Sophie Le Panse

Auxin Metabolism and Function in the Multicellular Brown Alga Ectocarpus siliculosus

Ectocarpus siliculosus is a small brown alga that has recently been developed as a genetic model. Its thallus is filamentous, initially organized as a main primary filament composed of elongated cells and round cells, from which branches differentiate. Modeling of its early development suggests the involvement of very local positional information mediated by cell-cell recognition. However, this model also indicates that an additional mechanism is required to ensure proper organization of the branching pattern. In this paper, we show that auxin indole-3-acetic acid (IAA) is detectable in mature E. siliculosus organisms and that it is present mainly at the apices of the filaments in the early stages of development. An in silico survey of auxin biosynthesis, conjugation, response, and transport genes showed that mainly IAA biosynthesis genes from land plants have homologs in the E. siliculosus genome. In addition, application of exogenous auxins and 2,3,5-triiodobenzoic acid had different effects depending on the developmental stage of the organism, and we propose a model in which auxin is involved in the negative control of progression in the developmental program. Furthermore, we identified an auxin-inducible gene called EsGRP1 from a small-scale microarray experiment and showed that its expression in a series of morphogenetic mutants was positively correlated with both their elongated-to-round cell ratio and their progression in the developmental program. Altogether, these data suggest that IAA is used by the brown alga Ectocarpus to relay cell-cell positional information and induces a signaling pathway different from that known in land plants.

Yeast Functional Analysis: Identification of Two Essential Genes Involved in ER to Golgi Trafficking

We screened for genes potentially involved in the secretory and vacuolar pathways a collection of 61 yeast strains, each bearing an essential orphan gene regulated by the tetO7‐CYC1 promoter that can be down‐regulated by doxycycline. After down‐regulating the expression of these genes, we performed systematic Western blot analysis for markers of the secretory and vacuolar pathways that undergo post‐translational modifications in their intracellular trafficking. Accumulation of protein precursors, revealed by Western immunoblot analysis, indicates defects in the secretory pathway or in associated biochemical modifications. After screening the whole collection, we identified two genes involved in ER to Golgi trafficking: RER2, a cis‐prenyl transferase, and USE1, the function of which was unknown. We demonstrated that repression of USE1 also leads to BiP secretion, and therefore likely affects retrograde, in addition to anterograde, ER to Golgi trafficking. The collection also includes two essential genes involved in intracellular trafficking that were conveniently repressed without resulting growth or trafficking defects.

ETOILE Regulates Developmental Patterning in the Filamentous Brown Alga Ectocarpus siliculosus

By means of a combination of experimental and modeling approaches applied to the hyperbranching mutant étoile, cell–cell communication, likely mediated by novel transmembrane proteins that share similarities with metazoan Notch receptors, was shown to account for the establishment of filament patterning and cell differentiation in the filamentous brown alga Ectocarpus siliculosus. Brown algae are multicellular marine organisms evolutionarily distant from both metazoans and land plants. The molecular or cellular mechanisms that govern the developmental patterning in brown algae are poorly characterized. Here, we report the first morphogenetic mutant, étoile (etl), produced in the brown algal model Ectocarpus siliculosus. Genetic, cellular, and morphometric analyses showed that a single recessive locus, ETL, regulates cell differentiation: etl cells display thickening of the extracellular matrix (ECM), and the elongated, apical, and actively dividing E cells are underrepresented. As a result of this defect, the overrepresentation of round, branch-initiating R cells in the etl mutant leads to the rapid induction of the branching process at the expense of the uniaxial growth in the primary filament. Computational modeling allowed the simulation of the etl mutant phenotype by including a modified response to the neighborhood information in the division rules used to specify wild-type development. Microarray experiments supported the hypothesis of a defect in cell–cell communication, as primarily Lin-Notch-domain transmembrane proteins, which share similarities with metazoan Notch proteins involved in binary cell differentiation were repressed in etl. Thus, our study highlights the role of the ECM and of novel transmembrane proteins in cell–cell communication during the establishment of the developmental pattern in this brown alga.

