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D C Thomas

Publications and source records attributed to D C Thomas.

At least 73 records · Page 4Linked to original sources
PubMed1994

Mispair-, site-, and strand-specific error rates during simian virus 40 origin-dependent replication in vitro with excess deoxythymidine triphosphate.

We have measured the fidelity of leading and lagging strand DNA replication in HeLa cell extracts. Providing an excess of one dNTP in reactions induces replication errors consistent with misincorporation of that dNTP. With excess dTTP, both substitutions and single-nucleotide frameshifts are induced. Error distribution is nonrandom; reproducible hot spots for a substitution and a frameshift error are observed. Measurements with two vectors having the origin of replication on opposite sides of the mutational target demonstrate that error rates for G.dTTP and C.dTTP mispairs depend on whether the strand is replicated as the leading or lagging strand. Also, the two hot spots are only observed in one origin-target orientation. Replication reactions reconstituted from two fractions derived from extracts are 3-fold less accurate, but the error specificity with excess dTTP is similar to that with extracts. This suggests that the processes responsible for the nonrandom error rates are not lost as a result of fractionation. Furthermore, the reconstituted system is devoid of mismatch repair activity. Thus, mismatch repair is not responsible for the mispair-, site-, and strand-specific differences observed.

Base Sequence↗
PubMed1994

Exposure to magnetic fields among electrical workers in relation to leukemia risk in Los Angeles County.

To address the hypotheses that electrical workers are exposed to higher magnetic fields and are at higher risk of leukemia than nonelectrical workers, we performed a registry-based case-control study among men aged 20-64 years with known occupation who were diagnosed with cancer in Los Angeles County between 1972 and 1990. Controls were men with cancers other than those of the central nervous system or leukemia. Magnetic field measurements on workers in each electrical occupation and in a random sample of occupations presumed to be nonelectrical were used to estimate magnetic field exposures for each occupation. Among men in electrical occupations, 121 leukemias were diagnosed. With the exception of electrical engineers, magnetic field exposures were higher among workers in electrical occupations than in nonelectrical occupations. A weakly positive trend in leukemia risk across average occupational magnetic field exposure was observed (odds ratio [OR] per 10 milligauss increase in average magnetic field = 1.2, 95% confidence interval [CI] 1.0-1.5). A slightly stronger association was observed for chronic myloid leukemia, although only 28 cases occurred among electrical workers (OR 10 milligauss increase = 1.6, 95% CI = 1.2-2.0). The results were not materially altered by adjustment for exposure to several agents known or suspected to cause leukemia. Although not conclusive, these results are consistent with findings from studies based on job title alone that electrical workers may be at slightly increased risk of leukemia.

Adult↗
PubMed1994

Censored survival models for genetic epidemiology: a Gibbs sampling approach.

Methods are proposed for the analysis of diseases with variable age at onset. The Cox proportional hazards model, widely used for epidemiologic analysis, is modified to include both measured (environmental) covariates and latent (genetic) variables, as well as their interactions. A Monte Carlo technique known as Gibbs sampling is utilized to generate observations from the posterior distributions of all model parameters. A correction to account for single ascertainment of pedigrees is also described. Simulation studies show that parameter estimation is nearly unbiased for a wide variety of models, and that moderate gene-environment interaction effects can be detected.

Age of Onset↗
PubMed1994

Processed meats and risk of childhood leukemia (California, USA).

