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R J Colonno

Publications and source records attributed to R J Colonno.

At least 73 records · Page 4Linked to original sources
PubMed1985

Molecular cloning and complete sequence determination of RNA genome of human rhinovirus type 14.

The genomic RNA of human rhinovirus type 14 was cloned in Escherichia coli and the complete nucleotide sequence was determined. The RNA genome is 7212 nucleotides long. A single large open reading frame of 6536 nucleotides was identified, which starts at nucleotide 678 and ends 47 nucleotides from the 3' end of the RNA genome. Comparisons of the specified proteins with those of other picornaviruses showed a striking homology (44-65%) between rhinovirus and poliovirus. The rhinovirus genomic RNA is rich in adenosine (32.1%) and strongly favors an adenosine or uridine in the third position of codons. The predicted map locations of all the rhinovirus structural and non-structural proteins and their proposed proteolytic cleavage sites are described.

Amino Acid Sequence↗
PubMed1985

In vitro synthesis of an infectious RNA from cDNA clones of human rhinovirus type 14.

Development of a novel infectious cDNA assay is described for human rhinovirus type 14. A full-length cDNA clone of the human rhinovirus type 14 genome RNA was assembled and transcribed in vitro by using the SP6 transcription system. Transfection of HeLa cells with the nascent RNA resulted in the production of rhinovirus indistinguishable from the parental virus by both immunological and polyacrylamide gel analysis.

Cloning, Molecular↗
PubMed1984

Many rhinovirus serotypes share the same cellular receptor.

Twenty-four human rhinovirus serotypes were grown and purified by centrifugation in metrizamide density gradients. These preparations had a lower buoyant density (1.24 g/cm3) and higher specific infectivities (1:24 to 1:240) than did rhinoviruses described previously (E. J. Stott and R. J. Killington, Annu. Rev. Microbiol. 26:503-524, 1972). Binding conditions in which the unique cellular receptors for virus attachment were saturated were determined for each serotype. Competition binding assays between pairs of serotypes allowed 20 of the 24 serotypes to be assigned to the same cellular receptor. The remaining four serotypes appeared to attach to a different cellular receptor. Since most serotypes were chosen for study at random, it seems likely that many of the yet unstudied rhinoviruses will share this common cellular receptor.

Electrophoresis, Polyacrylamide Gel↗
PubMed1983

Interferon induction of fibroblast proteins with guanylate binding activity.

Treatment of human diploid fibroblastic cells with interferon induces the synthesis of two guanylate binding proteins (GBP) with molecular weights of 67,000 and 56,000. The Mr = 67,000 protein (67K GBP) is synthesized upon treatment with either alpha-, beta-, or gamma-interferon. Among these interferons, gamma-interferon induces a higher level of 67K GBP synthesis. The 67K GBP synthesized in either beta- or gamma-interferon-treated cells has two charge forms with isoelectric points of 6.0 and 5.8, respectively. The synthesis of the Mr = 56,000 protein is induced by the treatment using either alpha- or beta-interferon, but its synthesis in gamma-interferon-treated cells is undetectable. The amounts of the radioactive GBPs synthesized in human fibroblasts are proportional to the amounts of the purified beta-interferon used for the inductions. Syntheses of GBPs require the transcription of cellular genes because their syntheses are completely blocked by actinomycin D treatments. The mRNA for the 67K GBP is found in fibroblasts that are treated by either alpha-, beta-, or gamma-interferon, but it is not detected in untreated cells. More 67K GBP mRNA is accumulated in the gamma-interferon-treated than in alpha- or beta-interferon-treated fibroblasts. This is consistent with more 67K GBP synthesis found in gamma-interferon-treated fibroblasts.

Cell Line↗
PubMed1983

Different mRNAs induced by interferon in cells from inbred mouse strains A/J and A2G.

Treatment of cells from inbred mouse strains A/J and A2G with interferon resulted in the development of different antiviral states for influenza viruses. A2G mice-derived cells that carry the resistance gene Mx were efficiently protected by interferon against influenza viruses, whereas the interferon protection against the same viruses in wild-type A/J mice-derived cells was only marginal. The two cell types, however, were equally protected by interferon against vesicular stomatitis virus and other non-orthomyxoviruses. The interferon-induced mRNAs of mouse embryonic fibroblast cells that carried either homozygous wild-type alleles or homozygous Mx alleles were compared. The isolated polysome-bound mRNAs from A/J (+/+) and A2G (Mx/Mx) cells were translated in a cell-free translation system, and the translation products were analyzed after two-dimensional gel electrophoresis. New mRNAs coding for at least eight proteins with molecular weights (MW) ranging from 30,000 to 80,000 were found in interferon-treated cells but not in control cells. Differences in the interferon-induced mRNAs from A/J and A2G cells were also found. An mRNA coding for a 72,000-MW protein was found in interferon-treated A2G cells but not in interferon-treated A/J cells. Interferon-treated A/J cells, on the other hand, contained an mRNA coding for a 65,000-MW protein that was not found in interferon-treated A2G cells. The in vitro-synthesized 65,000-MW protein efficiently bound to GMP. Cytoplasmic extracts prepared from interferon-treated A/J cells also contained a GMP-binding 65,000-MW protein that was undetectable in similarly treated A2G cells.

