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At least 1,009 records · Page 56Linked to original sources
PubMed2005

Murine gammaherpesvirus 68 open reading frame 45 plays an essential role during the immediate-early phase of viral replication.

Murine gammaherpesvirus 68 (MHV-68) has been developed as a model for the human gammaherpesviruses Epstein-Barr virus and human herpesvirus 8/Kaposi's sarcoma-associated herpesvirus (HHV-8/KSHV), which are associated with several types of human diseases. Open reading frame 45 (ORF45) is conserved among the members of the Gammaherpesvirinae subfamily and has been suggested to be a virion tegument protein. The repression of ORF45 expression by small interfering RNAs inhibits MHV-68 viral replication. However, the gene product of MHV-68 ORF45 and its function have not yet been well characterized. In this report, we show that MHV-68 ORF45 is a phosphorylated nuclear protein. We constructed an ORF45-null MHV-68 mutant virus (45STOP) by the insertion of translation termination codons into the portion of the gene encoding the N terminus of ORF45. We demonstrated that the ORF45 protein is essential for viral gene expression immediately after the viral genome enters the nucleus. These defects in viral replication were rescued by providing ORF45 in trans or in an ORF45-null revertant (45STOP.R) virus. Using a transcomplementation assay, we showed that the function of ORF45 in viral replication is conserved with that of its KSHV homologue. Finally, we found that the C-terminal 23 amino acids that are highly conserved among the Gammaherpesvirinae subfamily are critical for the function of ORF45 in viral replication.

Amino Acid Sequence↗
PubMed1995

Four dually resistant human cytomegalovirus strains from AIDS patients: single mutations in UL97 and UL54 open reading frames are responsible for ganciclovir- and foscarnet-specific resistance, respectively.

Four human cytomegalovirus strains with double resistance to both ganciclovir and foscarnet were recovered from acquired immunodeficiency syndrome (AIDS) patients. The four isolates were genetically unrelated and each of them consisted of a single viral population. In addition, for each isolate, UL97 and UL54 open reading frames sequencing showed a single aminoacid change in a conserved domain of each of the two genes.

AIDS-Related Opportunistic Infections↗
PubMed1987

A yeast nuclear gene, MRS1, involved in mitochondrial RNA splicing: nucleotide sequence and mutational analysis of two overlapping open reading frames on opposite strands.

We have cloned a 1.6-kb fragment of yeast nuclear DNA, which complements pet- mutant MK3 (mrs1). This mutant was shown to be defective in mitochondrial RNA splicing: the excision of intron 3 from the mitochondrial COB pre-RNA is blocked. The DNA sequence of the nuclear DNA fragment revealed two open reading frames (ORF1 with 1092 bp; ORF2 with 735 bp) on opposite strands, which overlap by 656 bp. As shown by in vitro mutagenesis, ORF1, but not ORF2, is responsible for complementation of the splice defect. Hence, ORF1 represents the nuclear MRS1 gene. Disruption of the gene (both ORFs) in the chromosomal DNA of the respiratory competent yeast strain DBY747 (long form COB gene) leads to a stable pet- phenotype and to the accumulation of the same mitochondrial RNA precursors as in strain MK3. The amino acid sequence of the putative ORF1 product does not exhibit any homology with other known proteins, except for a small region of homology with the gene product of another nuclear yeast gene involved in mitochondrial RNA splicing, CBP2. The function of the MRS1 (ORF1) gene in mitochondrial RNA splicing and the significance of the overlapping ORFs in this gene are discussed.

Base Sequence↗
PubMed1985

Subcellular localization of the product of the long open reading frame of human T-cell leukemia virus type I.

Human T-cell leukemia virus type I (HTLV-I) is a retrovirus associated with adult T-cell leukemia and lymphoma. In addition to containing the gag, pol, and env genes of the chronic leukemia viruses, the genome of HTLV-I contains a long open reading frame (LOR) located between the 3' end of the envelope gene and the 3' long terminal repeat sequence (LTR). It has been suggested that a protein of 42 kilodaltons that is encoded by the LOR region may participate in both trans-acting transcriptional regulation of the viral LTR as well as in the transforming properties of HTLV-I. It is reported here that a significant fraction of the 42-kilodalton HTLV LOR product is located in the nucleus of HTLV-I-infected transformed lymphocytes, a finding that is consistent with its proposed functions.

Cell Fractionation↗
PubMed1988

Accumulation of RNA homologous to human papillomavirus type 16 open reading frames in genital precancers.