The relation between the intake of certain food items thought to be precursors or inhibitors of N-nitroso compounds (NOC) and risk of leukemia was investigated in a case-control study among children from birth to age 10 years in Los Angeles County, California (United States). Cases were ascertained through a population-based tumor registry from 1980 to 1987. Controls were drawn from friends and by random-digit dialing. Interviews were obtained from 232 cases and 232 controls. Food items of principal interest were: breakfast meats (bacon, sausage, ham); luncheon meats (salami, pastrami, lunch meat, corned beef, bologna); hot dogs; oranges and orange juice; and grapefruit and grapefruit juice. We also asked about intake of apples and apple juice, regular and charcoal broiled meats, milk, coffee, and coke or cola drinks. Usual consumption frequencies were determined for both parents and the child. When the risks were adjusted for each other and other risk factors, the only persistent significant associations were for children's intake of hot dogs (odds ratio [OR] = 9.5, 95 percent confidence interval [CI] = 1.6-57.6 for 12 or more hot dogs per month, trend P = 0.01), and fathers' intake of hot dogs (OR = 11.0, CI = 1.2-98.7 for highest intake category, trend P = 0.01). There was no evidence that fruit intake provided protection. While these results are compatible with the experimental animal literature and the hypothesis that human NOC intake is associated with leukemia risk, given potential biases in the data, further study of this hypothesis with more focused and comprehensive epidemiologic studies is warranted.

Animals↗
PubMed1994

Correction of the non-uniform spatial sensitivity of electrical impedance tomography images.

One of the clinical applications of electrical impedance tomography (EIT) is the measurement of volume changes of body fluids. However, the spatial sensitivity of images produced by the Sheffield Mark I system is non-uniform and if quantitative comparisons of resistivity images resulting from volume changes are to be mad, this non-uniformity must be corrected for. All experimental data were collected from a phantom consisting of semi-permeable membrane tubes fixed vertically at different distances from the centre of a cylindrical tank which was filled with a saline solution of resistivity 5 omega m. Individual images were obtained when each tube was filled with a saline solution of resistivity 1.5 omega m. Analysis of these images, using the resistivity integral described by Thomas et al, yielded the sensitivity of EIT at different positions within the tank. Combinations of tubes were then filled with saline (1.5 omega m) and an image produced for each combination. Cross sections of the images were taken and used to confirm that the principle of superposition is valid for EIT. A simple pixel scaling correction, derived from the sensitivity data, was then applied to the images and the results re-analysed to demonstrate a reduction in volume measurement error. The application of this correction to in vivo images is discussed.

Electric Impedance↗
PubMed1993

A cohort study of thyroid disease in relation to fallout from nuclear weapons testing.

OBJECTIVE: To estimate individual radiation doses and current thyroid disease status for a previously identified cohort of 4818 schoolchildren potentially exposed to fallout from detonations of nuclear devices at the Nevada Test Site between 1951 and 1958. DESIGN: Cohort analytic study. SETTING: Communities in southwestern Utah, southeastern Nevada, and southeastern Arizona. PARTICIPANTS: Individuals who were still residing in the three-state area (n = 3122) were reexamined in 1985 and 1986, and information on the subjects' and their mothers' milk and vegetable consumption during the fallout period was obtained by telephone interview (n = 3545). After exclusions to eliminate missing data and confounding factors, 2473 subjects were available for analysis. MAIN OUTCOME MEASURES: Individual radiation doses to the thyroid were estimated by combining consumption data with radionuclide deposition rates provided by the US Department of Energy and a survey of milk producers. Relative risk models adjusted for age, sex, and state were fitted using maximum likelihood to period prevalence data for thyroid carcinomas, neoplasms, and nodules. RESULTS: Doses ranged from 0 mGy to 4600 mGy, and averaged 170 mGy in Utah. There was a statistically significant excess of thyroid neoplasms (benign and malignant; n = 19), with an increase in excess relative risk of 0.7% per milligray. A relative risk for thyroid neoplasms of 3.4 was observed among 169 subjects exposed to doses greater than 400 mGy. Positive but nonsignificant dose-response slopes were found for carcinomas and nodules. CONCLUSIONS: Exposure to Nevada Test Site-generated radioiodines was associated with an excess of thyroid neoplasms. The conclusions are limited by the small number of exposed individuals and the low incidence of thyroid neoplasms.

Adolescent↗
PubMed1993

Relative probability of mutagenic translesion synthesis on the leading and lagging strands during replication of UV-irradiated DNA in a human cell extract.

