compared with the sequences of the corresponding group A proteins. They exhibit a significant level of homology, allowing gene coding assignment for the group C rotavirus. The coding strategy of the group C virus RNA segment is the same as that for the group A large segments as one long open reading frame is present in each segment. The genome segment 1 encodes the structural protein VP1 which presents the RNA-dependent RNA polymerase consensus motifs. The VP1 protein is the most highly conserved between the rotaviruses of groups A and C. The genome segment 2 encodes the VP2 protein. The deduced protein sequence does not present the putative leucine zippers identified in the group A protein but its amino terminal is hydrophilic and highly charged as previously noted for the group A protein. The genome segment 3 encodes for a protein homologous to the group A outer capsid protein VP4. As observed among the various group A sequences, the amino terminal is the region presenting the fewest similarities. A cleavage region and a putative fusion motif similar to those present in the group A viruses have been identified. For this protein the comparison has been extended to the IDIRV [corrected] VP3 previously sequenced and indicates that groups A and C VP4 proteins are much more related to each other than to the group B equivalent. The genome segment 4 encodes for a protein showing an approximate 40% sequence identity to the minor core protein, VP3, of the group A rotavirus. This remarkable conservation of primary structures argues for severe functional constraint on the evolution of these proteins.     

Organization of the middle RNA segment of snowshoe hare Bunyavirus

The genetic organization of the M RNA segment of snowshoe hare (SSH) virus, a member of the Bunyavirus genus of the family Bunyaviridae, has been determined. The middle (M) RNA segment has a single open reading frame (ORF) of 1441 amino acids. We have used amino- and carboxy-terminus sequencing and synthetic peptides to map proteins within the ORF. The order of the proteins translated from the single large open reading frame is G2, NSm, G1. The G2 protein extends from amino acids 14 to 299. The molecule is 286 residues long, with a computed nonglycosylated molecular weight of 31,973 Da. It is preceded by a cleaved 13 amino acid signal sequence. G2 includes a long highly hydrophobic sequence and contains three potential N-linked glycosylation sites. The G1 protein occupies the C-terminal end of the open reading frame from amino acids 474 to 1441 (968 amino acid residues) and has a computed nonglycosylated, molecular weight of 108,981 kDa. It has two potential N-linked glycosylation sites, and a potential transmembrane region followed by a potential cytoplasmic domain at the C-terminal end. If membrane associated it has an orientation of N-terminus outer, C-terminus inner. Limited trypsin digestion removes a 33-kDa fragment from the N-terminal end, leaving a virion-associated truncated G1 molecule (amino acids 762 to 1441) with a single N-linked glycosylation site. Between the G2 and G1 molecules there are 174 amino acids, sufficient to code for 19 kDa of protein. Some antibodies raised against peptides within this region react with proteins of 11 kDa (NSm) and 10 kDa present in infected cell lysates, but the exact relationship of these proteins to the open reading frame remains to be determined.     

Comparative sequence analyses of the cognate structural protein VP6 genes of five US bluetongue viruses.

The S3 segment (the small segment 3), encoding the structural protein, VP6, from the five United States (US) prototype bluetongue virus (BTV) serotypes were amplified by the Clamp-R method and cloned as full-length entities. The complete nucleotide sequence of each cognate gene segment was determined. Each cognate S3 segment of BTV-10, 11, 13 and 17 was 1049 nucleotides long and contained an open reading frame (ORF) capable of encoding a 325-amino acid protein. However, the S3 segment of BTV-2, which also contained 1049 nucleotides, had a longer 5'-non-coding region of 99-nucleotide and contained an ORF capable only of encoding a 301-amino acid protein. Comparative analyses of the predicted amino acid sequences of S3 segments of BTV-2, 10, 11, 13 and 17 revealed that VP6 was unusually high in glycine and contained few aromatic amino acids, but a high concentration of charged amino acids, which is a characteristic of a hydrophilic protein. Phylogenetic analyses indicated that BTV-11, 13 and 17 were more closely related than the other two US BTV serotypes. BTV-2 was the most distantly related.     

