Identification of ASYNAPTIC4, a Component of the Meiotic Chromosome Axis

International audienceDuring the leptotene stage of prophase I of meiosis, chromatids become organized into a linear looped array via a protein axis that forms along the loop bases. Establishment of the axis is essential for the subsequent synapsis of the homologous chromosome pairs and the progression of recombination to form genetic crossovers. Here, we describe ASYNAPTIC4 (ASY4), a meiotic axis protein in Arabidopsis (Arabidopsis thaliana). ASY4 is a small coiled-coil protein that exhibits limited sequence similarity with the carboxyl-terminal region of the axis protein ASY3. We used enhanced yellow fluorescent protein-tagged ASY4 to show that ASY4 localizes to the chromosome axis throughout prophase I. Bimolecular fluorescence complementation revealed that ASY4 interacts with ASY1 and ASY3, and yeast two-hybrid analysis confirmed a direct interaction between ASY4 and ASY3. Mutants lacking full-length ASY4 exhibited defective axis formation and were unable to complete synapsis. Although the initiation of recombination appeared to be unaffected in the asy4 mutant, the number of crossovers was reduced significantly, and crossovers tended to group in the distal parts of the chromosomes. We conclude that ASY4 is required for normal axis and crossover formation. Furthermore, our data suggest that ASY3/ASY4 are the functional homologs of the mammalian SYCP2/SYCP3 axial components

Ribosomal RNA of Hyacinthus orientalis L. female gametophyte cells before and after fertilization

The nucleolar activity of Hyacinthus orientalis L. embryo sac cells was investigated. The distributions of nascent pre-rRNA (ITS1), 26S rRNA and of the 5S rRNA and U3 snoRNA were determined using fluorescence in situ hybridization (FISH). Our results indicated the different rRNA metabolism of the H. orientalis female gametophyte cells before and after fertilization. In the target cells for the male gamete, i.e., the egg cell and the central cell whose activity is silenced in the mature embryo sac (Pięciński et al. in Sex Plant Reprod 21:247–257, 2008; Niedojadło et al. in Planta doi:10.1007/s00425-012-1599-9, 2011), rRNA metabolism is directed at the accumulation of rRNPs in the cytoplasm and immature transcripts in the nucleolus. In both cells, fertilization initiates the maturation of the maternal pre-rRNA and the expression of zygotic rDNA. The resumption of rRNA transcription observed in the hyacinth zygote indicates that in plants, there is a different mechanism for the regulation of RNA Pol I activity than in animals. In synergids and antipodal cells, which have somatic functions, the nucleolar activity is correlated with the metabolic activity of these cells and changes in successive stages of embryo sac development

Integrating the Genetic and Physical Maps of Arabidopsis thaliana: Identification of Mapped Alleles of Cloned Essential (EMB) Genes

The classical genetic map of Arabidopsis includes more than 130 genes with an embryo-defective (emb) mutant phenotype. Many of these essential genes remain to be cloned. Hundreds of additional EMB genes have been cloned and catalogued (www.seedgenes.org) but not mapped. To facilitate EMB gene identification and assess the current level of saturation, we updated the classical map, compared the physical and genetic locations of mapped loci, and performed allelism tests between mapped (but not cloned) and cloned (but not mapped) emb mutants with similar chromosome locations. Two hundred pairwise combinations of genes located on chromosomes 1 and 5 were tested and more than 1100 total crosses were screened. Sixteen of 51 mapped emb mutants examined were found to be disrupted in a known EMB gene. Alleles of a wide range of published EMB genes (YDA, GLA1, TIL1, AtASP38, AtDEK1, EMB506, DG1, OEP80) were discovered. Two EMS mutants isolated 30 years ago, T-DNA mutants with complex insertion sites, and a mutant with an atypical, embryo-specific phenotype were resolved. The frequency of allelism encountered was consistent with past estimates of 500 to 1000 EMB loci. New EMB genes identified among mapped T-DNA insertion mutants included CHC1, which is required for chromatin remodeling, and SHS1/AtBT1, which encodes a plastidial nucleotide transporter similar to the maize Brittle1 protein required for normal endosperm development. Two classical genetic markers (PY, ALB1) were identified based on similar map locations of known genes required for thiamine (THIC) and chlorophyll (PDE166) biosynthesis. The alignment of genetic and physical maps presented here should facilitate the continued analysis of essential genes in Arabidopsis and further characterization of a broad spectrum of mutant phenotypes in a model plant

