Nucleotide sequence and structural features of a novel US-a junction present in a defective herpes simplex virus genome.

Defective genomes generated during serial propagation of herpes simplex virus type 1 (Justin) consist of tandem reiterations of sequences that are colinear with a portion of the S component of the standard viral genome. We determined the structure of the novel US-a junction, at which the US sequences of one repeat unit join the a sequences of the adjacent repeat unit. Comparison of the nucleotide sequence at this junction with the nucleotide sequence of the corresponding US region of the standard virus genome indicated that the defective genome repeat unit arose by a single recombinational event between an L-S junction a sequence and the US region. The recombinational process might have been mediated by limited sequence homology. The sequences retained within the US-a junction further define the signal for cleavage and packaging of 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.     

Nucleotide sequence stability of the genome of hepatitis delta virus.

Cultured cells were cotransfected with a fully sequenced 1,679-base cDNA clone of human hepatitis delta virus (HDV) RNA genome and a cDNA for the genome of woodchuck hepatitis virus (WHV). The HDV particles released were able to infect a woodchuck that was chronically infected with WHV. The HDV so produced was passaged a total of six times in woodchucks in order to determine the stability of the HDV nucleotide sequence. During a final chronic infection with such virus, liver RNA was extracted, and the HDV nucleotide sequence for the 352-base region, positions 905 to 1256, was obtained. By means of PCR, we obtained double-stranded cDNA both for direct sequencing and also for molecular cloning followed by sequencing. By direct sequencing, we found that a consensus sequence existed and was identical to the original sequence. From the sequences of 31 clones, we found 32% (10 of 31) to be identical to the original single nucleotide sequence. For the remainder, there were neither insertions nor deletions but there was a small number of single-nucleotide changes. These changes were predominantly transitions rather than transversions. Furthermore, the transitions were largely of just two types, uridine to cytidine and adenosine to guanosine. Of the 40 changes detected on HDV, 35% (14 of 40) occurred within an eight-nucleotide region that included position 1012, previously shown to be a site of RNA editing. These findings may have significant implications regarding both the stability of the HDV RNA genome and the mechanism of RNA editing.     

Nucleotide sequence of the bovine parainfluenza 3 virus genome: the genes of the F and HN glycoproteins.

By analysing complementary DNA clones constructed from genomic RNA of bovine parainfluenza 3 virus (BPIV3), we determined the nucleotide sequence of the region containing the entire F and HN genes. Their deduced amino acid sequences showed about 80% homologies with those of human parainfluenza 3 virus (HPIV3), about 45% with those of Sendai virus, and about 20% with those of SV5 and Newcastle disease virus (NDV), indicating, together with the results described in the preceding paper on the NP, P, C and M proteins of BPIV3, that BPIV3, HPIV3 and Sendai virus constitute a paramyxovirus subgroup, and that BPIV3 and HPIV3 are very closely related. The F and HN proteins of all these viruses, including SV5 and NDV, however, were shown to have protein-specific structures as well as short but well-conserved amino acid sequences, suggesting that these structures and sequences are related to the activities of these glycoproteins.     

Complete nucleotide sequence of the nonstructural protein genes of Semliki Forest virus.

The nucleotide sequence coding for the nonstructural proteins of Semliki Forest virus has been determined from cDNA clones. The total length of this region is 7381 nucleotides, it contains an open reading frame starting at position 86 and ending at an UAA stop codon at position 7379-7381. This open reading frame codes for a 2431 amino acids long polyprotein, from which the individual nonstructural proteins are formed by proteolytic processing steps, so that nsPl is 537, nsP2 798, nsP3 482 and nsP4 614 amino acids. In the closely related Sindbis and Middelburg viruses there is an opal stop codon (UGA) between the genes for nsP3 and nsP4. Interestingly, no stop codon is found in frame in this region of the Semliki Forest virus 42S RNA. In other aspects the amino acid sequence homology between Sindbis, Middelburg and Semliki Forest virus nonstructural proteins is highly significant.     

Nucleotide sequence of the gag gene and gag-pol junction of feline leukemia virus.

The nucleotide sequence of the gag gene of feline leukemia virus and its flanking sequences were determined and compared with the corresponding sequences of two strains of feline sarcoma virus and with that of the Moloney strain of murine leukemia virus. A high degree of nucleotide sequence homology between the feline leukemia virus and murine leukemia virus gag genes was observed, suggesting that retroviruses of domestic cats and laboratory mice have a common, proximal evolutionary progenitor. The predicted structure of the complete feline leukemia virus gag gene precursor suggests that the translation of nonglycosylated and glycosylated gag gene polypeptides is initiated at two different AUG codons. These initiator codons fall in the same reading frame and are separated by a 222-base-pair segment which encodes an amino terminal signal peptide. The nucleotide sequence predicts the order of amino acids in each of the individual gag-coded proteins (p15, p12, p30, p10), all of which derive from the gag gene precursor. Stable stem-and-loop secondary structures are proposed for two regions of viral RNA. The first falls within sequences at the 5' end of the viral genome, together with adjacent palindromic sequences which may play a role in dimer linkage of RNA subunits. The second includes coding sequences at the gag-pol junction and is proposed to be involved in translation of the pol gene product. Sequence analysis of the latter region shows that the gag and pol genes are translated in different reading frames. Classical consensus splice donor and acceptor sequences could not be localized to regions which would permit synthesis of the expected gag-pol precursor protein. Alternatively, we suggest that the pol gene product (RNA-dependent DNA polymerase) could be translated by a frameshift suppressing mechanism which could involve cleavage modification of stems and loops in a manner similar to that observed in tRNA processing.     