Gyp5p and Gyl1p are involved in the control of polarized exocytosis in budding yeast

We report here elements for functional characterization of two members of the Saccharomyces cerevisiae Ypt/Rab GTPase activating proteins family (GAP): Gyp5p, a potent GAP in vitro for Ypt1p and Sec4p, and the protein Ymr192wp/APP2 that we propose to rename Gyl1p (GYp like protein). Immunofluorescence experiments showed that Gyp5p and Gyl1p partly colocalize at the bud emergence site, at the bud tip and at the bud neck during cytokinesis. Subcellular fractionation and co-immunoprecipitation experiments showed that Gyp5p and Gyl1p co-fractionate with post-Golgi vesicles and plasma membrane, and belong to the same protein complexes in both localizations. We found by co-immunoprecipitation experiments that a fraction of Gyp5p interacts with Sec4p, a small GTPase involved in exocytosis, and that a fraction of Gyl1p associates at the plasma membrane with the Gyp5p/Sec4p complexes. We showed also that GYP5 genetically interacts with SEC2, which encodes the Sec4p exchange factor. Examination of the gyp5Δgyl1Δ mutants grown at 13°C revealed a slight growth defect, a secretion defect and an accumulation of secretory vesicles in the small-budded cells. These data suggest that Gyp5p and Gyl1p are involved in control of polarized exocytosis.

Temperature, water activity and pH during conidia production affect the physiological state and germination time of Penicillium species

Conidial germination and mycelial growth are generally studied with conidia produced under optimal conditions to increase conidial yield. Nonetheless, the physiological state of such conidia most likely differs from those involved in spoilage of naturally contaminated food. The present study aimed at investigating the impact of temperature, pH and water activity (aw) during production of conidia on the germination parameters and compatible solutes of conidia of Penicillium roqueforti and Penicillium expansum. Low temperature (5°C) and reduced aw (0.900 aw) during sporulation significantly reduced conidial germination times whereas the pH of the sporulation medium only had a slight effect at the tested values (2.5, 8.0). Conidia of P. roqueforti produced at 5°C germinated up to 45h earlier than those produced at 20°C. Conidia of P. roqueforti and P. expansum produced at 0.900 aw germinated respectively up to 8h and 3h earlier than conidia produced at 0.980 aw. Furthermore, trehalose and mannitol assessments suggested that earlier germination might be related to delayed conidial maturation even though no ultra-structural modifications were observed by transmission electron microscopy. Taken together, these results highlight the importance of considering environmental conditions during sporulation in mycological studies. The physiological state of fungal conidia should be taken into account to design challenge tests or predictive mycology studies. This knowledge may also be of interest to improve the germination capacity of fungal cultures commonly used in fermented foods.

Redescription and phylogenetic analyses of Durchoniella spp. (Ciliophora, Astomatida) associated with the polychaete Cirriformia tentaculata (Montagu, 1808)

Microscopic and phylogenetic analyses were performed on endocommensal astome ciliates retrieved from the middle intestine of a marine cirratulid polychaete, Cirriformia tentaculata, collected in the bay of Roscoff (English Channel, Northwest French coast) and on the Southwest English coast. Three morphotypes of the astome genus Durchoniella were identified, two corresponding to described species (the type species Durchoniella brasili (Léger and Duboscq, 1904) De Puytorac, 1954 and Durchoniella legeriduboscqui De Puytorac, 1954) while a third morphotype remains undescribed. Their small subunit (SSU) rRNA gene sequences showed at least 97.2% identity and phylogenetic analyses grouped them at the base of the subclass Scuticociliatia (Oligohymenophorea), as a sister lineage to all astomes from terrestrial oligochaete annelids. Ultrastructural examination by transmission electron microscopy and fluorescence in situ hybridization analyses revealed the presence of endocytoplasmic cocci and rod-shaped bacteria surrounded by a very thin membrane. These endocytoplasmic bacteria may play a role in the association between endocommensal astome ciliates and cirratulid polychaetes inhabiting in anoxic coastal sediments.