The arabidopsis DNA polymerase δ has a role in the deposition of transcriptionally active epigenetic marks, development and flowering

DNA replication is a key process in living organisms. DNA polymerase α (Polα) initiates strand synthesis, which is performed by Polε and Polδ in leading and lagging strands, respectively. Whereas loss of DNA polymerase activity is incompatible with life, viable mutants of Polα and Polε were isolated, allowing the identification of their functions beyond DNA replication. In contrast, no viable mutants in the Polδ polymerase-domain were reported in multicellular organisms. Here we identify such a mutant which is also thermosensitive. Mutant plants were unable to complete development at 28°C, looked normal at 18°C, but displayed increased expression of DNA replication-stress marker genes, homologous recombination and lysine 4 histone 3 trimethylation at the SEPALLATA3 (SEP3) locus at 24°C, which correlated with ectopic expression of SEP3. Surprisingly, high expression of SEP3 in vascular tissue promoted FLOWERING LOCUS T (FT) expression, forming a positive feedback loop with SEP3 and leading to early flowering and curly leaves phenotypes. These results strongly suggest that the DNA polymerase δ is required for the proper establishment of transcriptionally active epigenetic marks and that its failure might affect development by affecting the epigenetic control of master genes.Fil: Iglesias, Francisco Manuel. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Investigaciones Bioquimicas de Buenos Aires; Argentina. Fundación Instituto Leloir; ArgentinaFil: Bruera, Natalia Alejandra. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Investigaciones Bioquimicas de Buenos Aires; Argentina. Fundación Instituto Leloir; ArgentinaFil: Dergan Dylon, Leonardo Sebastian. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Investigaciones Bioquimicas de Buenos Aires; Argentina. Fundación Instituto Leloir; ArgentinaFil: Marino, Cristina Ester. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Investigaciones Bioquimicas de Buenos Aires; Argentina. Fundación Instituto Leloir; ArgentinaFil: Lorenzi, Hernán. J. Craig Venter Institute; Estados UnidosFil: Mateos, Julieta Lisa. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Investigaciones Bioquimicas de Buenos Aires; Argentina. Fundación Instituto Leloir; Argentina. Max Planck Institute for Plant Breeding Research; AlemaniaFil: Turck, Franziska. Max Planck Institute for Plant Breeding Research; AlemaniaFil: Coupland, George. Max Planck Institute for Plant Breeding Research; AlemaniaFil: Cerdan, Pablo Diego. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Investigaciones Bioquimicas de Buenos Aires; Argentina. Fundación Instituto Leloir; Argentina. Universidad de Buenos Aires. Departamento de Ciencias Exactas; Argentin

Differing Requirements for RAD51 and DMC1 in Meiotic Pairing of Centromeres and Chromosome Arms in Arabidopsis thaliana

During meiosis homologous chromosomes pair, recombine, and synapse, thus ensuring accurate chromosome segregation and the halving of ploidy necessary for gametogenesis. The processes permitting a chromosome to pair only with its homologue are not fully understood, but successful pairing of homologous chromosomes is tightly linked to recombination. In Arabidopsis thaliana, meiotic prophase of rad51, xrcc3, and rad51C mutants appears normal up to the zygotene/pachytene stage, after which the genome fragments, leading to sterility. To better understand the relationship between recombination and chromosome pairing, we have analysed meiotic chromosome pairing in these and in dmc1 mutant lines. Our data show a differing requirement for these proteins in pairing of centromeric regions and chromosome arms. No homologous pairing of mid-arm or distal regions was observed in rad51, xrcc3, and rad51C mutants. However, homologous centromeres do pair in these mutants and we show that this does depend upon recombination, principally on DMC1. This centromere pairing extends well beyond the heterochromatic centromere region and, surprisingly, does not require XRCC3 and RAD51C. In addition to clarifying and bringing the roles of centromeres in meiotic synapsis to the fore, this analysis thus separates the roles in meiotic synapsis of DMC1 and RAD51 and the meiotic RAD51 paralogs, XRCC3 and RAD51C, with respect to different chromosome domains