Animals↗
PubMed1982

Induction of unique mRNAs by human interferons.

Treatment of human fibroblast cells with human interferon (INF-alpha, IFN-beta, or IFN-gamma) resulted in the accumulation of at least four newly synthesized mRNAs. The mRNAs code for proteins having molecular weights of 56,000, 57,000, 62,000, and 68,000 when characterized in a wheat germ cell-free translation system. A direct relationship was observed between the amount of IFN used and the degree of both the accumulation of the induced mRNAs and the development of an antiviral state. In the case of IFN-alpha or IFN-beta, time course studies indicated that the induced mRNAs appeared as early as 40 min, accumulated for 2 h, then remained ribosome bound for up to 16 h. The ability of fibroblast cells to develop an antiviral state always coincided directly with both the appearance and the level of accumulation of the induced mRNAs. Further mRNA synthesis beyond 2 h had a minimal effect on the development of an antiviral state. Human IFN-gamma also induced the synthesis of the same four mRNAs but required higher interferon titers and a longer incubation time. In addition, IFN-gamma induced a disproportionate amount of the mRNA coding for the 68,000 molecular weight protein and three new mRNAs not detected in cells treated with IFN-alpha or IFN-beta. Mouse interferon induces the original four mRNAs in human cells but to a far lesser extent. This correlated with the inability of these cells to develop much resistance to viral infection.

Cell Line↗
PubMed1981

Accumulation of newly synthesized mRNAs in response to human fibroblast (beta) interferon.

Treatment of human fibroblast cells with human fibroblast (beta)interferon for up to 8 hr resulted in the accumulation of at least four mRNAs. The mRNAs were isolated from cellular polysomes and characterized by stimulation of translation in a wheat germ cell-free protein synthesis system. The mRNAs appear as early as 2 hr after exposure to interferon and can be translated in vitro into proteins having molecular weights of 61,000, 62,000, 64,000, and 68,000. The response is not elicited by mouse interferon or insulin and does not occur in the presence of actinomycin D. Chase experiments indicated that the induced mRNAs remain ribosome-associated for at least 3 hr after their synthesis. The appearance of the induced mRNAs correlated directly with the onset of an antiviral state. Velocity sedimentation of the induced mRNAs on sucrose gradients demonstrated that each of the four induced proteins are encoded by different-sized mRNAs.

Fibroblasts↗
PubMed1979

Interaction of a limited set of proteins with different mRNAs and protection of 5'-caps against pyrophosphatase digestion in initiation complexes.

A variety of 5'-3H-methyl-labeled, oxidized viral mRNAs were used as probes for detecting in wheat germ initiation complexes proteins that interact with, and can be cross-linked to, the 5'-cap structure. A limited and reproducible set of specific proteins was obtained with the different mRNAs. The binding of these proteins to the 5'-end of mRNA apparently results in protection against nucleotide pyrophosphatase digestion of the cap even in initiation complexes in which the 5'-end is susceptible to pancreatic RNase digestion. Cross-linked proteins from mammalian initiation complexes comigrated with several of the subunits of similarly treated eIF-3. A model for cap binding protein interaction with mRNA cap during initiation of translation is suggested.

Animals↗
PubMed1978

Intervening polyadenylate sequences in RNA transcripts of vesicular stomatitis virus.

Purified and partially resolved vesicular stomatitis virus (VSV) messenger RNA has been annealed to the VSV genomic RNA and visualized in the electron microscope under conditions in which duplex regions have a wider image width than single-stranded RNA. The locations of the intercistronic boundaries between the messages have been mapped on the VSV genome. The contour of the double-stranded regions is occasionally interrupted by looped-out single-stranded RNA. The loops are comprised of post transcriptionally synthesized polyadenylate. Most of these structures are found at the intercistronic boundaries and covalently bridge adjacent message sequences. In this paper, we discuss the possible significance of these loops.

Chromosome Mapping↗
PubMed1978

Complete nucleotide sequence of the leader RNA synthesized in vitro by vesicular stomatitis virus.

The complete nucleotide sequence of the leader RNA synthesized in vitro by the Indiana serotype of vesicular stomatitis virus is presented. The sequence was determined by the technique described by Donis-Keller, Maxam and Gilbert (1977) in combination with the standard two-dimensional fingerprint techniques described by Barrell (1971). The leader RNA contains 48 nucleotides variably terminating at the 3' terminus with cytosine (68%) and adenosine at position 47 (32%). Since the leader RNA is complementary to the 3' terminal portion of the viral genome RNA, the first 48 nucleotides from the 3' end of the genome RNA can be decuded. The leader RNA contains repetitive and palindromic sequences with a polypurine sequence at its 3' terminus. The possible role of some of the sequences is discussed.

