The complete nucleotide sequence of a common cold virus: human rhinovirus 14.

The complete nucleotide sequence of the single-stranded RNA genome of human rhinovirus 14, one of the causative agents of the common cold, has been determined from cDNA cloned in E. coli. The genome is typical of the picornaviridae family, comprising a 5' non-coding region of 624 nucleotides, a long open reading frame of 6537 nucleotides (90.8% of the genome) and a 3' non-coding region of 47 nucleotides. Comparison of the nucleotide sequence and the predicted amino acid sequence with those of the polioviruses reveals a surprising degree of homology which may allow recognition of regions of antigenic importance and prediction of the virus polyprotein cleavage sites. The results presented here imply a closer genetic relationship between the rhinovirus and enterovirus genera than previously suspected.     

Partial nucleotide sequence of the Murray Valley encephalitis virus genome. Comparison of the encoded polypeptides with yellow fever virus structural and non-structural proteins

The sequence of 5400 bases corresponding to the 5'-terminal half of the Murray Valley encephalitis virus genome has been determined. The genome contains a 5' non-coding region of about 97 nucleotides, followed by a single continuous open reading frame that encodes the structural proteins followed by the non-structural proteins. Amino acid sequence homology between the Murray Valley encephalitis and yellow fever (Rice et al., 1985) polyproteins is 42% over the region sequenced. The start points of the various Murray Valley encephalitis virus-coded proteins have been assigned on the basis of this homology and a consistent set of potential proteolytic cleavage sites identified, the sequences of which are similar in Murray Valley encephalitis and yellow fever. The deduced Murray Valley encephalitis gene order is 5'-C-prM (M)-E-NS1-ns2a-ns2b-NS3-3'. The genome organization of Murray Valley encephalitis and yellow fever appears to be identical and the sizes of the predicted virus-coded proteins similar between the two viruses. Both viruses encode a basic capsid protein followed by three glycoproteins; the glycoproteins appear to have the conventional topology of N terminus outside with a C-terminal membrane-spanning domain. There are conserved glycosylation sites in prM, the precursor to the M protein of the virion, and in NS1, a non-structural protein of uncertain function. The glycosylation sites in E, the major envelope protein of the virion, are not conserved as to position. We predict the existence, in flavivirus-infected cells, of two small, hydrophobic peptides, ns2a and ns2b, which show only limited amino acid sequence homology. Finally, about half of the amino acid sequence of NS3 has been obtained; NS3 is a hydrophilic non-structural protein that shows 55% amino acid sequence similarity between Murray Valley encephalitis and yellow fever over the region sequenced and is probably involved in RNA replication.     

Parvovirus genome: nucleotide sequence of H-1 and mapping of its genes by hybrid-arrested translation.

The nucleotide sequence of the parvovirus H-1 has been determined by the chain-terminating method of Sanger. The sequence is 5,176 nucleotides long. Two large open reading frames (1 and 2) and two smaller open reading frames (3 and 4) of potential importance were identified in the plus-strand sequence. Promoter sequences are located at map positions 4 and 38 when map positions are expressed as percent of genome length from the 3' end of the virion minus strand. The locations for the genes for the parvovirus capsid proteins and a 76,000-dalton noncapsid protein (NCVP1) were mapped by hybrid-arrested translation. The gene for the capsid proteins VP1 and VP2' is located in the 5' half of the virus genome. The gene for NCVP1 is located in the 3' half of the viral DNA.     

Nucleotide sequence and genome organization of foot-and-mouth disease virus.

A continuous 7802 nucleotide sequence spanning the 94% of foot and mouth disease virus RNA between the 5'-proximal poly(C) tract and the 3'-terminal poly(A) was obtained from cloned cDNA, and the total size of the RNA genome was corrected to 8450 nucleotides. A long open reading frame was identified within this sequence starting about 1300 bases from the 5' end of the RNA genome and extending to a termination codon 92 bases from its polyadenylated 3' end. The protein sequence of 2332 amino acids deduced from this coding sequence was correlated with the 260 K FMDV polyprotein. Its processing sites and twelve mature viral proteins were inferred from protein data, available for some proteins, a predicted cleavage specificity of an FMDV encoded protease for Glu/Gly(Thr, Ser) linkages, and homologies to related proteins from poliovirus. In addition, a short unlinked reading frame of 92 codons has been identified by sequence homology to the polyprotein initiation signal and by in vitro translation studies.     