Nigritoxin is a bacterial toxin for crustaceans and insects

The Tetraconata (Pancrustacea) concept proposes that insects are more closely related to aquatic crustaceans than to terrestrial centipedes or millipedes. The question therefore arises whether insects have kept crustacean-specific genetic traits that could be targeted by specific toxins. Here we show that a toxin (nigritoxin), originally identified in a bacterial pathogen of shrimp, is lethal for organisms within the Tetraconata and non-toxic to other animals. X-ray crystallography reveals that nigritoxin possesses a new protein fold of the α/β type. The nigritoxin N-terminal domain is essential for cellular translocation and likely encodes specificity for Tetraconata. Once internalized by eukaryotic cells, nigritoxin induces apoptotic cell death through structural features that are localized in the C-terminal domain of the protein. We propose that nigritoxin will be an effective means to identify a Tetraconata evolutionarily conserved pathway and speculate that nigritoxin holds promise as an insecticidal protein.The Tetraconata concept suggests that insects and crustaceans may share evolutionarily conserved pathways. Here, the authors describe the animal tropism and structure-function relationship of nigritoxin, showing that this protein is lethal for insects and crustaceans but harmless to other animals.

A novel species of the marine cyanobacterium Acaryochloris with a unique pigment content and lifestyle

All characterized members of the ubiquitous genus Acaryochloris share the unique property of containing large amounts of chlorophyll (Chl) d, a pigment exhibiting a red absorption maximum strongly shifted towards infrared compared to Chl a. Chl d is the major pigment in these organisms and is notably bound to antenna proteins structurally similar to those of Prochloron, Prochlorothrix and Prochlorococcus, the only three cyanobacteria known so far to contain mono- or divinyl-Chl a and b as major pigments and to lack phycobilisomes. Here, we describe RCC1774, a strain isolated from the foreshore near Roscoff (France). It is phylogenetically related to members of the Acaryochloris genus but completely lacks Chl d. Instead, it possesses monovinyl-Chl a and b at a b/a molar ratio of 0.16, similar to that in Prochloron and Prochlorothrix. It differs from the latter by the presence of phycocyanin and a vestigial allophycocyanin energetically coupled to photosystems. Genome sequencing confirmed the presence of phycobiliprotein and Chl b synthesis genes. Based on its phylogeny, ultrastructural characteristics and unique pigment suite, we describe RCC1774 as a novel species that we name Acaryochloris thomasi. Its very unusual pigment content compared to other Acaryochloris spp. is likely related to its specific lifestyle.

Toward Multiscale Modeling of Molecular and Biochemical Events Occurring at Fertilization Time in Sea Urchins

We review here previous theoretical and experimental works, which aim to model major events that occur at the time of fertilization in the sea urchin. We discuss works that perform experiments and develop hypotheses that link different scales of biological systems such as the intracellular Ca2+ concentration oscillations and the swimming behavior of sperm, the Ca2+ wave propagation and the fertilization membrane elevation of the egg, and the mRNA translational activation and the completion of the first mitotic division of the early embryo. The aim of this review is on one hand, to highlight the value of systems biology for understanding the mechanisms associated with fertilization and early embryonic development in sea urchins. On the other hand, this review attempts to illustrate, for mathematicians and bioinformaticians, the potential that represent these molecular and cellular events for modeling clear physiological processes.

13-P010 Auxin in the control of the development of the brown alga Ectocarpus siliculosus

cate that Shh signalling controls Laminin gene expression, a critical step in the formation of the myotomal basement membrane, and that Laminin-111 has specific and essential role in myotomal basement membrane assembly. Together, our data reveal a novel role for Shh signalling in the control of basement membrane assembly and provide a framework for understanding the complex interactions between extra-cellular matrix and intercellular signalling.