Sequence and expression of a gene from Theileria annulata coding for a 70-kilodalton heat-shock protein

A library consisting of randomly sheared Theileria annulata genomic DNA fragments in the vector lambda gt11 was screened with a Drosophila DNA segment containing the coding region of the 70-kDa heat-shock protein (hsp) gene. A positive recombinant was isolated and subjected to nucleotide sequence analysis. The DNA segment contains an open reading frame coding for a 71-kDa protein strongly homologous to hsp 70 from other organisms. Using this DNA as a probe, a homologous 2.5-kb RNA species was detected in sporozoites, piroplasms and a macroschizont-infected cell line, showing that this gene is constitutively expressed. The amount of this RNA increased following heat shock in the macroschizont-infected cell line. The T. annulata genome contains other sequences that hybridize weakly with this heat-shock gene.     

Nucleotide sequence of RNA segment 7 of influenza B/Singapore/222/79: maintenance of a second large open reading frame

The nucleotide sequence of RNA segment 7 of influenza B/Singapore/222/79 has been determined. The deduced amino acid sequence of the M1 protein indicates that it contains 248 amino acids of which 243 are identical in the B/Lee/40 M1 protein. A second overlapping open reading frame with a maximum coding region of 195 amino acids has also been maintained in the 39 years separating the two isolates. The deduced amino acid sequence of the second coding region displays 86% homology with the comparable B/Lee/40 sequence (27 changes). The conservation of the second open reading frame suggests that this region is important in the replicative cycle of influenza B viruses.     

A subtype of human papillomavirus 5 (HPV-5b) and its subgenomic segment amplified in a carcinoma: nucleotide sequences and genomic organizations.

A subtype of human papillomavirus 5 (HPV-5b) is closely associated with carcinomas in the disease epidermodysplasia verruciformis (EV). The complete genome was cloned from virus particles in benign lesions of a patient with EV and sequenced: it was 7779 nucleotides long and consisted of six open reading frames (ORFs) (E6, E7, E1, E2, E4, and E5) in the early region, three ORFs (L2, L3, and L1) in the late region, and a noncoding region, all existing on one DNA strand. The 40% segment of the HPV-5b genome specifically amplified in carcinomas was cloned from a primary carcinoma of the same EV patient and sequenced: it was 3143 nucleotides long and corresponded to a segment of the original HPV-5b genome containing the entire sequences of E6, E7, and the noncoding region and portions of E1 and L1. Compared to the whole genomic DNA, no mutations were detected in this probable malignancy-associated viral subgenomic segment cloned from carcinoma. These results suggest that amplification of the viral segment containing E6, E7, and the noncoding region may play a role in the malignant conversion of HPV-5b-infected benign lesions and that mutations in these genes or regions are not necessarily required.     

A long open reading frame in the mitochondrial LSU rRNA group-I intron of Cryphonectria parasitica encodes a putative S5 ribosomal protein fused to a maturase

A 4238-bp intervening sequence within the highly conserved U11 region of the mitochondrial large subunit ribosomal RNA gene of the fungus Cryphonectria parasitica Ep155 has been sequenced and identified to be a group-I intron. This is the largest group-I intron reported to-date for fungal mitochondrial genomes. The intron contains an 851-codon open reading frame encoding a putative, but complete, small-subunit ribosomal protein of 510 amino acids which is fused at its carboxyl terminus to a 311 amino-acid polypeptide representing a typical maturase-like protein. A short open reading frame of 83 amino acids with some similarity to maturases, but lacking a translation-initiation codon, was also noted at the 3′ end of the intron. The unusual size of the intron and the arrangement of the open and truncated reading frames suggest that this segment of the mtDNA of C. parasitica has arisen by a fusion of components from two or more different introns, possibly involving the re-location of intronic genes. Received: 7 August / 15 December 1998     

Human papillomavirus type 16 DNA sequence

The complete nucleotide sequence of HPV16 DNA (7904 bp) cloned from an invasive cervical carcinoma was determined. Homology comparisons allowed us to align the major open reading frames with the other published papilloma virus DNA sequences. The general organization of the open reading frames is similar to that of the other four papillomavirus (BPV1, HPV1a, HPV6b, CRPV) already sequenced. The sequence reveals an interruption of the reading frame coding for a suspected E1 protein.