We have previously demonstrated mutagenic bypass of pyrimidine dimers during SV40 origin-dependent replication of UV-irradiated DNA in human cell extracts [Thomas, D. C., & Kunkel, T. A. (1993) Proc. Natl. Acad. Sci. U.S.A. 90, 7744-7748]. Here we use two vectors having the origin of replication on opposite sides of a lacZ alpha reporter gene to examine the relative probability of mutagenic translesion synthesis on the leading and lagging strands. Although replication of both vectors is inhibited by UVB irradiation in a dose-dependent manner, the covalently closed DNA products of replication contain T4 endonuclease sensitive sites, indicating that bypass of cyclobutane pyrimidine dimers occurred. At fluences of 70 and 100 J/m2, the mutant frequencies obtained with both vectors are substantially higher than with control DNAs. Sequence analysis of mutants obtained with both vectors reveal three types of mutations at frequencies significantly above those obtained from replication of undamaged DNA. These are C-->T transitions, accounting for about two-thirds of the mutants, a small number of CC-->TT substitutions, and complex mutations. Comparing the distribution of C-->T substitutions in the two spectra permits an estimation of the probability of mutagenic translesion replication of the same sequence when replicated as the leading or lagging strand. The data suggest that the overall average UV-independent C-->T substitution probability per phenotypically detectable dipyrimidine site is the same during leading and lagging strand replication. However, statistically significant differences are observed when the distribution of C-->T substitutions is considered.(ABSTRACT TRUNCATED AT 250 WORDS)

Base Sequence↗
PubMed1993

Replication of UV-irradiated DNA in human cell extracts: evidence for mutagenic bypass of pyrimidine dimers.

We have examined the efficiency and fidelity of simian virus 40-origin-dependent replication of UV-irradiated double-stranded DNA in extracts of human cells. Using as a mutational target the alpha-complementation domain of the Escherichia coli lacZ gene in bacteriophage M13mp2 DNA, replication of undamaged DNA in HeLa cell extracts was highly accurate, whereas replication of DNA irradiated with UV light (280-320 nm) was both less efficient and less accurate. Replication was inhibited by irradiation in a dose-dependent manner. Nonetheless, covalently closed, monomer-length circular products were generated that were resistant to digestion by Dpn I, showing that they resulted from semiconservative replication. These products were incised by T4 endonuclease V, whereas the undamaged replication products were not, suggesting that pyrimidine dimers were bypassed during replication. When replicated, UV-irradiated DNA was used to transfect an E. coli alpha-complementation host strain to score mutant M13mp2 plaques, the mutant plaque frequency was substantially higher than that obtained with either unirradiated, replicated DNA, or unreplicated, UV-irradiated DNA. Both the increased mutagenicity and the inhibition of replication associated with UV irradiation were reversed by treatment of the irradiated DNA with photolyase before replication. Sequence analysis of mutants resulting from replication of UV-irradiated DNA demonstrated that most mutants contained C-->T transition errors at dipyrimidine sites. A few mutants contained 1-nt frameshift errors or tandem double CC-->TT substitutions. The data are consistent with the interpretation that pyrimidine dimers are bypassed during replication by the multiprotein replication apparatus in human cell extracts and that this bypass is mutagenic primarily via misincorporation of dAMP opposite a cytosine (or uracil) in the dimer.

DNA↗
PubMed1993

Fidelity of DNA replication by extracts of normal and malignantly transformed human cells.