The accumulation of human papillomavirus type 16 (HPV-16)-specific RNAs in tissue sections from biopsies of patients with genital precancers was studied by in situ hybridization with single-stranded 35S-labeled RNA. These analyses revealed that the most abundant early-region RNAs were derived from the E4 and E5 open reading frames (ORFs). RNAs homologous to the E6/E7 ORFs were also detected, whereas RNAs homologous to the intervening E1 ORF were not. This suggests that the E4 and E5 mRNAs are derived by splicing to the upstream E6/E7 ORFs, consistent with studies of HPV-11 in condylomata (L. T. Chow et al., Cancer Cells (Cold Spring Harbor) 5:55-72, 1987). Abundant RNAs homologous to the 5' portion of L1 were also detected. These RNAs were localized to the apical strata of the epithelium. HPV-16 RNAs accumulated in discrete regions of these lesions, and when present were most abundant in the upper cell layers of the precancerous epithelium. RNAs homologous to early ORFs were also detected in some germinal cells within the basal layer of the epithelium.

Epithelium↗
PubMed1989

Leader-encoded open reading frames modulate both the absolute and relative rates of synthesis of the virion proteins of simian virus 40.

Numerous viral and cellular RNAs are polycistronic, including several of the late mRNA species encoded by simian virus 40 (SV40). The functionally bicistronic major late 16S and functionally tricistronic major late 19S mRNA species of SV40 contain the leader-encoded open reading frames (ORFs) LP1, located upstream of the sequence encoding the virion protein VP1, and LP1*, located upstream of the sequence encoding the virion proteins VP2 and VP3. To determine how these leader ORFs affect synthesis of the virion proteins, monkey cells were transfected with viral mutants in which either the leader-encoded translation initiation signal was mutated or the length and overlap of the leader ORF relative to the ORFs encoding the virion proteins were altered. The levels of initiation at and leaky scanning past each initiation signal were determined directly by quantitative analysis of the viral proteins synthesized in cells transfected with these mutants. Novel findings from these experiments included the following. (i) At least one-third of ribosomes bypass the leader-encoded translation initiation signal, GCCAUGG, on the SV40 major late 16S mRNA. (ii) At least 20% of ribosomes bypass even the consensus translation initiation signal, ACCAUGG, when it is situated 10 bases from the 5' end on the major late 16S mRNA. (iii)O The presence of the leader ORF on the bicistronic 16S mRNA species reduces VP1 synthesis threefold relative to synthesis from a similar RNA that lacks it. (iv) At least half and possibly all VP1 synthesized from the bicistronic 16S mRNA species is made by a leaky scanning mechanism. (v) LP1 and VP1 are synthesized from the bicistronic 16S mRNA species at approximately equal molar ratios. (vi) Approximately half of the VP1 synthesized in SV40-infected cells is synthesized from the minor, monocistronic 16S mRNA even though it accounts for only 20% of the 16S mRNA present. (vii) The presence and site of termination of translation of the leader ORF on the late 19S mRNAs affect the relative as well as absolute rates of synthesis of VP2 and VP3.

Genes, Viral↗
PubMed1988

Overlapping open reading frames revealed by complete nucleotide sequencing of turnip yellow mosaic virus genomic RNA.

The complete nucleotide sequence of turnip yellow mosaic virus (TYMV) genomic RNA has been determined on a set of overlapping cDNA clones using a sequential sequencing strategy. The RNA is 6318 nucleotides long, excluding the cap structure. The genome organization deduced from the sequence confirms previous results of in vitro translation. A novel open reading frame (ORF) putatively encoding a Pro-rich and very basic 69K (K = kilodalton) protein is detected at the 5' end of the genome. It is initiated at the first AUG codon on the RNA and overlaps the major ORF that encodes the non structural 206K (previously referred to as 195K) protein of TYMV; its function is unknown. Several amino acid consensus sequences already described among plant and animal viruses are also found in the TYMV-encoded polypeptides. A comparison with other viruses whose RNA sequence is known leads to the conclusion that TYMV belongs to the "Sindbis-like" supergroup of viruses and could be related to Semliki forest virus.

Amino Acid Sequence↗
PubMed1988

Escherichia coli gpt gene provides dominant selection for vaccinia virus open reading frame expression vectors.

Mycophenolic acid, an inhibitor of purine metabolism, was shown to block the replication of vaccinia virus in normal cell lines. This observation led to the development of a dominant one-step plaque selection system, based on expression of the Escherichia coli gpt gene, for the isolation of recombinant vaccinia viruses. Synthesis of xanthine-guanine phosphoribosyltransferase enabled only the recombinant viruses to form large plaques in a selective medium containing mycophenolic acid, xanthine, and hypoxanthine. To utilize the selection system efficiently, we constructed a series of plasmids that contain the E. coli gpt gene and allow insertion of foreign genes into multiple unique restriction endonuclease sites in all three reading frames between the translation initiation codon of a strong late promoter and synthetic translation termination sequences. The selection-expression cassette is flanked by vaccinia virus DNA that directs homologous recombination into the virus genome. The new vectors allow high-level expression of complete or partial open reading frames and rapid construction of recombinant viruses by facilitating the cloning steps and by simplifying their isolation. The system was tested by cloning the E. coli beta-galactosidase gene; in 24 h, this enzyme accounted for approximately 3.5% of the total infected-cell protein.

