New insights into carbon transport and incorporation to wood

Wood formation in trees requires carbon import from the photosynthetic tissues. In several tree species the majority of this carbon is derived from sucrose transported in the phloem. This thesis describes experimental work on the mechanism of radial sucrose transport from phloem to developing wood and subsequent incorporation of carbon into wood polymers. I investigated the role of active sucrose transport during secondary cell wall formation in hybrid aspen (Populus tremula x tremuloides). Reduction of a plasma membrane localised sucrose transporter (PttSUT3) decreased carbon allocation to secondary walls of wood fibers. The results show the importance of active sucrose transport for wood formation in a symplasmic phloem loading tree species, and identify PttSUT3 as a principal transporter for carbon delivery into secondary cell wall forming wood fibers. To investigate the temporal dynamics of carbon transport and wood polymer biosynthesis, I labelled two-month-old hybrid aspen trees with ¹³CO₂ and tracked the flux of ¹³C from leaves to developing wood. Analysis of the cell wall polymer labelling patterns using 2D-NMR revealed temporal differences in the labelling of carbohydrates and lignin subunits. Further analysis showed that ¹³C incorporation into different wood polymers is influenced by the diurnal cycle. Primary metabolism enzymes in the developing wood play an important role in carbon partitioning to wood cell wall polymers. In this part of the study, the activity of eight primary metabolism enzymes linking sucrose to cell wall precursor biosynthesis was determined in phloem, cambium and in different stages of wood development. Comparison of enzyme activity measurements with transcript and metabolite profiles across the developing wood suggested a central role for transcriptional regulation of carbon allocation to wood. Combined, the results of the three projects provide new insights into the mechanism and regulation of carbon allocation to developing wood

Cauliflower mosaic virus protein P6 is a multivalent node for RNA granule proteins and interferes with stress granule responses during plant infection

Cauliflower mosaic virus intersects with components of the stress granule pathway within its replication factories and has a global capacity to suppress stress granule assembly in plants.Biomolecular condensation is a multipurpose cellular process that viruses use ubiquitously during their multiplication. Cauliflower mosaic virus replication complexes are condensates that differ from those of most viruses, as they are nonmembranous assemblies that consist of RNA and protein, mainly the viral protein P6. Although these viral factories (VFs) were described half a century ago, with many observations that followed since, functional details of the condensation process and the properties and relevance of VFs have remained enigmatic. Here, we studied these issues in Arabidopsis thaliana and Nicotiana benthamiana. We observed a large dynamic mobility range of host proteins within VFs, while the viral matrix protein P6 is immobile, as it represents the central node of these condensates. We identified the stress granule (SG) nucleating factors G3BP7 and UBP1 family members as components of VFs. Similarly, as SG components localize to VFs during infection, ectopic P6 localizes to SGs and reduces their assembly after stress. Intriguingly, it appears that soluble rather than condensed P6 suppresses SG formation and mediates other essential P6 functions, suggesting that the increased condensation over the infection time-course may accompany a progressive shift in selected P6 functions. Together, this study highlights VFs as dynamic condensates and P6 as a complex modulator of SG responses

Arabidopsis RNA processing body components LSM1 and DCP5 aid in the evasion of translational repression during Cauliflower mosaic virus infection

Viral infections impose extraordinary RNA stress, triggering cellular RNA surveillance pathways such as RNA decapping, nonsense-mediated decay, and RNA silencing. Viruses need to maneuver among these pathways to establish infection and succeed in producing high amounts of viral proteins. Processing bodies (PBs) are integral to RNA triage in eukaryotic cells, with several distinct RNA quality control pathways converging for selective RNA regulation. In this study, we investigated the role of Arabidopsis thaliana PBs during Cauliflower mosaic virus (CaMV) infection. We found that several PB components are co-opted into viral factories that support virus multiplication. This pro-viral role was not associated with RNA decay pathways but instead, we established that PB components are helpers in viral RNA translation. While CaMV is normally resilient to RNA silencing, dysfunctions in PB components expose the virus to this pathway, which is similar to previous observations for transgenes. Transgenes, however, undergo RNA quality control-dependent RNA degradation and transcriptional silencing, whereas CaMV RNA remains stable but becomes translationally repressed through decreased ribosome association, revealing a unique dependence among PBs, RNA silencing, and translational repression. Together, our study shows that PB components are co-opted by the virus to maintain efficient translation, a mechanism not associated with canonical PB functions.Arabidopsis RNA processing body components LSM1 and DCP5 are co-opted by Cauliflower mosaic virus to maintain efficient virus translation in the presence of RNA DEPENDENT POLYMERASE6-governed silencing