The complete nucleotide sequence of the RNA coding for the primary translation product of foot and mouth disease virus.

The complete nucleotide sequence of the coding region of foot and mouth disease virus RNA (strain A1061) is presented. The sequence extends from the primary initiation site, approximately 1200 nucleotide from the 5' end of the genome, in an open translational reading frame of 6,999 nucleotides to a termination codon 93 nucleotides from the 3' terminal poly (A). Available amino acid sequence data correlates with that predicted from the nucleotide sequence. The amino acid sequence around cleavage sites in the polyprotein shows no consistency, although a number of the virus-coded protease cleavage sites are between glutamate and glycine residues.     

Capsid coding sequence is required for efficient replication of human rhinovirus 14 RNA.

Mechanisms by which the plus-sense RNA genomes of picornaviruses are replicated remain poorly defined, but existing models do not suggest a role for sequences encoding the capsid proteins. However, candidate RNA replicons (delta P1 beta gal and delta P1Luc), representing the sequence of human rhinovirus 14 virus (HRV-14) with reporter protein sequences (beta-galactosidase or luciferase, respectively) replacing most of the P1 capsid-coding region, failed to replicate in transfected H1-HeLa cells despite efficient primary cleavage of the polyprotein. To determine which P1 sequences might be required for RNA replication, HRV-14 mutants in which segments of the P1 region were removed to frame from the genome were constructed. Mutants with deletions involving the 5'proximal 1,489 nucleotides of the P1 region replicated efficiently, while those with deletions involving the 3' 1,079 nucleotides did not. Reintroduction of the 3' P1 sequence into the nonreplicating delta P1Luc construct resulted in a new candidate replicon, delta P1Luc/VP3, which replicated well and expressed luciferase efficiently. Capsid proteins provided in trans by helper virus failed to rescue the nonreplicating delta P1Luc genome but were able to package the larger-than-genome-length delta P1Luc/VP3 replicon. Thus, a 3'-distal P1 capsid-coding sequence has a previously unrecognized cis-active function related to replication of HRV-14 RNA.     

The nucleotide sequence of tobacco rattle virus RNA-2 (CAM strain).

The nucleotide sequence of the smaller genomic strand (RNA-2) of the bipartite tobacco rattle virus (CAM strain) has been determined. RNA-2 is capped at the 5' terminus and contains 1799 nucleotide residues. There is a single 223 codon long open reading frame extending from nucleotide 574 to 1242 which designates a protein of Mr 23,654. The derived amino acid composition, in percent, matches that previously determined for the virus capsid protein. The long open reading frame is flanked by 5' and 3' untranslated regions of 573 and 554 nucleotides, respectively. The 5' leader sequence contains two different sets of direct repeats, one of 119 nucleotides and the other of 76. It also contains 13 apparently unused AUG codons, four of which lie in the same frame as the capsid protein cistron. The 3' terminal sequence of RNA-2 is identical to that of the larger genomic strand (RNA-1) for 459 nucleotides.     

The nucleotide sequence of poliovirus type 3 leon 12 a1b: comparison with poliovirus type 1.

The complete nucleotide sequence of the genome of the Sabin vaccine strain of poliovirus type 3 (P3/Leon 12 a1 b) has been determined from cDNA cloned in E. coli. The genome comprises a 5' non-coding region of 742 nucleotides, a large open reading frame of 6618 nucleotides (89% of the sequence) and a 3' non-coding region of 72 nucleotides. There is 77.4% base-sequence homology and 89.6% predicted amino-acid homology between types 1 and 3. Conservation of all glutamine-glycine and tyrosine-glycine cleavage sites suggests a mechanism of polyprotein processing similar to that established for poliovirus type 1.     

Amino acid sequence of human respiratory syncytial virus nucleocapsid protein.

Amino acid sequence of the human respiratory syncytial (RS) virus nucleocapsid (NC) protein, deduced from the DNA sequence of a recombinant plasmid, is presented. The cDNA plasmid (pRSB11) has 1412 bp of RS viral NC sequence and lacks six nucleotides of the 5' end of mRNA. There is a single long open reading frame encoding 467 amino acids. This 51540 dal protein is rich in basic amino acids and has no homologies with other known viral capsid proteins.     