To test the hypothesis that a mutator phenotype may be associated with carcinogenesis (L. A. Loeb, Cancer Res., 51: 3074-3079, 1991), we have compared the fidelity of double-stranded DNA replication and the efficiency of mismatch repair in extracts from normal diploid and malignantly transformed human cells. Included was a diploid fibroblast strain and its transformed derivative, as well as a second diploid fibroblast strain and HeLa cells. The fidelity of DNA replication by cytoplasmic extracts in the presence of simian virus 40 large tumor antigen (SV40 T-antigen) was measured using a forward mutagenesis assay. The replicated DNA consisted of double-stranded M13 mp2 DNA containing the SV40 origin of replication and the lacZ alpha complementation gene as a target sequence for scoring mutations. T-antigen-dependent replication was detected in all cell extracts, with those from transformed cells having the greatest activity. No differences in replication fidelity were detected between normal and transformed cell extracts. Using a heteroduplex containing a G.G mispair, we also detected mismatch repair activity in the cell extracts, including efficient repair in extracts from malignantly transformed cells. While these data do not eliminate the possibility that a mutator phenotype may be associated with carcinogenesis, they do suggest that genetic instability associated with transformation does not involve reduced fidelity of replication of undamaged DNA or reduced mismatch repair efficiency.

Cell Line, Transformed↗
PubMed1993

Vesicular stomatitis virus glycoprotein contains a dominant cytoplasmic basolateral sorting signal critically dependent upon a tyrosine.

To investigate the contribution of the cytoplasmic domain of the vesicular stomatitis virus G glycoprotein to its basolateral expression in polarized epithelial cells, chimeric proteins containing the external and transmembrane domains of an apically targeted protein, the influenza virus hemagglutinin (HA), and either the G cytoplasmic domain or an unrelated cytoplasmic sequence, were introduced into Madin-Darby canine kidney (MDCK) cells. Addition of the cytoplasmic tail of G to a truncated HA resulted in delivery of greater than 95% of the chimeric protein to the basolateral cell surface, indicating that the G cytoplasmic domain contains a dominant basolateral sorting signal. A similar chimera, containing the cytoplasmic tail of herpes simplex I glycoprotein gC, was not sorted basolaterally. Deletion of the cytoplasmic tail from G protein itself decreased the fidelity of sorting to the basolateral surface, but not the extent to which the protein reached the plasma membrane. Mutation of cytoplasmic tyrosine 501 of G caused an identical loss of basolateral targeting, suggesting that the tyrosine, or the sequence surrounding it, is required for efficient basolateral transport of G. Mutation of tyrosine 501 had no effect on internalization of G, which was much slower than that of endocytic receptors. Thus, VSV G protein contains an efficient cytoplasmic basolateral targeting signal that is not an efficient internalization signal.

Amino Acid Sequence↗
PubMed1993

A generalized estimating equations approach to fitting major gene models in segregation analysis of continuous phenotypes.

This paper discusses the application of estimating equations methods based on a quadratic exponential model [Prentice and Zhao, 1991] as a potential competitor with full likelihood approaches to estimating the effect of major genes in a segregation analysis [Elston, 1981] of continuous phenotypes, in the single allele problem. We show that while the estimating equations methods based on the quadratic exponential family cannot be used by themselves to estimate all parameters of interest, an iterative two-stage approach can be used, in which the population allele frequency is first considered to be a known parameter, which permits the estimating equations method to estimate the remaining parameters. At the second stage a "pseudo-profile" loglikelihood based only on the founders is used to estimate the allele frequency. After each maximization of the pseudo-profile loglikelihood at the second stage, the parameters in the first stage are reestimated using a new value of the allele frequency, and a new value of the second stage pseudo-profile loglikelihood is obtained. We used simulated pedigree data for illustrations.

Alleles↗
PubMed1993

Use of generalized estimating equations in segregation analysis of continuous outcomes.

The use of generalized estimating equations (GEE) [Liang and Zeger, 1986] is explored as a potential alternative to maximum likelihood methods for segregation analysis of continuous phenotypes. We discuss the number of sample moments required by GEE to estimate parameters in simple major gene models, and the computational burden of the GEE methods applied to large pedigrees.

Humans↗
PubMed1993

A bivariate genetic analysis of HDL- and LDL-cholesterol incorporating measured covariates: a Gibbs sampling application.

