Metabolic Engineering of Yeast and Plants for the Production of the Biologically Active Hydroxystilbene, Resveratrol

Resveratrol, a stilbenic compound deriving from the phenyalanine/polymalonate route, being stilbene synthase the last and key enzyme of this pathway, recently has become the focus of a number of studies in medicine and plant physiology. Increased demand for this molecule for nutraceutical, cosmetic and possibly pharmaceutic uses, makes its production a necessity. In this context, the use of biotechnology through recombinant microorganisms and plants is particularly promising. Interesting results can indeed arise from the potential of genetically modified microorganisms as an alternative mechanism for producing resveratrol. Strategies used to tailoring yeast as they do not possess the genes that encode for the resveratrol pathway, will be described. On the other hand, most interest has centered in recent years, on STS gene transfer experiments from various origins to the genome of numerous plants. This work also presents a comprehensive review on plant molecular engineering with the STS gene, resulting in disease resistance against microorganisms and the enhancement of the antioxidant activities of several fruits in transgenic lines

Mineral content and biomechanical properties of fibrolamellar bone

editorial reviewe

A Molecular-Scale Understanding of Misorientation Toughening in Corals and Seashells.

peer reviewedBiominerals are organic-mineral composites formed by living organisms. They are the hardest and toughest tissues in those organisms, are often polycrystalline, and their mesostructure (which includes nano- and microscale crystallite size, shape, arrangement, and orientation) can vary dramatically. Marine biominerals may be aragonite, vaterite, or calcite, all calcium carbonate (CaCO3 ) polymorphs, differing in crystal structure. Unexpectedly, diverse CaCO3 biominerals such as coral skeletons and nacre share a similar characteristic: Adjacent crystals are slightly misoriented. This observation is documented quantitatively at the micro- and nanoscales, using polarization-dependent imaging contrast mapping (PIC mapping), and the slight misorientations are consistently between 1° and 40°. Nanoindentation shows that both polycrystalline biominerals and abiotic synthetic spherulites are tougher than single-crystalline geologic aragonite. Molecular dynamics (MD) simulations of bicrystals at the molecular scale reveal that aragonite, vaterite, and calcite exhibit toughness maxima when the bicrystals are misoriented by 10°, 20°, and 30°, respectively, demonstrating that slight misorientation alone can increase fracture toughness. Slight-misorientation-toughening can be harnessed for synthesis of bioinspired materials that only require one material, are not limited to specific top-down architecture, and are easily achieved by self-assembly of organic molecules (e.g., aspirin, chocolate), polymers, metals, and ceramics well beyond biominerals

Le harpon biologique de la mante marine (lysiosquillina maculata): comparaison structurale et biomécanique avec les cuticules du corps et d'autres épines