Identification of the initiation site of poliovirus polyprotein synthesis.

The complete nucleotide sequence of poliovirus RNA has a long open reading frame capable of encoding the precursor polyprotein NCVP00. The first AUG codon in this reading frame is located 743 nucleotides from the 5' end of the RNA and is preceded by eight AUG codons in all three reading frames. Because all proteins that map at the amino terminus of the polyprotein (P1-1a, VP0, and VP4) are blocked at their amino termini and previous studies of ribosome binding have been inconclusive, direct identification of the initiation site of protein synthesis was difficult. We separated and identified all of the tryptic peptides of capsid protein VP4 and correlated these peptides with the amino acid sequence predicted to follow the AUG codon at nucleotide 743. Our data indicate that VP4 begins with a blocked glycine that is encoded immediately after the AUG codon at nucleotide 743. An S1 nuclease analysis of poliovirus mRNA failed to reveal a splice in the 5' region. We concluded that synthesis of the poliovirus polyprotein is initiated at nucleotide 743, the first AUG codon in the long open reading frame.     

Sequence of 3,687 nucleotides from the 3' end of Sendai virus genome RNA and the predicted amino acid sequences of viral NP, P and C proteins.

The sequence of 3,687 nucleotides from the 3' end of the Sendai virus genome (Z strain) was determined by a molecular cloning technique followed by rapid sequence analysis. Two large open reading frames, one consisting of 1,572 nucleotides and the other of 1,704 nucleotides, were observed in the region, that is OP-1 and OP-2 from the 3' end of the genome. The amino acid sequences of the gene products were predicted from the observed sequence. Determination of amino acid compositions of viral proteins, P, HN, Fo, NP and M, led us to conclude that NP and P are the gene products of OP-1 and OP-2, respectively. An additional open reading frame consisting of 612 nucleotides (OP-3) was discovered in the 3' most proximal region of OP-2. The predicted product of OP-3 was considered to be viral non-structural protein C. The leader sequence of 51 nucleotides at the 3' terminal of the genome and consensus sequences at 3' and 5' ends of each gene for proteins NP and P were identified.     

Nucleotide sequence and genome organization of canine parvovirus.

The genome of a canine parvovirus isolate strain (CPV-N) was cloned, and the DNA sequence was determined. The entire genome, including ends, was 5,323 nucleotides in length. The terminal repeat at the 3' end of the genome shared similar structural characteristics but limited homology with the rodent parvoviruses. The 5' terminal repeat was not detected in any of the clones. Instead, a region of DNA starting near the capsid gene stop codon and extending 248 base pairs into the coding region had been duplicated and inserted 75 base pairs downstream from the poly(A) addition site. Consensus sequences for the 5' donor and 3' acceptor sites as well as promotors and poly(A) addition sites were identified and compared with the available information on related parvoviruses. The genomic organization of CPV-N is similar to that of feline parvovirus (FPV) in that there are two major open reading frames (668 and 722 amino acids) in the plus strand (mRNA polarity). Both coding domains are in the same frame, and no significant open reading frames were apparent in any of the other frames of both minus and plus DNA strands. The nucleotide and amino acid homologies of the capsid genes between CPV-N and FPV were 98 and 99%, respectively. In contrast, the nucleotide and amino acid homologies of the capsid genes for CPV-N and CPV-b (S. Rhode III, J. Virol. 54:630-633, 1985) were 95 and 98%, respectively. These results indicate that very few nucleotide or amino acid changes differentiate the antigenic and host range specificity of FPV and CPV.     

Molecular and biological characterization of deformed wing virus of honeybees (Apis mellifera L.)

Deformed wing virus (DWV) of honeybees (Apis mellifera) is closely associated with characteristic wing deformities, abdominal bloating, paralysis, and rapid mortality of emerging adult bees. The virus was purified from diseased insects, and its genome was cloned and sequenced. The genomic RNA of DWV is 10,140 nucleotides in length and contains a single large open reading frame encoding a 328-kDa polyprotein. The coding sequence is flanked by a 1,144-nucleotide 5' nontranslated leader sequence and a 317-nucleotide 3' nontranslated region, followed by a poly(A) tail. The three major structural proteins, VP1 (44 kDa), VP2 (32 kDa), and VP3 (28 kDa), were identified, and their genes were mapped to the N-terminal section of the polyprotein. The C-terminal part of the polyprotein contains sequence motifs typical of well-characterized picornavirus nonstructural proteins: an RNA helicase, a chymotrypsin-like 3C protease, and an RNA-dependent RNA polymerase. The genome organization, capsid morphology, and sequence comparison data indicate that DWV is a member of the recently established genus Iflavirus.     

