Mário Rui P. F. N. Costa

Pollen grain development is compromised in Arabidopsis agp6 agp11 null mutants

Arabinogalactan proteins (AGPs) are structurally complex plasma membrane and cell wall proteoglycans that are implicated in diverse developmental processes, including plant sexual reproduction. Male gametogenesis (pollen grain development) is fundamental to plant sexual reproduction. The role of two abundant, pollen-specific AGPs, AGP6, and AGP11, have been investigated here. The pollen specificity of these proteoglycans suggested that they are integral to pollen biogenesis and their strong sequence homology indicated a potential for overlapping function. Indeed, single gene transposon insertion knockouts for both AGPs showed no discernible phenotype. However, in plants homozygous for one of the insertions and heterozygous for the other, in homozygous double mutants, and in RNAi and amiRNA transgenic plants that were down-regulated for both genes, many pollen grains failed to develop normally, leading to their collapse. The microscopic observations of these aborted pollen grains showed a condensed cytoplasm, membrane blebbing and the presence of small lytic vacuoles. Later in development, the generative cells that arise from mitotic divisions were not seen to go into the second mitosis. Anther wall development, the establishment of the endothecium thickenings, the opening of the stomium, and the deposition of the pollen coat were all normal in the knockout and knockdown lines. Our data provide strong evidence that these two proteoglycans have overlapping and important functions in gametophytic pollen grain development.

Early germination of Arabidopsis pollen in a double null mutant for the arabinogalactan protein genes AGP6 and AGP11.

The pollen specificity of the Arabidopsis arabinogalactan protein (AGP) genes AGP6 and AGP11 suggests that they are integral to pollen biogenesis, and their high percent of sequence similarity may indicate a potential for overlapping function. Arabidopsis agp6 agp11 double null mutants have been studied in our laboratory, and in the present work, we characterize the germination and growth of its pollen. When compared to wild type, mutant agp6 agp11 pollen displayed reduced germination and elongation, both in vivo and in vitro, and precocious germination inside the anthers, provided that sufficient moisture was available. This characteristic was not observed in wild type plants, even in water content conditions which for the mutant were sufficient for pollen germination. Therefore, an additional distinctive phenotypic trait of arabinogalactan proteins AGP6 and AGP11 may be to avert untimely germination of pollen. Such AGPs may control germination through water uptake, suggesting an important biological function of this gene family in pollen.

Expression-based and co-localization detection of arabinogalactan protein 6 and arabinogalactan protein 11 interactors in Arabidopsis pollen and pollen tubes

BackgroundArabinogalactan proteins (AGPs) are cell wall proteoglycans that have been shown to be important for pollen development. An Arabidopsis double null mutant for two pollen-specific AGPs (agp6 agp11) showed reduced pollen tube growth and compromised response to germination cues in vivo. A microarray experiment was performed on agp6 agp11 pollen tubes to search for genetic interactions in the context of pollen tube growth. A yeast two-hybrid experiment for AGP6 and AGP11 was also designed.ResultsThe lack of two specific AGPs induced a meaningful shift in the gene expression profile. In fact, a high number of genes showed altered expression levels, strengthening the case that AGP6 and AGP11 are involved in complex phenomena. The expression levels of calcium- and signaling-related genes were found to be altered, supporting the known roles of the respective proteins in pollen tube growth. Although the precise nature of the proposed interactions needs further investigation, the putative involvement of AGPs in signaling cascades through calmodulin and protein degradation via ubiquitin was indicated. The expression of stress-, as well as signaling- related, genes was also changed; a correlation that may result from the recognized similarities between signaling pathways in both defense and pollen tube growth.The results of yeast two-hybrid experiments lent further support to these signaling pathways and revealed putative AGP6 and AGP11 interactors implicated in recycling of cell membrane components via endocytosis, through clathrin-mediated endosomes and multivesicular bodies.ConclusionsThe data presented suggest the involvement of AGP6 and AGP11 in multiple signaling pathways, in particular those involved in developmental processes such as endocytosis-mediated plasma membrane remodeling during Arabidopsis pollen development. This highlights the importance of endosomal trafficking pathways which are rapidly emerging as fundamental regulators of the wall physiology.

Operational and Design Study of RIM Machines

Abstract Particle Image Velocimetry (PIV) and Laser Doppler Anemometry (LDA) data is presented and used to characterize the dynamic behaviour of the flow field in a RIM machine, and the main flow patterns and mixing mechanisms are examined and established. A two-dimensional CFD model is used for the simulation of mass transfer and chemical reaction in the mixing chamber of a RIM machine. Some design and operational parameters are considered and its influence on the mixing behaviour of a RIM machine is analysed.

A general kinetic analysis of non-linear irreversible copolymerizations

Abstract Irreversible non-linear copolymerizations with arbitrarily great numbers of monomers are described specifying the stoichiometric coefficients of the reactions involving end-groups. Rate equations for the vectorial generating function or discrete transform of the distribution of end-groups and repeating units and its derivatives with respect to logarithms of Laplace parameters can thus be written. Similar rate equations can be written for the discrete transforms of the distributions of descendants of each linking group with respect to their numbers of end-groups. This allows the prediction of gel properties, such as the concentration of elastically active network chains and mass fraction of pendant material. Mass balance equations using these rate laws are non-linear partial differential equations solvable by the method of characteristics. Numerical methods are developed for computing molecular weight distributions and average molecular weights before and after gelation, as well as the weight fraction of sol ang gel properties related to its elasticity behaviour. The curing of a tetraepoxide with a primary diamine is taken as a case study and results are compared with predictions by the theory of branching processes.