Spearing mantis shrimps are aggressive crustaceans impaling fish at high speed thanks to a spiky raptorial appendage. The multiple spikes of this appendage must stand strong and repetitive mechanical constraints during the impact with the fish skin and the prey retention. To face those constraints the mantis shrimp spikes must have deeply adapt their exoskeleton (or cuticle) to tune their mechanical properties. To understand how millions of years of evolution has forged this formidable harpoon multiple approaches will be combined in this thesis. In a first time, the internal organization and composition of the cuticles of the spike and the body of the mantis shrimp were finely described thanks to microscopy methods to determine the basic and derived state of the cuticle ultrastructure and composition. Secondly, cytochemical treatments were used on moulting time-series samples to elucidate the matrix properties of the mantis shrimp cuticle as well as its dynamical ultrastructural changes. Then, a multiscale study was conducted on the raptorial spike to observe its adaptation at macro-(shape), micro- and composition-scales and link them with mechanical properties. Finally, a larger vision on cuticular adaptations of the mantis shrimp cuticle was obtained thanks to an ultrastructural and compositional study of three other spike-like cuticular structures found on the animal body (maxilliped dactyl, propodus spike and uropod spike). This multidisciplinar study of this specific biological material answers diverse questions concerning biological as well as mechanical thematic. The two first approaches show unique cuticular features, gave a comparison of the mantis shrimp cuticle with the other crustacean in the light of the group phylogeny and finally elucidate the specific secretive features giving rise to this specialized structure. The third approach highlights the keys structural and compositional adaptations of the spike cuticle and their impact on the mechanical properties, giving insight into the production of bioinspired tough materials. Finally, the last approach of this work broadened our understanding of the evolutive history of the mantis shrimp cuticle bringing to light other structural and compositional features interesting in biological material sciences.Les mantes marines ravisseuses (ou squilles) sont d’agressifs crustacés qui empalent les poissons à grande vitesse grâce à un appendice de prédation muni d’épines. Les multiples épines de cet appendice doivent supporter de fortes contraintes mécaniques répétitives lors de l'impact avec la peau du poisson et durant la rétention de la proie. Pour faire face à ces contraintes, les épines de la crevette mante doivent avoir profondément adapté leur exosquelette (ou cuticule) pour ajuster leurs propriétés mécaniques. Pour comprendre comment des millions d'années d'évolution ont forgé ces formidables harpons, plusieurs approches seront combinées dans cette thèse. Dans un premier temps, l'organisation interne et la composition des cuticules de l'épine et du corps de la crevette mante ont été finement décrites grâce à des méthodes de microscopie pour déterminer l'état initial et dérivé de l'ultrastructure et de la composition de la cuticule. Deuxièmement, des traitements cytochimiques ont été utilisés sur des échantillons de séries chronologiques de mue pour élucider les propriétés de la matrice de la cuticule de la crevette mante ainsi que ses changements ultrastructuraux dynamiques durant la mue. Ensuite, une étude à multiples échelles a été menée sur l'épine pour observer son adaptation aux échelles macrométrique (forme), micrométrique et composition les reliant ensuite aux propriétés mécaniques. Enfin, une vue plus large sur les adaptations cuticulaires de la cuticule de la crevette mante a été obtenue grâce à une étude ultrastructurale et compositionnelle de trois autres épines cuticulaires trouvées sur le corps animal (maxillipède du dactyle, épine du propodite et épine de l’uropode). Cette étude multidisciplinaire de ce matériel biologique spécialisé répond à diverses questions concernant des thématiques aussi bien biologiques que mécaniques. Les deux premières approches montrent les caractéristiques structurelles uniques, comparent la cuticule de la crevette mante avec celles des autres crustacé tenant compte de la phylogénie du groupe et enfin élucident les caractéristiques de sécrétion spécifiques à l'origine de cette structure spécialisée. Enfin, la troisième approche met en évidence les principales adaptations structurelles et compositionnelles de la cuticule de l’épine et leur impact sur les propriétés mécaniques, pointant des adaptations ayant un potentiel intérêt dans la production de matériaux résistants bio-inspirés. Enfin, la dernière approche de ce travail a élargi notre compréhension de l'histoire évolutive de la cuticule de la crevette mante en mettant en lumière d'autres caractéristiques structurelles et compositionnelles intéressantes pour les sciences des matériaux biologiques

étude d'un harpon biologique; la microstructure de l'appendice ravisseur de la mante marine

Mantis shrimps (or Stomatopods) form one of the most surprising crustacean order in earth. Separated from decapods since Cretacean, species composing this group display unique features that makes them formidable marine benthic predators. In these features can be found the widest visual spectrum, a transformed telson used both as a shield and as a fin, an antennal pallet used as a rudder but also a pair of enhanced predatorial limbs. Two kinds of predatorial limbs exists, dividing mantis shrimps in two groups: smashing limbs used to brake carapaces and to knock out preys and spearing limbs used to impale fishes. Both these limbs are deployed at high speed thanks to their ability to store and release elastic energy and are reinforced to endure impacts. This study will focus on one spearing mantis shrimp, the striped mantis shrimp (Lysiosquillina maculata (Fabritius,1973)) and the cuticle of its raptorial appendage. The main goal is to understand how spines found in this appendage are internally arranged to face the mechanical stress that occurs when harpooning prey. Techniques as optical microscopy and scanning electron microscopy will allow to define how is arranged the cuticle found in the spine. Techniques of micro-analysis will then superimpose composition to the structural information. These analyses highlight a complex assembly of four layers which can’t be directly linked to the classic succession of layers found in arthropods (e.g. endocuticle, exocuticle and epicuticle). These layers were named lamellar layer, parallel layer, soft helicoidal layer and highly mineralised layer. Each of them differs by fibres orientations, mineralisation rate or compositions and they are thought to play precise roles in the mechanical behaviour of the stomatopod spine

Renforcement et adaptation de l'épine de la mante marine: comment la cuticule d'un crustacé devient un parfait harpon?