Nucleotide sequence of a cloned woodchuck hepatitis virus genome: evolutional relationship between hepadnaviruses.

We have determined the complete nucleotide sequence of a cloned DNA of woodchuck hepatitis virus (WHV), the most oncogenic virus among hepadnaviruses. The genome, designated WHV2, is 3,320 base pairs long and contains four major open reading frames (ORFs) coded on the same strand of nucleotide sequence as in the human hepatitis B virus (HBV) genome. Comparison of the nucleotide sequence and amino acid sequences deduced from it among the genomes of various hepadnaviruses demonstrates that each protein shows an intrinsic property in conserving its amino acid sequence. A parameter, the ratio of the number of triplets with one-letter change but no amino acid substitution to the total number of triplets in which one-letter change occurred, was introduced to measure the intrinsic properties quantitatively. For each ORF, the parameter gave characteristic values in all combinations. Therefore, the relative evolutional distance between these hepadnaviruses can be measured by the amino acid substitution rate of any ORF. These comparisons suggest that (i) the difference between two WHV clones, WHV1 and WHV2, corresponds to that among clones of a HBV subtype, HBVadr, and (ii) WHV and ground squirrel hepatitis virus can be categorized in a way similar to the subgroups of HBV.     

Nucleotide sequence of the ipaBCD structural genes of Shigella dysenteriae

A 9 kb EcoRI and two PstI fragments from the virulence plasmid of Shigella dysenteriae CG097 were shown to contain all ipa genes by probing with Shigella flexneri ipaB, -C, -D and -A gene probes. The DNA sequences of S. dysenteriae ipaBC genes were very similar to those of S. flexneri M90T and S. flexneri YSH6000, but ipaD differed by 22 codons from that of S. flexneri. The differences in ipaD may account for the different in vitro host specificities shown by S. dysenteriae and S. flexneri. The nucleotide composition of ipa genes revealed an unusually large number of codons that are rarely used in Escherichia coli chromosomal genes, indicating a different origin.     

Nucleotide sequence of a cloned woodchuck hepatitis virus genome: comparison with the hepatitis B virus sequence.

The complete nucleotide sequence of a woodchuck hepatitis virus genome cloned in Escherichia coli was determined by the method of Maxam and Gilbert. This sequence was found to be 3,308 nucleotides long. Potential ATG initiator triplets and nonsense codons were identified and used to locate regions with a substantial coding capacity. A striking similarity was observed between the organization of human hepatitis B virus and woodchuck hepatitis virus. Nucleotide sequences of these open regions in the woodchuck virus were compared with corresponding regions present in hepatitis B virus. This allowed the location of four viral genes on the L strand and indicated the absence of protein coded by the S strand. Evolution rates of the various parts of the genome as well as of the four different proteins coded by hepatitis B virus and woodchuck hepatitis virus were compared. These results indicated that: (i) the core protein has evolved slightly less rapidly than the other proteins; and (ii) when a region of DNA codes for two different proteins, there is less freedom for the DNA to evolve and, moreover, one of the proteins can evolve more rapidly than the other. A hairpin structure, very well conserved in the two genomes, was located in the only region devoid of coding function, suggesting the location of the origin of replication of the viral DNA.     

Nucleotide sequence of human endogenous retrovirus genome related to the mouse mammary tumor virus genome.

We determined the complete nucleotide sequence of the human endogenous retrovirus genome HERV-K10 isolated as the sequence homologous to the Syrian hamster intracisternal A-particle (type A retrovirus) genome. HERV-K10 is 9,179 base pairs long with long terminal repeats of 968 base pairs at both ends; a sequence 290 base pairs long, however, was found to be deleted. It was concluded that a composite genome having the 290-base-pair fragment is the prototype HERV-K provirus gag (666 codons), protease (334 codons), pol (937 codons), and env (618 codons) genes. The size of the protease gene product of HERV-K is essentially the same as that of A- and D-type oncoviruses but nearly twice that of other retroviruses. A comparison of the deduced amino acid sequences encoded by the pol region showed HERV-K to be closely related to types A and D retroviruses and even more so to type B retrovirus. It was noted that the env gene product of HERV-K structurally resembles the mouse mammary tumor virus (type B retrovirus) env protein, and the possible expression of the HERV-K env gene in human breast cancer cells is discussed.