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C S Young

Publications and source records attributed to C S Young.

At least 73 records · Page 4Linked to original sources
PubMed1983

The genetic analysis of adenovirus recombination in triparental and superinfection crosses.

Previous genetic and molecular data suggest that adenovirus genomes can undergo several rounds of recombination before being encapsidated (C. S. H. Young and S. J. Silverstein, Virology 101, 503-515). Two predictions of this hypothesis have been tested. The first is that infection with three differentially marked parental viruses should lead to the appearance of recombinants with genetic contributions from all three parents. In a triparental cross, involving two strains with different ts mutations in chimeric Ad5/Ad2+ND1 backgrounds, and a third strain containing both ts mutations in an Ad5 background, it was demonstrated that multiple recombinations, involving distinguishable restriction endonuclease sites and host range markers from all three parents, were common. The second prediction, from previous kinetic data, is that cells are recombinationally proficient from the eclipse period well into the exponential rise period. This has been tested by superinfecting singly infected cultures, both during eclipse and in early exponential phase. Recombinant viruses were produced in these superinfections, demonstrating that the early to late switch in the replicative cycle does not inhibit recombination. From the temporal appearance of recombinants moreover, it seems likely that recombination functions, and the DNA structures necessary to initiate recombination, are present well into the late phase of replication.

Adenoviruses, Human↗
PubMed1983

The genetic analysis of recombination using adenovirus overlapping terminal DNA fragments.

We have studied the consequences of genetic recombination between overlapping terminal fragments of adenovirus genomes with respect to markers in the overlapping sequence. The findings are consistent with general recombination occurring approximately isotonically within the interval. In particular, single markers within the overlap, scored nonselectively, showed frequencies of recovery dependent on the position of their locus in relation to the ends of the overlap. Pairs of ts markers recombined to form ts+ progeny in proportion to their distance apart, provided the markers were oriented so that a single crossover between them would produce a full-length genome bearing both ts+ alleles. In the opposite orientation, where such a single crossover would be expected to produce ts/ts recombinants, the ts+ frequency was much lower, indicating that multiple recombination events are rare in this system. These findings rule out site-specific recombination, recombination occurring exclusively at the cleaved ends of the overlap, and recombination by means of mismatch repair of a heteroduplex the length of the overlap. They also indicate either that any heteroduplex junction region formed in the course of this reaction is quite short or that it is not subject to heteroduplex repair. Finally, our results demonstrate the efficacy of overlap recombination as a genetic and physical mapping tool and as a method of strain construction, and they suggest other applications, such as using overlap recombination to demonstrate that closely spaced pairs of markers (e.g., putative second-site reversions and their accompanying ts lesions) can be segregated.

Adenoviridae↗
PubMed1983

Detection, rescue, and mapping of mutations in the adenovirus DNA binding protein gene.

r(ts107)202 was isolated in HeLa cells as a temperature-independent revertant of H5ts107, an adenovirus mutant that maps in the structural gene of the viral DNA-binding protein. r(ts107)202 is a host-range temperature-conditional mutant: it is temperature independent for growth in HeLa cells but temperature sensitive for growth in 293 cells, a type 5 adenovirus-transformed human cell line. Marker rescue experiments using H5ts107 DNA and restriction enzyme fragments from r(ts107)202 DNA demonstrated that the mutations causing the r(ts107)202 phenotype were localized in HindIII fragment A containing the entire DNA-binding protein gene. To obtain a fine structure map of the r(ts107)202 mutations, overlap recombination between EcoRI fragment A (0-75.9 map units) from either Ad5wt or r(ts107)202 and BamHI fragment B (59.5-100 map units) from either Ad5wt or r(ts107)202 was performed. Segregation of the H5ts107 primary-site mutation away from the accompanying reversion mutation could be demonstrated in 5 of 200 plaques when r(ts107)202 EcoRI fragment A was crossed with the Ad5wt BamHI fragment. In the reciprocal cross, none of 200 plaques contained the H5ts107 mutant. These results permitted a determination of the order of the primary-site mutation (H5ts107) and secondary-site mutation in r(ts107)202, and the frequency of recombination predicted a distance of 50-340 base pairs between these two mutations. This experimental result agrees with the nucleotide sequence of the r(ts107)202 mutant which shows that 182 base pairs separate the primary-site and secondary-site mutations in r(ts107)202.

Adenoviruses, Human↗
PubMed1983

Expression of adenovirus E1a and E1b gene products and the Escherichia coli XGPRT gene in KB cells.

The recombinant plasmid pSV2-gpt, which contains the Escherichia coli XGPRT gene under the control of a simian virus 40 early promoter, was modified to contain the type 2 adenovirus (Ad2) XhoI-C (0 to 15.5 map units) restriction endonuclease fragment. Plasmid (pLB206) DNA was introduced into human KB cells by Ca2+-mediated DNA transfection, and transformants were selected in medium containing xanthine, aminopterin, and mycophenolic acid, as a consequence of expression of the dominant, selectable XGPRT gene. A series of 13 gpt+ cell lines were isolated and tested for their ability to complement Ad5 deletion mutants in E1a (H5dl312) and E1b (H5dl315). Four classes of gpt+ KB cell lines were identified, including clones constitutively expressing both E1a and E1b, only E1a, or only E1b or not expressing either E1a or E1b. DNA and RNA filter transfer hybridization analysis substantiated the conclusions that those cell lines capable of complementing viral host range mutants contained the appropriate viral DNA sequences and cytoplasmic polyadenylated RNA species. DNA filter transfer hybridization studies also revealed that the transfected vector DNA was stably integrated into chromosomal DNA in the KB transformants and the number of integrated sites ranged from 1 to 3. The gpt+ KB cell line that only expressed E1b gene functions only contained viral E1b gene sequences; those cell lines that expressed neither E1a nor E1b gene function contained only small or no regions of Ad2 DNA. When weaned off the selective medium, transformed KB cell lines stably maintained their inserted DNA in the absence of selective pressure and could easily be adapted to growth in suspension culture.