Introduction: In the field of bioinspired material, the crustacean cuticle is seen as an example of organo-mineral biomaterial able to endure strong stress thanks to the combination of fibrilar organisation and mineral deposition. Stomatopoda is a crustacean order represented by two groups of species; smashing mantis shrimps and spearing mantis shrimps. Hence, the first group is already well studied for the mechanical abilities of their smashing limbs, this study will focus on the spearing mantis shrimps and their spearing appendage. These appendage present spikes able to impale fish in a fraction of second and are therefore designed to penetrate at high speed, to avoid escape of the prey but also to resist bending during the capture. Objective: The aim of this study is to determine how the mantis shrimp cuticle adapts its shape, internal organization and composition to endure the intense stress occurring during attacks. Materials and methods: Specimens of Lysiosquillina maculata were dissected and their spikes were conserved in ethanol or fixed with glutaraldehyde. The samples in ethanol were then embedded in resin and polished to be observed in µCTscan and SEM and analyzed by EDS and nanoindentation. Sample fixed in glutaraldehyde were contrast with OsO4 and embedded in resin to be observed in TEM. Results: Firstly, the µCTscan highlighted particular features in the external structure, the spike presenting serrations at its edges linked by grooves at its sides. It also show a curvature linked to the angle of attack of the mantis shrimp. But its internal organization also shows important rearrangements in comparison to the classic crustacean cuticle. These rearrangements affect even the arrangement of chitino-proteic fibers as the composition of the mineral. SEM-BSE and TEM observation highlight that the spike cuticle does not show a clear subdivision in the three classical layer found in arthropod cuticle i.e. endo-, exo, and epicuticle but consist in four layer with different organization. These layers are respectively called from the surface to the epidermis: the highly mineralized layer (HML), the outer helicoidal layer (OHL), the striated layer (STL) and the inner helicoidal layer (IHL). EDS analysis also shows that theses layers also present different composition and mineralization rate; the HML has the higher mineralization rate and is composed of flurapatite, the HML has the lower mineralization rate and is, as the OHL and STL, composed of a mix between calcium phosphate and calcium carbonate. The three innermost layers are also characterized by variation in the substitution rates of minor element as Na, K, Mg and F. Analysis of the spike cuticle with nanoindentation tests highlight variation in the reduced modulus between the layers. Conclusion: During its evolution, the spearing mantis shrimp has strongly modified its exoskeleton either in structure and mineral composition in order to perfectly suit its mechanical constraint. Variation of its internal organization seems to be adapted to endure with anisotropic stresses. On its side, modification of the composition in the mineral part raised its surface reduced modulus and hardness at value comparable with vertebrate teeth. Finally, by both structural and mineral variation, the cuticle spike is also thought to cope with cracks propagations.Architecture et tests mécaniques d'une structure organo-minérale, la cuticule de mante marine : comparaison par impressions 3D biomimétique

The spiky exoskeleton of the spearing mantis shrimp; comparative study of 3 puncturing bio-tools.