A global model for the hot-pressing of MDF

The hot-pressing operation is the final stage in medium-density fiberboard (MDF) manufacture, where the mattress of fibers is compressed and heated to promote the cure of the resin. In MDF hot-pressing, many physical, chemical and mechanical processes are involved; the complexity of this operation arises from the fact that they are coupled. A global model is presented for this operation that integrates all mechanisms involved in the panel formation (heat and mass transfer, chemical reaction and mechanical behavior). This approach results in a two-dimensional unsteady state problem, which involves the knowledge of the polymerization kinetics of the resin, the transport properties and material properties, which are position and time dependent. This dynamic model was used to predict the evolution of the variables relating to heat and mass transfer (temperature, moisture content, gas pressure and relative humidity), as well as the variables relating to mechanical behavior (pressing pressure, strain, modulus of elasticity and density). The model performance was analyzed using the typical operating conditions for the hot-pressing of MDF and the results were compared to the experimental data from an industrial MDF press. We concluded that the model could predict in an acceptable way the behavior of the key variables for the control of the pressing cycle, as well as some physico–mechanical properties of the final product. The improvement of this model will permit the scheduling of the press cycle to fulfill objectives of minimization of energy consumption, better quality of the board and increased process flexibility.

Immunolocalization of arabinogalactan proteins (AGPs) in reproductive structures of an early-divergent angiosperm, Trithuria (Hydatellaceae)

BACKGROUND AND AIMS Trithuria is the sole genus of Hydatellaceae, a family of the early-divergent angiosperm lineage Nymphaeales (water-lilies). In this study different arabinogalactan protein (AGP) epitopes in T. submersa were evaluated in order to understand the diversity of these proteins and their functions in flowering plants. METHODS Immunolabelling of different AGPs and pectin epitopes in reproductive structures of T. submersa at the stage of early seed development was achieved by immunofluorescence of specific antibodies. KEY RESULTS AGPs in Trithuria pistil tissues could be important as structural proteins and also as possible signalling molecules. Intense labelling was obtained with anti-AGP antibodies both in the anthers and in the intine wall, the latter associated with pollen tube emergence. CONCLUSIONS AGPs could play a significant role in Trithuria reproduction, due to their specific presence in the pollen tube pathway. The results agree with labellings obtained for Arabidopsis and confirms the importance of AGPs in angiosperm reproductive structures as essential structural components and probably important signalling molecules.

Characterization of urea-formaldehyde resins by GPC/SEC and HPLC techniques: Effect of ageing

During the last 40 years, several analytical techniques have been developed/adapted to characterize urea-formaldehyde (UF) resins. However, a great part of the research about this kind of wood adhesives has been performed by industrial producers and, thus, the main part of the existing knowledge is retained within those companies. This work describes a methodology for determining the molecular weight distribution (MWD) of UF resins using Gel Permeation Chromatography (GPC)/Size Exclusion Chromatography (SEC) with 2 detectors (differential refractive index (RI) and differential viscosity). This method permitted to characterize/distinguish commercial UF resins produced with different F/U molar ratios and to monitor the molecular weight and MWD with ageing. An HPLC method was additionally used to evaluate the fraction of unreacted urea, monomethylolurea and dimethylolurea present in commercial UF resins and measure the evolution of these three compounds with ageing.

Optimization of the Synthesis of Urea-Formaldehyde Resins using Response Surface Methodology

In the near future, companies will face the need to produce low formaldehyde emission resins, i.e., not above the emission level of natural wood. However, for producing this new generation of urea-formaldehyde resins (UF), it is necessary to optimize the synthesis process. This work describes an optimization procedure for UF resin synthesis, following an alkaline–acid process, focusing on the conditions of the condensation step. A design of experiments methodology was employed to optimize the 3 selected factors (number of urea additions, time span between urea additions, and condensation pH), in order to produce particleboards with maximum internal bond strength and minimum formaldehyde release. The condensation pH played a significant role in increasing the Internal Bond (IB) strength and reducing the Formaldehyde Emission (FE). The sequential addition of urea also has a noticeable influence on resin performance. Optimum conditions for production of UF resins have been proposed and tested by the response surface methodology using the desirability function.

Synthesis of waterborne polyurethane-urea dispersions with chain extension step in homogeneous and heterogeneous media

HYPOTHESIS The possibility of tailoring the final properties of environmentally friendly waterborne polyurethane and polyurethane-urea dispersions and the films they produce makes them attractive for a wide range of applications. Both the reagents content and the synthesis route contribute to the observed final properties. EXPERIMENTS A series of polyurethane-urea and polyurethane aqueous dispersions were synthesized using 1,2-ethanediamine and/or 1,4-butanediol as chain extenders. The diamine content was varied from 0 to 4.5wt%. Its addition was carried out either by the classical heterogeneous reaction medium (after phase inversion step), or else by the alternative homogeneous medium (prior to dispersion formation). Dispersions as well as films prepared from dispersions have been later extensively characterized. FINDINGS 1,2-Ethanediamine addition in heterogeneous medium leads to dispersions with high particle sizes and broad distributions whereas in homogeneous medium, lower particle sizes and narrow distributions were observed, thus leading to higher uniformity and cohesiveness among particles during film formation. Thereby, stress transfer is favored adding the diamine in a homogeneous medium; and thus the obtained films presented quite higher stress and modulus values. Furthermore, the higher uniformity of films tends to hinder water molecules transport through the film, resulting, in general, in a lower water absorption capacity.