Error-free Process Research Articles

Induction of error-free DNA repair inEscherichia coli by heat shock or nutritional stress

The thermal induction of resistance to UV inEscherichia coli JE1011 has now been shown to depend on an error-free DNA repair process. Heat shock causes a simultaneous reduction in the UV-induced frequency of mutation to prototrophy of two nutritional markers. In contrast, induction of SOS repair by thymine starvation causes the expected increase in mutation frequency upon UV irradiation. Thiamine deprivation produces an effect similar to heat shock, leading to increased UV resistance and a large decrease in the radiation-induced mutation frequency. Thus, two very different types of stress are able to induce error-free DNA repair in these cells.

Excision repair of UV- or benzo[ a]pyrene diol epoxide-induced lesions in xeroderma pigmentosum variant cells is ‘error free’

It is known that cells from one class of xeroderma pigmentosum (XP) patients, called XP variants, carry out excision repair of UV-induced DNA damage at a normal rate and are only slightly more sensitive than normal cells to the cytotoxic effect of UV radiation, but are much more sensitive to the mutagenic effect of UV. To see if this hypermutability were the result of an ‘error-prone’, excision repair process, we irradiated fibroblasts derived from an XP variant patient, XP4BE, under conditions that allowed the cells various lengths of time for excision repair before the onset of DNA synthesis (S phase) and assayed the frequency of 6-thioguanine (TG)-resistant mutants. Cells synchronized by release from confluence (G 0 state ) and irradiated just prior to S phase showed a dose-dependent increase in mutants at very high frequencies; cells irradiated in early G 1, ≈ 12 h before the onset of S phase, showed frequencies 4 times lower. Cells irradiated in the G 0 state and allowed 24 h or 48 h for excision repair before the onset of S phase showed still lower frequencies. A comparison of the relative rates of decrease in mutant frequency with time for excision repair before the onset of S phase in XP variant cells and normal human fibroblasts after a dose of 4 or 6 J/m 2 showed that these were equal. However, for every time point, the frequency of mutants induced per dose of UV was significantly higher in the XP variant population than in the normal, suggesting that the XP variant cells have an abnormally error-prone process of replicating DNA on a template containing unexcised lesions or normal cells are by-passing many of such lesions using an error-free process. A similar comparative study in synchronized populations of XP4BE cells and normal cells, using the anti 7,8-diol-9,10-epoxide of benzo[ a]pyrene, showed that excision repair prior to the onset of S phase also decreased the frequency of mutants induced in XP variant cells by this agent. But for every dose and time point, the frequencies induced in XP4BE cells and normal cells were identical. Thus, the hypermutability of the XP4BE cells was specific to UV radiation-induced DNA lesions.

Computer Aided Design System for Instrumentation Engineering

The utilization of computer in the design work is particularly based on three facts: fastness, errorfree processing and integration of data. Great advantages can be achieved by integrating the data management system and the graphic system into a functional unit.The paper describes a computer-aided instrumentation design system developed by EKONO Consulting Engineers. In step by step progressing design work, the data entered earlier are utilized in all phases of planning. The work advances by adding and revising the data stored in the system. The same piece of information can appear in several forms. Any data is, however, fed and updated only once. The systems support engineers throughout the design process and produce most of the documents needed.

Dynamic Measurement of Computer Generated Image Segmentations

This paper introduces a general purpose performance measurement scheme for image segmentation algorithms. Performance parameters that function in real-time distinguish this method from previous approaches that depended on an a priori knowledge of the correct segmentation. A low level, context independent definition of segmentation is used to obtain a set of optimization criteria for evaluating performance. Uniformity within each region and contrast between adjacent regions serve as parameters for region analysis. Contrast across lines and connectivity between them represent measures for line analysis. Texture is depicted by the introduction of focus of attention areas as groups of regions and lines. The performance parameters are then measured separately for each area. The usefulness of this approach lies in the ability to adjust the strategy of a system according to the varying characteristics of different areas. This feedback path provides the means for more efficient and error-free processing. Results from areas with dissimilar properties show a diversity in the measurements that is utilized for dynamic strategy setting.