Cloning, Molecular↗
PubMed1987

Integration of human papillomavirus type 16 into cellular DNA of cervical carcinoma: preferential deletion of the E2 gene and invariable retention of the long control region and the E6/E7 open reading frames.

The integration patterns of human papillomavirus (HPV) type 16 in the cellular DNA of six cervical carcinoma samples were analyzed by the Southern blot procedure. None of the HPV integrants retained the entire viral genome. Double HPV integration was found in one case while all other cases were single integrants. In some samples, internal deletion and selective amplification of the viral sequences were observed. On integration, the E2 open reading frame (ORF) was invariably lost but the E6/E7 ORFs and the long control region of the HPV-16 genome were retained in all seven integrations analyzed and may play a role in cellular transformation and/or maintenance of the transformed phenotype.

Base Sequence↗
PubMed1999

Primary structure of open reading frame 2 and 3 of the hepatitis E virus isolated from Morocco.

The nucleotide sequence from position 5,014 to 7,186 of the hepatitis E virus (HEV) genome was determined using a set of 10 polymerase chain reaction (PCR) fragments amplified directly from a pool of fecal specimens obtained from patients with well-documented epidemic HEV infection in Morocco. This sequence contains the 3'-terminal region of open reading frame 1 (ORF1), full length ORF2 and ORF3, and a portion of the 3'-noncoding region. The HEV Morocco nucleotide sequence was compared with the corresponding sequences of 13 HEV strains. A region of ORF2 that overlaps with ORF3 was found to be the most conserved region of ORF2, whereas a protein segment encoded by this region was found to be the most variable. Theoretical RNA secondary structure analysis predicted that this region may be folded into a strong secondary structure that may constrain nucleotide sequence variability. In addition, the nucleotide sequence comparison revealed that the HEV Morocco sequence is most homologous to the sequences of the HEV Asian strains compared with the HEV Mexico, swine, and US strains. Phylogenetic analysis performed on the entire ORF2 and ORF3 sequences and on a small fragment of ORF2 allowed classification of the HEV Morocco strain together with a few other known African strains as a separate subtype within the Asian-African genotype.

Base Sequence↗
PubMed1989

Genetic analysis of CRPV pathogenesis: the L1 open reading frame is dispensable for cellular transformation but is required for papilloma formation.

Genetic studies to elucidate the role of papillomaviruses in the development and progression of tumors have been severely hampered because the viruses cannot be grown in tissue culture and therefore mutants are not available. We have employed recombinant DNA for papilloma induction to identify essential sequences involved in papillomavirus pathogenesis. Here, we demonstrated that deleting most of the open reading frame (ORF) L2 did not affect the potential of viral cottontail rabbit papillomavirus (CRPV) DNA to induce papillomas. The extrachromosomally maintained DNA in the papillomas was not rearranged and the major transcripts of 1.3 and 2.0 kb encoded E7 and E6, respectively. A recombinant DNA containing a larger deletion lacking the 3' terminal half of ORF L2 and all of ORF L1 (pdlBc/l) did not induce papillomas. The results indicate that sequences in the late region not required for transformation of NIH 3T3 cells by bovine papillomavirus type-1 essential in CRPV for induction of papillomas.

Animals↗
PubMedDate not supplied

Sequencing of coronavirus IBV genomic RNA: a 195-base open reading frame encoded by mRNA B.

DNA sequencing of genomic cDNA clones of avian infectious bronchitis virus (IBV) has been carried out. 770 bases have been determined which include genomic sequences spanning the 5' termini of the two smallest mRNAs of the 3'-coterminal "nested" set: mRNA A and mRNA B. This region contains the complete coding sequences for mRNA B which are additional to those present in mRNA A. Two open reading frames are present, predicting proteins of Mrs 7500 and 9500.

Animals↗
PubMed1988

Transactivating potential of the 3' open reading frame of murine mammary tumor virus.

The procaryotic chloramphenicol acetyltransferase (CAT) gene controlled by the murine mammary tumor virus (MMTV) promoter shows reduced activity in a rat mammary tumor cell line after infection with MMTV but to a considerably lesser extent than the CAT gene controlled by a heterologous promoter, indicating trans-acting regulation of promoter activity by MMTV. Cotransfection of vectors capable of expressing RNA from the 3' open reading frame (orf) of MMTV with the CAT-MMTV construct resulted in enhanced CAT activity, suggesting that orf carries a transactivating potential. Since transactivation was also found with a vector containing only orf and part of the viral env gene, it was concluded that a separate transcriptional unit exists for the orf message.