Complete nucleotide sequences of all three poliovirus serotype genomes. Implication for genetic relationship, gene function and antigenic determinants

The complete nucleotide sequences of the genomes of the type 2 ( P712 , Ch, 2ab ) and type 3 (Leon 12a1b ) poliovirus vaccine strains were determined. Comparison of the sequences with the previously established genome sequence of type 1 (LS-c, 2ab ) poliovirus vaccine strain revealed that 71% of the nucleotides in the genome RNAs were common, that the 5' and 3' termini of the genomes were highly homologous, and that more than 80% of the nucleotide differences in the coding region occurred in the third letter position of in-phase codons, resulting in a low frequency of amino acid difference. These results strongly suggested that the serotypes of poliovirus derived from a common prototype. A comparison of the amino acid sequences predicted from the genome sequences showed highest variation in the capsid protein region, whereas non-structural proteins are highly conserved. Initiation of polyprotein synthesis occurs in all three strains more than 740 nucleotides downstream from the 5' end. An analysis of the non-coding region suggests that small peptides that could potentially originate from this region are conserved. The amino acid sequences immediately surrounding the cleavage signals, however, show a higher than average degree of variation. The analysis of the amino acid sequences of the capsid protein VP1 of all serotypes has led to the prediction of potential antigenic sites on the virion involved in neutralization.     

Kelp fly virus: a novel group of insect picorna-like viruses as defined by genome sequence analysis and a distinctive virion structure

The complete genomic sequence of kelp fly virus (KFV), originally isolated from the kelp fly, Chaetocoelopa sydneyensis, has been determined. Analyses of its genomic and structural organization and phylogeny show that it belongs to a hitherto undescribed group within the picorna-like virus superfamily. The single-stranded genomic RNA of KFV is 11,035 nucleotides in length and contains a single large open reading frame encoding a polypeptide of 3,436 amino acids with 5' and 3' untranslated regions of 384 and 343 nucleotides, respectively. The predicted amino acid sequence of the polypeptide shows that it has three regions. The N-terminal region contains sequences homologous to the baculoviral inhibitor of apoptosis repeat domain, an inhibitor of apoptosis commonly found in animals and in viruses with double-stranded DNA genomes. The second region contains at least two capsid proteins. The third region has three sequence motifs characteristic of replicase proteins of many plant and animal viruses, including a helicase, a 3C chymotrypsin-like protease, and an RNA-dependent RNA polymerase. Phylogenetic analysis of the replicase motifs shows that KFV forms a distinct and distant taxon within the picorna-like virus superfamily. Cryoelectron microscopy and image reconstruction of KFV to a resolution of 15 A reveals an icosahedral structure, with each of its 12 fivefold vertices forming a turret from the otherwise smooth surface of the 20-A-thick capsid. The architecture of the KFV capsid is unique among the members of the picornavirus superfamily for which structures have previously been determined.     

The nucleotide and deduced amino acid sequences of the encephalomyocarditis viral polyprotein coding region.

The nucleotide sequence of 7200 bases of encephalomyocarditis (EMC) viral RNA, including the complete polyprotein-coding region, was determined. The polyprotein is encoded within a unique translational reading frame, 6870 bases in length. Protein synthesis begins with the sequence Met-Ala-Thr, and ends with the sequence Leu-Phe-Trp, 126 bases from the 3' end of the RNA. Viral capsid and noncapsid proteins were aligned with the deduced amino acid sequence of the polyprotein. The proteolytic processing map follows the standard 4-3-4 picornaviral pattern except for a short leader peptide (8 kd), which precedes the capsid proteins. Identification of the proteolytic cleavage sites showed that EMC viral protease, p22, has cleavage specificity for gln-gly or gln-ser sequences with adjacent proline residues. The cleavage specificity of the host-coded protease(s) includes both tyr-pro and gln-gly sequences.