Repair of lethal, mutagenic and transforming damage induced by X-rays in golden hamster embryo cells.

We compared the frequencies of mutation and morphological transformation in relation to the repair of potentially lethal damage (PLD) and sublethal damage (SLD) in X-irradiated golden hamster embryo (GHE) cells. The repair of PLD induced in GHE cells by X-rays was shown to be an error-free process which reduced the frequency of mutation and morphological transformation. However, if cells were irradiated with doses divided into two halves separated by various time intervals to study repair of SLD, the frequency of mutants and morphological transformants depended upon the length of the interval between the doses. If the interval was 2 h, the frequency of mutants and transformants was lower than if the total dose were delivered in a single exposure. If the interval was 4 h, these frequencies were higher than the latter. The transformation frequencies remained high even if this interval was increased to 12 h, but the frequency of mutants decreased again. Thus, enhancement of mutant or transformant frequency by a second exposure to X-rays, 4 h after the first irradiation, correlated with recovery of DNA synthesis in the X-irradiated cell populations. Studies with synchronized cells also showed that induction of mutants and transformants was highest in cells irradiated at the DNA synthesis (S) phase. These results suggest that DNA synthesis plays a critical role in the process of X-ray-induced morphological transformation and mutagenesis.

Mechanisms of Teratogenesis

The mechanisms of teratogenesis faJl into two broad categories based on the etiology of the congenital malformations: (a) errors in genetic programming based on deviations in the genotype of the embryo or the low probability for error of a normal genotype; and (b) environmental agents or factors that interact with an embryo during the period of development (drugs, chemicals, radiation, hyperthermia, infections, abnormal maternal metabolic states, or mechanical factors). The etiology of human malformations includes both genetic and environ­ mental factors, but it also includes a large category labelled unknown (Table 1). Many geneticists believe that a significant proportion of congenital malforma­ tions of unknown etiology are polygenic (l, 2). Thus, malformations with an increased recurrent risk, such as cleft palate, anencephaly, spina bifida, con­ genital heart disease, pyloric stenosis, talipes equinovarus, and congenital dislocation of the hip, fit the criteria for polygenic inherited disease. They also happen to fit the criteria for multifactorial disease (1, 3). Included in the unknown category are malformations that occur spontaneously at a very low frequency simply on the basis of the probability of spontaneous errors of development. Thus, particular genotypes have an inherent malformation rate based on the fact that embryonic development is a very complicated process and has a variable but low probability of going awry just as DNA duplication, for example, is not an error-free process. The explanation for the etiology and mechanisms of teratogenesis for this large group of malformations with un­ known etiology will depend upon identifying the genes involved in polygenic or plurogenic processes, identifying the interacting genetic and environmental

Influence of confluent holding time on UV light mutagenesis in human diploid fibroblasts

We have investigated the induction of mutants resistant to 6-thioguanine (6TG) following 254 nm ultraviolet light exposure of density-inhibited cultures of human diploid fibroblasts. Phenotypic expression of 6TG resistance was maximal within 9 days and remained stable through 19 days after irradiation. In reconstruction studies, complete recovery of 6TG-resistant mutants occurred at cell densities of up to 35 000 cells per 100-mm petri dish. The induced mutation frequency increased linearly with dose over the range of 3–9 J/m 2; the D 0 of the survival curve was 4.2 J/m 2. Delaying subculture to low density for 1.5–24 h after irradiation produced unexpected alterations in induced mutation frequencies. An increase in UV-induced mutations of approximately 3-fold was observed in cultures maintained in confluence for 3 h. This trend was reversed with longer holding times: the mutation frequency declined sharply in cultures held for 6 h compared to the 3-h value, and thereafter showed a steady and gradual diminution to background levels. These data suggest that the repair of potentíally mutagenic damage is a complex phenomenon which can lead to an increase or decrease in mutation frequency as a function of holding time. Although the decline in mutation frequency observed following longer holding intervals is consistent with the notion of an error-free process, we hypothesize that the increased mutation frequency produced by a short holding period reflects the existence of a cell-mediated process which enhances the mutagenic potential of at least some UV-induced DNA photoproducts.