Base Sequence↗
PubMed1978

Nucleotide sequence of the leader RNA of the New Jersey serotype of vesicular stomatitis virus.

Sequence for the leader RNA Synthesized by the New Jersey serotype of vesicular stomatitis virus is presented and its complementary sequence representing the 3'-terminal sequence of the genome RNA is deduced. Comparison with the leader RNA sequence of the serologically distinct Indiana strain reveals that the 3'-terminal region of the genomes of two viruses is highly conserved.

Base Sequence↗
PubMed1978

In vitro RNA transcription by the New Jersey serotype of vesicular stomatitis virus. II. Characterization of the leader RNA.

The New Jersey serotype of vesicular stomatitis virus (VSV) was able to synthesize a small RNA (leader RNA) approximately 70 bases in length similar to the leader RNA synthesized in vitro by the genetically distinct Indiana serotype of VSV. Also, the New Jersey leader RNA contained the same 5'-terminal sequence, ppA-C-G, as the Indiana leader RNA and had a very similar base composition, with 42% AMP, 16% CMP, 18.6% GMP, and 23.4% UMP. The 3'-terminal sequence of the VSV New Jersey genome RNA was detemined and found to contain the sequence- Py-G-UOH, again the same as that of the Indiana serotype of VSV. Evidence that the New Jersey leader RNA is transcribed from the 3' end of the genome RNA was obtained from the fact that it can protect the 3'-terminal base of [3H]borohydride-labeled New Jersey genome RNA from RNase digestion. Although the New Jersey and Indiana leader RNAs were similar in many respects, they were unable to form RNase-resistant hybrids when annealed to heterologous genome RNA.

Base Sequence↗
PubMed1977

In vitro synthesis of messenger RNA by a defective interfering particle of vesicular stomatitis virus.

A defective interfering particle derived from the heat-resistant strain of vesicular stomatitis virus was analyzed for the presence of virion-associated RNA polymerase (nucleosidetriphosphate:RNA nucleotidyltransferase, EC 2.7.7.6) activtiy. The RNA synthesizing capacity of the defective particles in vitro was similar to that of the wild-type virus. Characterization of the RNA produced in vitro indicated that the defective particles were able to synthesize vesicular stomatitis virus leader RNA and four virus mRNA species that sediment at 12-18 S. These RNA products were identical to the mRNAs synthesized in vitro by the wild-type virus in regard to size, polyadenylation, capping, and methylation. In contrast to the wild-type virus, the purified defective particles did not synthesize the large mRNA species sedimenting at 31 S in vitro. Possible mechanisms of homotypic and heterotypic interferences shown by this defective particle are discussed.

Chromosome Mapping↗
PubMed1977

Vesicular stomatitis virus: mode of transcription.

Recent studies on the mechanism by which the virion-associated RNA polymerase of vesicular stomatitis virus transcribes RNA have revealed several new biological features of general interest. The mode of synthesis of the 5'-terminal cap structure of the mRNAs, the sequential transcription of the genes and the presence of a transcribed "leader" RNA segment are properties which are either not shown by other viruses, or have not yet been described. These features are probably inter-related with the primary transcription process, which itself may be a useful model for future studies on mRNA biosynthesis in eukaryotic systems.

DNA-Directed RNA Polymerases↗
PubMed1977

Characterization of vesicular stomatitis virus mRNA species synthesized in vitro.

The smallest size class of mRNA (12S) synthesized in vitro by the virion-associated RNA polymerase of vesicular stomatitis virus contains two mRNA species of similar molecular weight that code for the viral M and NS proteins. The resolution of these mRNA species was achieved by converting them to duplexes by annealing with the genome RNA, followed by RNase T2 treatment and separation in a polyacrylamide gel. Using this separation technique, the mRNA's were identified by comparing the relative resistance of their syntheses to UV irradiation of the virus. The molecular weights of these two mRNA species calculated as duplex RNAs were smaller than expected. The possible reasons for this discrepancy are discussed.

Cell-Free System↗
PubMed1976

A unique RNA species involved in initiation of vesicular stomatitis virus RNA transcription in vitro.

Purified virions of vesicular stomatitis virus (VSV) are capable of synthesizing two distinct types of virus-specific RNA in vitro. The first consists of several viral mRNAs which have been previously shown to contain the blocked 5' terminal sequence GpppApApCpApGp and 3' terminal poly(A). The second type of RNA has an unblocked 5' terminus and does not contain poly(A) stretches long enough to bind to oligo (dT)-cellulose columns. It migrates in 20% polyacrylamide gels as a single homogeneous peak with an estimated chain length of 68 nucleotides. Base analysis demonstrated that this small RNA molecule is composed of 48% AMP, 20% CMP, 11% GMP, and 21% UMP. The 5' terminal sequence of the small RNA is ppApCpGp, which appears to be complementary to the 3' terminal sequence of the VSV genome RNA (...PypGpU). These results indicate that this small RNA molecule probably represents the intitiated lead-in RNA segment which is removed during formation of VSV mRNAs by a possible processing mechanism.

Adenosine↗