From the defensives cacti spines to the offensive felid claws and harpooning honeybee stinger, puncturing tools are found in a wide diversity of living organisms. They fulfil various roles and are based on different biological materials. Among those materials, the arthropod cuticle is often pointed out as the most versatile one. It is made of a complex helicoidal arrangement of chitin-protein fibres embedded into an organo-mineral matrix giving rise to an excellent multifunctional material serving for the organism protection and support and for all other secondary tasks as walking, sensing, or hunting. Among arthropods, impressive examples of puncturing tools can be found in the spearing mantis shrimp, an aggressive hunting crustacean known for its fast spiky appendage designed to impale prey within a fraction of second. This animal has the peculiarity to possess three puncturing tools with different roles but derived from the same basic material. Indeed, in addition to the spearing spikes used for hunting, mantis shrimps also possess defensive back spikes (the uropod spikes) to protect against predators, and maxilliped spikes for food treatment. Each of these tools must cope with specific mechanical challenges and therefore shows distinct structural and compositional adaptations. Following a multiscale structural and compositional analysis, we highlight three key adaptive strategies. All three spikes have the endocuticle subdivided into 3 functional regions. Nevertheless, the ultrastructure and the composition of the superior cuticular layers show divergent features, ranging from highly mineralized exocuticle and reduced epicuticle in the raptorial spike, to an organic and prismatic exocuticle surrounded by a bromine enriched epicuticle in the maxilliped spike and an intermediate structure in the uropod spike. This study gave a unique comparison of three different puncturing tools derived from the same structure, highlighting the incredible versatility of the arthropod cuticle

Microstructural and compositional variation in pacu and piranha teeth related to diet specialization (Teleostei: Serrasalmidae)

In any vertebrate group, tooth shape is known to fit with a biological function related to diet. However, little is known about the relationships between diet and tooth microstructure and composition in teleost fishes. In this work, we describe the external morphology, internal microstructure and elemental composition of the oral teeth of three representative species of the family Serrasalmidae having different feeding habits (herbivorous vs. omnivorous vs. carnivorous). We used backscattered-electron imaging and low vacuum environmental scanning electron microscope to compare the organization and mineralization of tooth layers as well as energy dispersive X-ray microanalysis and Raman microspectrometry to investigate the elemental composition, Ca/P ratio and mineralogy of the most superficial layers. Oral teeth of each serrasalmid species have the same internal orga- nization based on five distinctive layers (i.e. pulp, dentine, inner enameloid, outer enameloid and cuticle) but the general tooth morphology is different according to diet. Microstructural and compositional variation of the cuticle and iron-enrichment of superficial layers were highlighted between herbivorous and carnivorous species. Iron is more concentrated in teeth of the herbivorous species where it is associated with a thicker cuticle ex- plaining the more intense red-pigmentation of the cutting edges of oral teeth. The iron-enrichment is interpreted as a substitution of Ca by Fe in the hydroxyapatite. These traits are discussed in the light of the evolutionary history of the family. Further considerations and hypotheses about the formation and origin of the mineralized tooth layers and especially the iron-rich superficial layers in teleost fishes are suggested

Structure and mineralization of the spearing mantis shrimp (Stomatopoda; Lysiosquillina maculata) body and spike cuticles

Stomatopoda is a crustacean order including sophisticated predators called spearing and smashing mantis shrimps that are separated from the well-studied Eumalacotraca since the Devonian. The spearing mantis shrimp has developed a spiky dactyl capable of impaling fishes or crustaceans in a fraction of second. In this high velocity hunting technique, the spikes undergo an intense mechanical constraint to which their exoskeleton (or cuticle) has to be adapted. To better understand the spike cuticle internal architecture and composition, electron microscopy, X-ray microanalysis and Raman spectroscopy were used on the spikes of 7 individuals (collected in French Polynesia and Indonesia), but also on parts of the body cuticle that have less mechanical stress to bear. In the body cuticle, several specificities linked to the group were found, allowing to determine the basic structure from which the spike cuticle has evolved. Results also highlighted that the body cuticle of mantis shrimps could be a model close to the ancestral arthropod cuticle by the aspect of its biological layers (epi- and procuticle including exo- and endocuticle) as well as by the Ca-carbonate/phosphate mineral content of these layers. In contrast, the spike cuticle exhibits a deeply modified organization in four functional regions overprinted on the biological layers. Each of them has specific fibre arrangement or mineral content (fluorapatite, ACP or phosphate-rich Ca-carbonate) and is thought to assume specific mechanical roles, conferring appropriate properties on the entire spike. These results agree with an evolution of smashing mantis shrimps from primitive stabbing/spearing shrimps, and thus also allowed a better understanding of the structural modifications described in previous studies on the dactyl club of smashing mantis shrimps.Architecture and mechanical testing of resistant organo-mineral structures of the mantis shrimp cuticle: comparison with biomimetic 3D imprint