Michel Dron

Structural and transcription analysis of two homologous genes for the P700 chlorophyll a-apoproteins in Chlamydomonas reinhardii: evidence for in vivo trans-splicing.

The two homologous genes for the P700 chlorophyll a‐apoproteins (ps1A1 and ps1A2) are encoded by the plastom in the green alga Chlamydomonas reinhardii. The structure and organization of the two genes were determined by comparison with the homologous genes from maize using data from heterologous hybridizations as well as from DNA and RNA sequencing. While the ps1A2 (736 codons) gene shows a continuous gene organization, the ps1A1 (754 codons) gene possesses some unusual features. The discontinuous gene is split into three separate exons which are scattered around the circular chloroplast genome. Exon 1 (86 bp) is separated by ∼50 kb from exon 2 (198 bp), which is located ∼ 90 kb apart from exon 3 (1984 bp). All exons are flanked by intronic sequences of group II. Transcription analysis reveals that the ps1A2 gene hybridizes with a 2.8‐kb transcript, while all exon regions of the ps1A1 gene are homologous to a mature mRNA of 2.7 kb. From our data we conclude that the three distantly separated exonic sequences of the ps1A1 gene constitute a functional gene which probably operates by a trans‐splicing mechanism.

Characterization of a chloroplast mutation in the psaA2 gene of Chlamydomonas reinhardtii

SummaryThe synthesis of polypeptides related to the CPI chlorophyll-protein complex of photosystem I has been studied by pulse-labeling experiments in twenty chloroplast mutants of Chlamydomonas reinhardtii. Three mutations of the same locus (Girard-Bascou 1987) result in the absence of these CPI-related polypeptides. Among these mutations one, (FUD26) leads to the synthesis of a new polypeptide presumed to be a truncated CPI apoprotein. The molecular characterization of this mutation in the psaA2 gene has been achieved by DNA sequencing the 3′ end of this gene. The FUD26 mutation is a 4 base pair deletion resulting in frameshift and premature termination of the protein.

The Arabidopsis SHAGGY-related protein kinase (ASK) gene family: structure, organization and evolution.

Higher plants contain a multigene family encoding proteins that share a highly conserved catalytic protein kinase domain about 70% identical to SHAGGY protein kinase (SGG) and glycogen synthase kinase-3 (GSK-3), respectively, from Drosophila and mammals. In this study we have characterized the structure and evolution of the Arabidopsis SHAGGY-related protein kinase (ASK) gene family. At least ten ASK genes are present per haploid genome of Arabidopsis. The genomic sequences of five ASK genes show a strikingly high conservation of intron positions and exon lengths. Phylogenetic analyses suggested that the Arabidopsis gene family contains at least three ancient classes of genes that diverged early in land plant evolution. The different classes may reflect specificity of substrates and/or biological functions. Eight out of the ten predicted ASK genes were mapped and shown to be dispersed over the five Arabidopsis chromosomes. A tentative model for the organization and evolution of the Arabidopsis ASK genes is presented.

Organelle genome stability in anther-derived doubled haploids of wheat (Triticum aestivum L., cv. ‘Moisson’)

SummaryChloroplast and mitochondrial compartments of a parental line of wheat (Triticum aestivum L., cv. ‘Moisson’) and its anther-derived doubled haploid lines have been analyzed and compared on the basis of their DNA restriction patterns. The results obtained show that no noticeable difference can be detected between doubled haploid lines and parental line at the level of ctDNA and mtDNA organization. It may be concluded that in vitro culture by itself does not systematically generate a cytoplasmic variation in germ cells.

VARIATION IN GENOME ORGANIZATION OF THE PLANT PATHOGENIC FUNGUS COLLETOTRICHUM LINDEMUTHIANUM

Abstract The genome structure of Colletotrichum lindemuthianum in a set of diverse isolates was investigated using a combination of physical and molecular approaches. Flow cytometric measurement of genome size revealed significant variation between strains, with the smallest genome representing 59% of the largest. Southern-blot profiles of a cloned fungal telomere revealed a total chromosome number varying from 9 to 12. Chromosome separations using pulsed-field gel electrophoresis (PFGE) showed that these chromosomes belong to two distinct size classes: a variable number of small (< 2.5 Mb) polymorphic chromosomes and a set of unresolved chromosomes larger than 7 Mb. Two dispersed repeat elements were shown to cluster on distinct polymorphic minichromosomes. Single-copy flanking sequences from these repeat-containing clones specifically marked distinct small chromosomes. These markers were absent in some strains, indicating that part of the observed variability in genome organization may be explained by the presence or absence, in a given strain, of dispensable genomic regions and/or chromosomes.