Replication and mutagenesis of irradiated single-strand phage DNA.

Mutagens may be divided into two classes on the basis of the mechanism of their mutagenic action and the requirement for cellular repair genes for their mutagenic effect. Direct mutagens cause subtle modifications of the bases in DNA (or its precursors) and give rise to mutations by mispairing during DNA replication. Direct mutagens do not require cellular DNA repair activity for their mutagenic effect (i.e. act independently of the bacterial recA and lexA genes) and normally have a high degree of mutagenic specificity, reflecting the specificity of mispairing. Examples of direct mutagens are deaminating agents, such as hydroxylamine and bisulfite ; some base analogs, such as 2-aminopurine ; and alkylating agents, such as ethylmethane sulfonate. Indirect mutagens destroy the coding properties of DNA templates, thereby blocking DNA replication, and, unless repaired, are lethal. In situations where there is a redundancy in the DNA sequence information error-free repair processes may eliminate (e.g. by excision repair) or ‘tolerate’ (e.g. by postreplication recombinational exchanges) these lesions. However, when the mutagen destroys a unique piece of DNA sequence information, repair by error-free processes is no longer possible, and recA lexA-dependent error-prone repair is the only alternative to lethality. Such a situation may occur when a non-coding lesion is located within an overlapping excision gap (1), or in an overlapping gap in nascent DNA (2), or in a single-stranded genome (3). Examples of indirect mutagens are ultraviolet (UV) and ionizing radiation and many chemical mutagens and carcinogens, such as aflatoxin B1, benzo(a)pyrene, and mitomycin C.

Elimination of MNU-induced mutational lesions in V79 Chinese hamster cells.

Studies were performed on the non-linear dose response for gene mutations induced by low doses of monofunctional methylating agents in V79 Chinese hamster cells. When treatment with methylnitrosourea was applied at the beginning of the S phase in synchronized cells, a linear dose-response curve was obtained, whereas application of the dose after gene replication resulted in a strong reduction of the number of induced mutations. Additional time for repair resulted in reduced dose response of MNU, indicating that an error-free repair process operates on methylated DNA in V79 Chinese hamster cells.

Host cell reactivation by excision repair is error-free in human cells

Do host cell repair processes affect the mutagenesis of UV-irradiated virus in human cells? The answer was obtained by investigating the mutagenesis of UV-irradiated herpes simplex virus after the irradiated virus was grown in human cells that possess normal repair capacity (normal) or lack excision repair (XPA) or post-replication repair (XP var). Evidence is presented which indicate that XPA cells express no host cell reactivation, while XP var cells express the normal level. Viral mutagenesis was measured as the fraction of the progeny of the surviving virus capable of plaque formation in the presence of iododeoxycytidine. In the normal and XPA cells mutagenesis of the irradiated virus increased linearly with UV exposure. The UV exposure needed to yield a given mutagenesis level for virus grown in XPA cells was much lower than that for virus grown in normal cells. However, when the mutation frequencies were compared at similar virus survival levels, the data from virus grown in normal cells and in XPA cells were indistinguishable. Mutagenesis in XP var cells increased as dose squared and was similar in magnitude to that in normal cells. Thus the excision repair of normal cells which provided host cell reactivation by removing lethal UV damage also removed mutagenic lesions from the virus with the same efficiency, while the repair deficiency of XP var cells had a minor role in host cell reactivation and in mutagenesis. This demonstrates that in human cells host cell reactivation by excision repair is primarily an error-free process.

Antimutators of mitochondrial and nuclear DNA in Saccharomyces cerevisiae. Relationship with gamma-ray sensitivity.