We analyzed HDL- and LDL-cholesterol levels as a bivariate phenotype in 27 families as a function of major genes, polygenes, and measured covariates using a Monte Carlo sampling technique called Gibbs sampling. Major genes and polygenes exhibited strong effects, when considered separately. While a major gene versus polygene model could not be clearly differentiated for HDL-C, polygenes appeared to play a stronger role than a major gene for LDL-C. There was no evidence of linkage between the two major genes for HDL- and LDL-C, and the correlation in polygenes was negative. The analysis illustrate the potential applicability of Gibbs sampling to such complex problems as the multivariate analysis of continuous phenotypes.

Cholesterol, HDL↗
PubMed1993

Does the gender difference in interferon production seen in picornavirus-infected spleen cell cultures from ICR Swiss mice have any in vivo significance?

Splenocyte cultures from female ICR Swiss mice produced greater interferon (IFN) levels, particularly IFN-gamma, than did cultures from males by 12 h post-infection (pi) with the D variant of encephalomyocarditis virus (EMCV-D). This early IFN-gamma is produced by natural killer (NK)-like cells and is dependent on plastic adherent cells and IFN-alpha/beta. In this study, we evaluated the significance of this observation on the innate resistance of ICR Swiss females to EMCV-D-mediated disease. Treatment of females with rabbit anti-mouse IFN-alpha/beta serum rendered them susceptible to the diabetogenicity of EMCV-D. Although sera from both sexes of ICR Swiss mice exhibited peak IFN levels day 3 pi, IFN-gamma was present in the sera of males at only 1 day pi and in the sera of females at days 1-3 pi. Females cleared virus from the circulation by day 2 pi, 1 day earlier than did males. Flow cytometric evaluations of lymphoid cell phenotypes in spleens and pancreata of infected mice revealed that percentages of L3T4+ cells were significantly decreased only in spleens from males at day 1 pi and were diminished along with Ly2+ cells in pancreata of males at 7 days pi, suggesting that T-cell responses were impaired in virus-infected males.

Animals↗
PubMed1993

Effects of 3'-azido-3'-deoxythymidine metabolites on simian virus 40 origin-dependent replication and heteroduplex repair in HeLa cell extracts.

Although the capacity of 3'-azido-3'-deoxythymidine (AZT) triphosphate, an active metabolite of the antiviral agent zidovudine (AZT), to inhibit polymerization by a variety of purified DNA polymerases has been described, it is important to understand its effect on replication by the more complex protein assemblies responsible for DNA replication in human cells. In the present study, we have determined the effects of AZT metabolites on the efficiency of simian virus 40 origin-dependent bidirectional replication of double-stranded DNA in extracts of human HeLa cells. Replication was inhibited by AZT diphosphate and AZT triphosphate, but only at concentrations exceeding those thought to be present in vivo. However, replication was inhibited by AZT monophosphate at concentrations previously reported to accumulate in human cells cultured in the presence of AZT, suggesting that AZT monophosphate may contribute to cytotoxicity by inhibiting chromosomal replication. In an attempt to determine whether AZT treatment could have longer term mutagenic effects on cells, we also determined the effects of these AZT derivatives on replication fidelity and on the efficiency of repair of DNA substrates containing single-base mismatches. Despite the ability of a normal deoxynucleoside monophosphate to reduce the fidelity of DNA replication, presumably by reducing exonucleolytic proofreading of errors, neither the mono-, di-, nor triphosphate form of AZT reduced base substitution fidelity when present in replication reactions. Similarly, the efficiency of repair of DNA substrates containing single-base mismatches was unaffected by these compounds. However, replication fidelity was affected by perturbations in relative and absolute concentrations of deoxynucleoside triphosphate substrates similar to those reported to occur in AZT-treated cells. Thus, AZT treatment could potentially be mutagenic in vivo via reduced replication fidelity resulting from alterations in deoxynucleoside triphosphate pools.

Antiviral Agents↗