Diversity and dynamics of plant genome size: An example of polysomaty from a cytogenetic study of Tahitian vanilla (Vanilla ×tahitensis, Orchidaceae)

PREMISE OF THE STUDY Abnormal mitotic behavior with somatic aneuploidy and partial endoreplication were previously reported for the first time in the plant kingdom in Vanilla planifolia. Because vanilla plants are vegetatively propagated, such abnormalities have been transmitted. This study aimed to determine whether mitotic abnormalities also occur in Vanilla hybrid or are suppressed by sexual reproduction. METHODS Twenty-eight accessions of Vanilla ×tahitensis, one V. planifolia, and hybrid V. planifolia × V. ×tahitensis were analyzed by chromosome counts, cytometry, and fluorescent in situ hybridization of 18S-5.8S-26S rDNA. KEY RESULTS In a single root meristem of V. ×tahitensis, chromosome number varied from 22 to 31 in diploids (mean 2C = 5.23 pg), 31 to 41 in triploids (2C = 7.82 pg) and 43 to 60 in tetraploids (2C = 10.27 pg). Morphological diversity is apparently related to ploidy changes. Aneuploidy and partial (asymmetrical) endoreduplication were observed in root meristems of both V. ×tahitensis and the hybrid V. planifolia × V. ×tahitensis, but pollen grains had the euploid chromosome number (n = 15 in diploids). CONCLUSIONS Genome irregularities may be transmitted not only during vegetative propagation but also by sexual reproduction in Vanilla. However, there must be a complex regulation of genome size and organization between the aneuploidy in somatic tissues and subsequently euploid gametic tissue. This is a novel example of polysomaty with developmentally regulated partial endoreplication.

Is transcription of higher plant chloroplast ribosomal operons regulated by premature termination

Three stem‐loop structures (H1, H2, H3) can be formed in the leader transcript of the chloroplast rDNA of spinach, tobacco and maize. H2 and H3 partially overlap and cannot exist simultaneously. These potential hairpins lead us to postulate that higher plant chloroplast rDNA is regulated by premature termination. This mechanism could be controlled by the presence or absence of a ribosome translating an hypothetical leader peptide encoded in the rDNA leader sequence of these 3 higher plants.

Chloroplast Gene Function: Combined Genetic and Molecular Approach in Chlamydomonas Reinhardii

Chlamydanonas reinhardii is a green unicellular algae that is particularly well-suited for the study of chloroplast gene function and regulation. It contains a single, large chloroplast which comprises about 40% of the cell volume. Moreover, the life cycle of this alga involves sexual recanbination in which haploid gametes of opposite mating type (+ and −) fuse to form a diploid zygote (Fig. 1). At this point, the chloroplasts from both gametes fuse, but the chloroplast genaue is not inherited in mendelian fashion. Instead, the zygote retains only the chloroplast genome from the mating type + or maternal parent (Gillham, 1978). Fortunately, for those interested in chloroplast genetics, this uniparental maternal inheritance occurs about 95% of the time, so that approximately 5% of the zygotes are either uniparental paternal or biparental with respect to the chloroplast genome. And it is in these exceptional, biparental zygotes that recanbination occurs between the maternal and paternal chloroplast genes, at a frequency that allows for genetic mapping of chloroplast DNA. Thus, coupling the classical genetic approach and the ability to obtain and map mutants with the techniques of molecular biology and the ability to isolate, analyze and in some cases modify individual genes, their messages and their protein products, provides a powerful framework for studying the function and expression of chloroplast genes.

Light and developmental regulation of the Anp-controlled anthocyanin phenotype of bean pods

In the presence of the dominant allele of the Anp gene, bean pods present a purple-mottled phenotype. The purple pigmentation is variable from cell to cell in the pod epidermal layer and develops as a random mosaic. Three anthocyanidins, delphinidin, petunidin and malvidin, are involved in this purple pigmentation. Anthocyanins accumulated in vacuoles; anthocyanoplasts and cristal bodies were also observed occasionally. A developmental switch is a prerequisite for anthocyanin accumulation in the pods. This does not occur before day 4 after pollination and is controlled by light in competent pods. mRNAs for PAL, CHS, CHI, DFR and UFGT are induced in the pods, indicating that the general anthocyanin biosynthetic pathway is well conserved at both the biochemical and molecular levels in this species. mRNA steady-state level studies of PAL and CHS suggest that the light regulation occurs at the transcriptional level.

Structure and Expression of Chloroplast and Nuclear Genes in Chlamydomonas reinhardii

Chlamydomonas reinhardii, a facultative photoautotroph, can also be grown heterotrophically on acetate in the dark. Thus, mutants impaired in photosynthetic functions can be recovered as acetate requirers and their specific defects analyzed. While a part of the genetic information necessary for chloroplast function is found in the nucleus and displays classical Mendelian inheritance, another part is found in the chloroplast and undergoes non-Mendelian, uniparental transmission. In the occasional biparental zygotes, chloroplast gene markers can be observed to recombine. This unique property of Chlamydomonas has led to the derivation of a formal genetic map of the chloroplast genome (Sager 1972; Gilharn 1978). An extensive nuclear genetic map also exists (Harris 1982).