Victor Nigon

Quantitative estimations of chloroplast DNA in bleached mutants of Euglena gracilis

SummaryThe level of chloroplast DNA has been estimated in bleached mutants of Euglena by the increase of the renaturation rate of a radioactive chloroplast DNA probe in response to the addition of total mutant DNA.Two classes of bleached mutants differ from each other by their level of chloroplast DNA:- a few bleached mutants contain chloroplast DNA in amounts similar to those of wild type strains; all sequences of the wild type chloroplast DNA seem to be present but in non stoechiometric proportions.- most bleached mutants have about 100 to 1,000 times less chloroplast DNA than wild type cells. In these mutants chloroplast DNA sequences form two frequency classes:- one class has about 5 to 12% of the complexity of the wild type genome; these sequences are reiterated from 30 to 160 times per cell and hybridize with wild type ribosomal cistrons. They are expressed as chloroplast ribosomal RNA in all bleached mutants analyzed so far.- the second class shows at least 40% of the complexity of the wild type genome; these sequences are present in 2 to 7 copies per cell.

Comparative studies of chloroplastic and nuclear DNA repair abilities after ultraviolet irradiation of Euglena gracilis

SummaryStudies of nuclear and chloroplastic-DNA repair after ultraviolet irradiation of Euglena gracilis show that photoreactivation is very efficient at both the nuclear and chloroplastic level. Liquid-holding or split-dose experiments and treatment with caffeine reveal, furthermore, that dark-repair is very efficient in nuclear DNA but not in chloroplastic DNA (ctDNA). The possibility of a chloroplastic dark-repair of restricted efficiency is discussed.Determination of chloroplastic DNA content by reassociation kinetics indicates that an important degradation follows UV irradiation during liquid holding in the dark.

Modifications of chloroplast DNA during streptomycin induced mutagenesis in Euglena gracilis

SummaryThe evolution of chloroplast DNA was analysed during streptomycin induced mutagenesis in Euglena gracilis strain bacillaris and strain Z. In addition to a massive reduction of the cellular level of chloroplast DNA, several structural modifications have been observed in early stages of mutagenesis but they are generally eliminated during the later stages. The ribosomal cistrons are regularly rearranged: two of the three tandemly arranged cistrons occuring in wild type chloroplast DNA decrease while the third one is relatively more conserved and amplified during mutagenesis and in bleached mutants.

Minicircular DNA having sequence homologies with chloroplast DNA in a bleached mutant of Euglena gracilis

SummaryChloroplast DNA was isolated from total cellular DNA of a bleached mutant of Euglena gracilis (Y3BUD) by enrichment of the light component (p = 1.686) by repeated CsCl equilibrium centrifugations. Electron microscope visualization of this DNA showed minicircular DNA molecules in addition to large circular molecules (42 pm) identical to wild type chloroplast DNA. They were heterogenous in size and their contour lengths ranged from 0.8 to 8.5 μm. Fractionnation by agarose gel electrophoresis gave several discrete bands. Some of them hybridized with pure chloroplast DNA and with several cloned chloroplast DNA fragments, particularly to ribosomal fragments, while others did not show homology with chloroplast DNA being probably of extrachloroplatic origin.

Rearrangement of chloroplast ribosomal cistrons by unequal crossing-over in Euglena gracilis.

SummaryThe chloroplast DNA of a wild type photosynthetic variant of Euglena gracifs (ATCC n° 10616) with five ribosomal cistrons has been analyzed by restriction mapping. The results complete the electron microscope study of Koller and Delius (MGG 188, 305, 1982); they support a model of formation of the variant DNA by rearrangement of the wild type ribosomal cistrons through unequal crossing-over. The recombination sites have been determined. The recombination model proposed also explains the formation of the “Z-S” variant with a single ribosomal cistron (Wurtz and Buetow 1981).

Somatic segregation and rate of greening after ultraviolet irradiation of Euglena gracilis

Summary1.During multiplication of irradiated cells, a segregation may take place between bleached cells, whose progeny is unable to green, and green ones. Some of the green cells give progenies exclusively made of green cells; the progeny of others is partly composed of bleached cells.2.If one assumes that greening results from the activity of functional units endowed with genetic continuity (Plastidial Segregating Units=PSU), segregation of these units seems to occur according to a model involving random sorting out during the three first divisions. During the following divisions, functional units seem to multiply faster than those impaired by irradiation.3.The greening rate of colonies issued from irradiated cells seems to be conditioned mostly by the number of functional PSU remaining in the mother cell of the colony.

Fitting survival curves with theoretical models; goodness-of-fit tests and parameter estimation. application to the clonal survival of Chlorella and the chloroplastic survival of Euglena gracilis after irradiation

Abstract (1) Maximum-likelihood methods are proposed for fitting survival curves with theoretical models. The methods give estimates of the parameters and goodness-of-fit tests. (2) The underlying theory is reviewed. (3) Application of these methods to the chloroplastic survival of Euglena gracilis led to the following results: Different models, based on target theory, give an adequate fit to the observations when each experiment is considered separately. However, there is a large variation between the estimate of the number of targets obtained from different experiments. Lack of a satisfactory explanation for this variation thus throws doubt on the validity of these simplest models. With more complex models one can postulate a constant number of target sites. These models consider the target sites as the sites of secondary effects for which variation between experiments is more easily explained. However, these models contain additional parameters which cannot be estimated from the available data, thus depriving the models of predictive value. If it were possible to obtain further experimental data from which these additional parameters could be estimated, then the predictive value of the models would be restored.

Production of plastidial antibiotic-resistant mutants by ultraviolet irradiation of Euglena gracilis

SummaryUltraviolet-irradiation of Euglena gracilis drastically increases the proportion of green colonies resistant to chloroplast-inhibiting antibiotics. Some green colonies persist at high UV doses. These results might indicate the occurrence in the chloroplastic DNA of repair processes, of which one could be error-prone.