Adenoviruses, Human↗
PubMed1982

The role of imaging modalities in diagnosis and management of pyomyositis.

A case of tropical pyomyositis in a temperate climate is reported and 29 more cases are reviewed from the literature. Because of its rarity and deceiving clinical presentation, the disorder may go unrecognized for weeks in nontropical regions. The role of various imaging modalities in timely and accurate diagnosis and nonsurgical management of this disorder is discussed.

Child↗
PubMed1981

Tumor promoters alter the temporal program of adenovirus replication in human cells.

In this study we evaluated the effect of phorbol ester tumor promoters on the kinetics of adenovirus type 5 (Ad5) replication in human cells. When added at the time of infection, 12-O-tetradecanoyl-phorbol-13-acetate (TPA) accelerated the appearance of an early virus antigen (72,000-molecular-weight [72K] deoxyribonucleic acid-binding protein), the onset of viral deoxyribonucleic acid synthesis, and the production of infectious virus. The appearance of an Ad5-specific cytopathic effect (CPE) was also accelerated in infected cultures exposed to TPA, whereas phorbol, 4 alpha-phorbol-12,13-didecanoate and 4-OmeTPA, which are inactive as tumor promoters, were ineffective in inducing this morphological change. The acceleration of the CPE seen in TPA-treated Ad5-infected cells was not caused by TPA induction of the protease plasminogen activator, since the protease inhibitors leupeptin and antipain do not inhibit the earlier onset of this CPE and, in contrast, epidermal growth factor, which induces plasminogen activator in HeLa cells, does not induce an earlier CPE. Evidence for a direct effect of TPA on viral gene expression was obtained by analyzing viral messenger ribonucleic acid (mRNA) synthesis. TPA accelerated the appearance of mRNA from all major early regions of Ad5, transiently stimulated the accumulation of region III mRNA, and accelerated the appearance of late Ad5 mRNA. Thus, TPA altered the temporal program of Ad5 mRNA production and accelerated the appearance of at least some Ad5-specific polypeptides during lytic infection of human cells. These effects presumably explain the earlier onset of the Ad5-specific CPE in TPA-treated cells and may have relevance to the effects of TPA on viral gene expression in nonpermissive cells carrying integrated viral deoxyribonucleic acid sequences.

Adenoviridae↗
PubMed1975

Extrachromosomal elements in a super-suppression system of yeast. III. Enhanced detection of weak suppressors in certain non-suppressed psi-strains.

Individual colonies of an adenine-requiring strain of the genotype ade 2-1, SUPQ5, [psi minus] exhibit high and uniform frequencies of ADE+ revertants (5-1 times 10-5). The frequencies for strains of the genotypes ade 2-1, SUPQ5+, [psi+] and ade 2-1 SUPQ5+, [psi minus] are by contrast 1-2 times 10-7 and 0-5 times 10-7 respectively. Mutation from [psi minus] to [psi+] can occur. This suggests that the original mutation from [psi+] to [psi minus] is not necessarily a physical loss of, or extensive deletion, the extrachromosomal elements. Although mutation from [psi minus] to [psi+] occurs, the majority of ADE+ revertants from ade 2-1, SUPQ5, [psi minus] strains contain other types of mutation. Crosses between most ADE+ revertants and ade 2-1, SUPO5+, [psi minus] strains yield adenine-requiring diploids and tetrad segregations of 0:4, 1:3 and 2:2 ADE+ :ade minus. It is argued that most mutations give rise to weak recessive suppressors (SUPX) which are themselves incapable of suppressing the ochre allele ade 2-1, unless the allele SUPQ5 is also present. Tests with two suppressors, S-theta and S-zeta, isolated by Dr R. A. Gilmore, which are themselves incapable of suppressing ade 2-1, show that they do not do so even in the presence of SUPQ5. Thus the set of suppressors isolated by us is a special class which can perhaps only be observed in the genetic background ade 2-1, SUPQ5, [psi minus]. There is evidence that some of the new suppressors are capable, in the absence of SUPQ5, of weakly suppressing trp 5-48 and perhaps can 1-100. All ochre alleles are, however, suppressed by the combination SUPQ5, SUPX. SUPQ5 is able to suppress trp 5-48, his 5-2, lys 1-1 and can 1-100 in a [psi minus] background, although it does so variably. It is incapable of suppressing ade 2-1 in such a background. It is argued that the alteration in the [psi] determinant results in a lowered efficiency of suppression by SUPQ5 rather than a changed specificity.

Adenine↗
PubMed1975

Action of interferon in enucleated cells.

Interferon induces protection of enucleated BSC-1 cells against infectious vesicular stomatitis virus production if cells are treated before, but not after, enucleation.

Animals↗
PubMed1975

Heat-stable variant of human adenovirus type 5: characterization and use in three-factor crosses.

A variant of human adenovirus type 5 which is heat stable (hs) in vitro has been isolated following three rounds of heat inactivation at 52 C. The variant is genetically stable, both through vegetative viral passage and through recombination into other genetic backgrounds, which suggests that it arises from a single mutation. Three-factor crosses, using this mutant in conjunction with previously described temperature-sensitive mutants, suggest the hs mutation lies near the left-hand end of the genetic map. The mutant has been used to demonstrate the production of reciprocal recombinants in two-factor crosses. The mutational lesion is unknown, but phenotypic mixing occurs in hs times hs-+ infections, which suggests that it lies in a gene specifying a viri on structural protein. Other biological parameters examined have shown no differences from the wild-type hs-+.

Adenoviridae↗