Crossover interference mechanism: New lessons from plants

Plants are the source of our understanding of several fundamental biological principles. It is well known that Gregor Mendel discovered the laws of Genetics in peas and that maize was used for the discovery of transposons by Barbara McClintock. Plant models are still useful for the understanding of general key biological concepts. In this article, we will focus on discussing the recent plant studies that have shed new light on the mysterious mechanisms of meiotic crossover (CO) interference, heterochiasmy, obligatory CO, and CO homeostasis. Obligatory CO is necessary for the equilibrated segregation of homologous chromosomes during meiosis. The tight control of the different male and female CO rates (heterochiasmy) enables both the maximization and minimization of genome shuffling. An integrative model can now predict these observed aspects of CO patterning in plants. The mechanism proposed considers the Synaptonemal Complex as a canalizing structure that allows the diffusion of a class I CO limiting factor linearly on synapsed bivalents. The coarsening of this limiting factor along the SC explains the interfering spacing between COs. The model explains the observed coordinated processes between synapsis, CO interference, CO insurance, and CO homeostasis. It also easily explains heterochiasmy just considering the different male and female SC lengths. This mechanism is expected to be conserved in other species

Caractérisation génétique et moléculaire de mutants d'embryogenèse précoce d'Arabidopsis thaliana

Chez les plantes à fleurs, le développement des graines est initié par une double fécondation qui conduit à la formation de l'embryon et l'albumen. Ce processus a été décrit il a plus d'un siècle mais les mécanismes moléculaires par lesquels la fécondation active le programme embryogène sont encore peu compris. La voie la plus étudiée concerne la répression du développement de la cellule centrale avant la fécondation. Nous avons utilisé une approche de génétique classique dans le but de caractériser des gènes essentiels impliqués dans l'activation de l'embryogenèse précoce chez la plante modèle Arabidopsis thaliana. Les analyses ont porté sur des mutants embryons défectueux (emb) bloqués dans les toutes premières étapes de l'embryogenèse et étiquetés par une insertion d'ADN-T. Ces mutations sont sporophytiques et récessives. Notre étude s'est concentrée sur la caractérisation phénotypique et moléculaire des mutants zeus (zeu) arrêtés après l'élongation du zygote et des mutants cyclops (cyl) bloqués après la première division asymétrique du zygote. Un gène ZEU et deux gènes CYL ont été identifiés et vérifiés par complémentation fonctionnelle. Les profils d'expression de chaque gène suggèrent une régulation transcriptionnelle au moment de la fécondation cohérente avec leur rôle dans l'activation du développement de la graine. Les protéines ZEU1 et CYL2 sont impliquées dans la réplication de l'ADN. La fonction potentielle de CYL1 suggère qu'une voie de signalisation similaire à celle nécessaire pour l'induction de l'embryogenèse somatique serait impliquée in vivo pour l'induction du développement de l'embryon zygotique.In flowering plants, the seed development is initiated by a double fertilization which induces the formation of the embryo and the endosperm. This process has been described more than one century ago but the molecular mechanisms by which the fertilization activates the embryogenic programm are poorly understood. The most studied pathway concerns the repression of the central cell development before fertilization. We have used a forward genetic approach in order to characterize essential genes involved in the activation of early embryogenesis in the model plant Arabidopsis thaliana. The analysis have been made on embryo-defective (emb) mutants arrested during the first steps of embryogenesis and tagged by a T-DNA insertion. The mutations are sporophytic and recessive. We focused on the phenotypic and molecular characterization of zeus (zeu) mutants arrested after the zygote elongation and cyclops (cyl) mutants stopped after the first assymetric division of the zygote. One ZEU and two CYL genes have been identified and confirmed by fonctional complementation. The expression profil of each gene suggest a transcriptional regulation at the moment of fertilization consistent with their role in the activation of the seed development. The ZEU1 and CYL2 proteins are involved in DNA replication. The putative function of CYL1 suggests that a signalling pathway similar to the one involved in the induction of somatic embryogenesis could be implicated in vivo to induce the zygotic embryo development.?PERPIGNAN-BU Sciences (661362101) / SudocSudocFranceF