Animals↗
PubMed1985

Characterization of the ileS-lsp operon in Escherichia coli. Identification of an open reading frame upstream of the ileS gene and potential promoter(s) for the ileS-lsp operon.

The preceding paper presents evidence for the co-transcriptional expression of the ileS and lsp genes in Escherichia coli. To identify the promoter for the ileS-lsp operon, we have determined the nucleotide sequence of an 1.8-kilobase DNA fragment between the rpsT and IleS genes. The sequence data have revealed an open reading frame, designated gene X, which encodes a polypeptide with 312 amino acid residues. Both in vivo and in vitro expressions of the x gene result in the synthesis of a soluble protein with an apparent Mr of 35,000. The x gene is transcribed in the same direction as that of the ileS-lsp operon and opposite to that of the upstream adjacent rpsT gene. No transcription termination sequence can be discerned in the intercistronic region between the x and ileS genes. DNase I footprinting experiment revealed a RNA polymerase binding site at 170-151 base pairs upstream of the x gene.

Bacterial Proteins↗
PubMed1988

The gag gene products of human immunodeficiency virus type 1: alignment within the gag open reading frame, identification of posttranslational modifications, and evidence for alternative gag precursors.

Seven human immunodeficiency virus gag polypeptides were identified in the purified virus and in infected CD4+ lymphocytes by peptide mapping and limited amino acid sequencing of immune-purified proteins. Two gag polyproteins of 55,000 (p55) and 41,000 (p41) daltons were rapidly labeled and readily processed into the major internal gag proteins that were aligned within the gag open reading frame (ORF) as NH2-p16 (MA)-p24 (CA)-p9 (NC)-p7-COOH. The myristoylated p16 (matrix, MA) protein was processed from the myristoylated p55 gag precursor protein. The immunoreactivity of the p16 (MA) protein with region-specific gag antisera and the conservation of the N-terminal myristyl group of the p55 precursor protein in p16 (MA) confirmed its position as the N-terminal-most protein. The p9 (nucleocapsid, NC) protein was localized to residue 378 of the gag ORF, next to the C terminus of the p24/p25 (core antigen, CA) protein. The p9 protein had a repeating Cys residue containing motif which is found in the nucleic acid-binding Cys residue-containing proteins of retroviruses. The p24 (CA) protein, which was localized to residue 133 of the gag ORF, was apparently derived by C-terminal processing of an intermediate polypeptide, p25. Both the mature p24 (CA) and p16 (MA) proteins were phosphorylated at Ser residue(s). We also identified two forms of gag p41 species, one resulting from the C-terminal processing of p55 and the other originating either from N-terminal processing of p55 or from de novo synthesis.

Amino Acid Sequence↗
PubMed2005

Structure and variability of the UL149 open reading frame from low-passage clinical isolates of human cytomegalovirus.

Human cytomegalovirus (HCMV), a ubiquitous herpesvirus, causes a lifelong subclinical infection in healthy adults but leads to significant morbidity and mortality in neonates and immunocompromised individuals. A region (referred to as UL/b') present in the Toledo strain of HCMV and low passage clinical isolates contains 19 additional genes, which are absent in the highly passage laboratory strain AD169. One of these genes, UL149 open reading frame, was amplified by PCR and sequenced from isolates obtained from infants with congenital HCMV infection, to determine whether genetic variation of this gene could influence the signs of the virus infection. The major finding is that the UL149 is a variable gene in all 26 clinical isolates, and the sequences from clinical isolates were classified into three major groups. It is concluded that the HCMV UL149 sequence is variable at the nucleotide level and it might play an important role in HCMV infection.

Amino Acid Sequence↗
PubMed1987

Three suppressor mutations which cure a mitochondrial RNA maturase deficiency occur at the same codon in the open reading frame of the nuclear NAM2 gene.

Dominant mutations of the nuclear NAM2 gene are able to compensate for a deficiency of the maturase encoded by the fourth intron of the mitochondrial cytochrome b gene. We have determined the complete nucleotide sequence of the NAM2-1 suppressor allele. The results of S1 nuclease protection experiments show that two overlapping poly(A)+ RNAs are transcribed from the gene using different promoters. The longer transcript contains two open reading frames (ORFs), a long ORF which could encode a protein of 894 amino acids, mol. wt 102,000 daltons, and a short ORF of 51 codons which is omitted from the shorter transcript. The wild-type nam2+ and two other suppressor alleles, NAM2-6 and NAM2-7, have been cloned. A comparison of the sequence of the wild-type and the three suppressor alleles shows that on three separate occasions the same codon specifying glycine was mutated (once to serine and twice to cysteine). Finally sequence comparisons identified two regions in the long ORF, distinct from the position of the suppressor mutations, that could correspond to binding domains for a nucleotide and a nucleic acid.

Alleles↗