In Saccharomyces cerevisiae ten antimutator mutants have been isolated. The spontaneous occurrence of mitochondrial mutants resistant to erythromycin, oligomycin, and diuron is decreased 2-60-fold in these strains. The rate of forward and reverse spontaneous mutations of the nuclear genome is also reduced. The meiotic progenies arising from the crosses of seven mutants (LB1, LB2, LB4, LB5, LB6, LB7, LB10) with an isogenic parental strain exhibit 2 : 2 segregations and therefore are the result of mutations in a single nuclear gene. The six mutants LB1, LB2, LB4, LB6, LB7, LB10 are semidominant and determine six complementation groups. The mutant LB5 is dominant and therefore cannot be assigned to any complementation group. The mutants, LB1, LB4 and LB10 are gamma-ray sensitive and, by tetrad analysis, it has been shown that gamma-ray sensitivity and spontaneous antimutability are the result of a single nuclear gene mutation. The other three mutants LB3, LB8, and LB9 exhibit complex tetrad segregations, typical of cytoplasmic inheritance and do not complement each other. However, although the mutations are semidominant, it has not been possible to detect any antimutator cytoductant among some 500 cytoductants carrying the karl-1 nucleus. These results suggest that either several nuclear genes are involved in the expression of the antimutator phenotype or that the antimutator gene is located on nonchromosomal elements of the nucleus. The present study leads to the conclusion that a large number of nuclear genes are able to control simultaneously the spontaneous mutation rate of nuclear and mitochondrial genes. Since out of the ten antimutator mutants, three are also deficient in the repair of gamma-ray damage, it is also concluded that spontaneous and gamma-ray-induced lesions of DNA can be repaired by the same error-free process.

Inducible DNA polymerase in cultured rat fibroblasts treated with skin carcinogen mitomycin C

DNA polymerases are involved in the filling of excision [ 1,2] and postreplication [3,4] single-strand gaps in DNA in ultraviolet-irradiated bacteria. Gapfilling with the constitutive DNA polymerase is an error-free process that cannot be the source of ultraviolet-induced mutations [ 51. It has been reported that heat treatment of tif strain of E. coli causes the appearance of a mutagenic DNA polymerase responsible for the fixation of premutational lesions during heat-induced ‘SOS repair’ [6] . An inducibility has been shown for minor branch of postreplication repair in E. coli [7] , Micrococcus luteus [8] and in human fibroblasts [9] . The question remains: Is inducible repair in mammalian cells coupled with the induction of a DNA polymerase with a reduced fidelity in repair replication? Mitomycin C (MMC) studied extensively for its mutagenic [lo] and carcinogenic [ 111 properties can induce in E. cob the synthesis of X protein [ 121. This property usually correlates with the ability of a chemical to induce SOS repair [5] . In connection with the question above the observation of an increased DNA polymerase activity in extracts from MMC-treated mammalian cells [ 13,141 is of interest. We therefore investigated the origin and reaction properties of the DNA polymerase in cells affected by MMC.

Automatic printed circuit board routeing with intermediate control of the tracking: APRICOT

For several years computers have been used to produce artwork for printed circuit board manufacturer in basically two different modes. In the automatic mode the computer decides most, if not all, of the track routes and relies on the human to add the unrouted connections. This can be done in an essentially offline manner but it is error-prone. In the interactive mode, which may follow the application of an automatic program, the unrouted connections are manipulated on a graphical display. This is essentially an error-free process but it is performed online. This paper describes a new approach to automatic routeing that allow ‘error-free’ manipulation of the routeing in an offline manner. The automatic routine uses a planned, as opposed to a search, technique and this allows intermediate control (offline) of the automatic routeing process. It also enables the user to change component positions at any time during the routeing process in an offline mode.

Cognate Recognition across Dialects

Abstract Synchronic cognates are subject to recognition failures across dialects, if processing through different determining features leads to phonemic misclassification. But phonemes common to all dialects can also cause recognition failures if they differ in function. These findings support a theoretical distinction between phone, phoneme, and morpho phone, the last being an essential unit for the cognate reconstruction necessary for error-free processing of unfamiliar dialects.