Philippe Heizmann

Sporophytic self-incompatibility systems: Brassica S gene family.

Publisher Summary Incompatibility systems represent an important element of control of sexual reproduction in higher plants. By preventing self-pollination they act to promote outbreeding and therefore may exert profound effects on the genetic structures of populations and perhaps the evolution of species. Incompatibility phenotypes are genetically controlled by a single, major gene designated “S.” Examples include the gametophytically controlled system of the Solanaceae and the sporophytic system that is characteristic of the Brassicaceae. In the species Brassica oleracea, a single highly polymorphic locus S determines both the incompatibility phenotype of pollen via expression in the diploid tapetum of the anthers and also that of the stigma papillae cells. The system would thus appear to be founded on a specialized form of cell-to-cell interaction between the surface of the pollen grain and that of the style. Any identity in expressed S alleles between these two partners results in a rapid and specific inhibition of self-pollen germination at the stigmatic surface. The high relative abundance and strict developmental and tissue-specific regulation of the Brassica S locus-specific glycoproteins (SLSGs) enable a simple strategy for the molecular cloning of the SLG-specific messages.

Changes in protein patterns and protein synthesis during anther development in Brassica oleracea

Summary To study male gametophyte development in Brassica oleracea L., the various stages of microspores and pollen in fresh anthers were assessed with the use of fluorescent dyes and by cytological observations. Developmental variations in SDS-PAGE protein patterns of anthers and pollen grains were then analysed from the late vacuolate microspore stage to the end of the pollen-maturation period. Total protein staining and concanavalin A-binding glycoprotein detection showed that a specific set of developmental polypeptides appears during the tricellular pollen stages. Protein synthesis was studied by [ 31 S] methionine incorporation, SDS-PAGE, fluorography. Two periods of protein synthetic activity were detected: the first one corresponding to the microspore and bicellular pollen stages; the second one corresponding to the mid, late, mature tricellular pollen stages. During this second period, new polypeptides were synthesized and most of them could be correlated with the developmental polypeptides that appeared in total and Con A-binding protein patterns. These results suggest the occurrence of a metabolic reorientation after the second pollen mitosis, at the time of sperm cell maturation. The biochemical data are discussed in terms of diploid and haploid genome expression.

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).

Sequence of a mitochondrial plasmid of sunflower (Helianthus annuus) and its relationship to other mitochondrial plasmids

Abstract A circular plasmid called plT is found in mitochondria of sunflower ( Helianthus annuus ). Its nucleotide sequence has been determined and analyzed in order to understand its possible role and origin. The nucleotide sequence exhibits a 159 base pair region with highly organized repeats. The sequence of plT shows no major homology with other plasmids of higher plants although organized patterns are present in the circular plasmids. We also report the existence of two related mitochondrial plasmids in a line of H. annuus and in the H. petiolaris fallax species. In addition, plT plasmid has been detected in total cellular DNA of male-sterile sunflower with a copy number of a hundred times lower than in the male-fertile one.

Sporophytic Self-Incompatibility

The anemogamic or entomogamic pollination processes passively sustained by land plants and the wide occurence of hermaphrodism among most plant families, convergerge to favor a natural propensity for self-pollination, inbreeding and homozigosity. In fact, many plants species are known to be able to identify and to reject their own pollen. Indeed, Darwin already described an heteromorphic form of self-incompatibility (SI) in Primula, where size differences between style and anthers (heterostyly) severely hamper self-pollination. Since then, a large number of species was identified to be able to recognize and to reject self-pollen exclusively by physiological reactions (homomorphic SI). Population geneticists consider that the natural selection of SI early in the rise of flowering plants has been determinant for their successful espansion and domination in the present living world (Whitehouse, 1950).

Expression of SLG self-incompatibility genes in the anthers of Brassica oleracea: new members in the SLG gene family

Abstract The model of self-incompatibility proposed by Lewis in 1960 assumed that identical or similar ‘S-products’ produced symmetrically in the male and female flower tissues are involved in self recognition. This review analyzes the results of two converging methodologies used by two different groups, to demonstrate the transcription, suspected to be very low, of the Brassica S-gene in the anthers: (1) Chimeric constructions made of the β-glucuronidase (GUS) and of the diptheria toxin A (DT-A) reporter genes, placed under the control of the promoter region of an SLG gene from Brassica oleracea were used to transform Arabidopsis thaliana and Nicotiana tabacum . The transgenic plants showed a high level of expression of the reporter genes in the mature female tissues, and a low level of activity of these genes in the male tissues. (2) The polymerase chain reaction (PCR) allowed to demonstrate amplifications from S-transcripts from anthers of Brassica oleracea at early microsporogenesis, having high sequence homologies with the S-transcripts found in the mature stigmas. These two combined sets of data characterize the control segments (by trasngeny) and the coding segment (PRC) of the SLG gene; they tend thus to support the model proposed several years ago by Lewis. In addition, some PRC reactions amplified several transcripts from vegetative tissues (roots, leaves), displaying various degrees of homology with the SLG sequence. These results suggest that the S-complex might be larger than previously described, involved in various kinds of plant cell signaling systems, and expressed in many different tissues.