Evaluation of Nutritional Composition of Pure Filamentous Fungal Biomass as a Novel Ingredient for Fish Feed

The rapid growth of aquaculture and the lack of fish meal demand new sustainable ingredients. Although fungal biomass is found to be a promising sustainable fish feed supplementation candidate, the characteristics of this protein-rich source are closely influenced by the quality of the applied growth medium. In this work, the nutritional properties of pure filamentous fungal biomass provided from the cultivation of Aspergillus oryzae, Neurospora intermedia and Rhzopus oryzae were evaluated to assess their potential as alternative novel protein sources in fish feed. In this regard, fungal biomass yields of up to 0.19 ± 0.005 (g dry biomass/g substrate glucose) were obtained during submerged cultivation of fungal strains. The pure fungal biomass acquired could contain significant amounts of protein up to 62.2 ± 1.2% (w/w). The obtained protein had a high quality with notable inclusion of essential amino acids such as lysine, arginine, methionine and threonine with comparable concentrations to those of fish meal. Fungal biomass is mainly considered as protein source, however, entitlement of 6.9 ± 0.5, 4.0 ± 0.7 and 17.2 ± 1.1% (w/w) of lipids and ratio of polyunsaturated fatty acids (PUFA) to saturated fatty acids (SFA) of 1.37:1, 1.74:1 and 1.47:1 in A. oryzae, N. intermedia and R. oryzae, respectively, signal health benefits for the fish. Considering the results, protein-rich pure fungal biomass with amino acid composition is greatly compatible with fish meal, and contains essential nutrients such as fatty acids and minerals. This pure biomass constitutes a promising sustainable alternative supplement to be introduced in fish feed industry

Distributed Deployment Strategies for Improved Coverage in a Network of Mobile Sensors With Prioritized Sensing Field

Efficient deployment strategies are proposed for a mobile sensor network, where the coverage priority of different points in the field is specified by a given function. The multiplicatively weighted Voronoi (MW-Voronoi) diagram is utilized to find the coverage holes of the network for the case where the sensing ranges of different sensors are not the same. Under the proposed strategies, each sensor detects coverage holes within its MW-Voronoi region, and then moves in a proper direction to reduce their size. Since the coverage priority of the field is not uniform, the target location of each sensor is determined based on the weights of the vertices or the points inside the corresponding MW-Voronoi region. Simulations validate the theoretical results

Immersed flat-sheet membrane bioreactors for lignocellulosic bioethanol production

The rising awareness of the environmental, economic and socio-political impacts of over-exploitation of fossil-based fuel and energy sources, have motivated the transition toward more sustainable and renewable energy sources. Lignocellulosic materials (e.g. agricultural residues) are potential candidates for sustainable bioethanol production that contributes to the replacement of fossil fuels. However, to have an economically feasible and commercialized process, issues associated with lignocellulosic bioethanol production in upstream, fermentation and downstream processing stages should be alleviated. Membrane bioreactors with their great capabilities in semi-selective separation are promising options for making a breakthrough in lignocellulosic biorefinery processes. Therefore, in this thesis, different membrane modules and immersed membrane bioreactors (iMBRs) set-ups were developed and applied to take advantage of this long-matured water and wastewater treatment technique in remediation of challenges in the lignocellulosic bioethanol production. Thus, In order to intensify and optimize the lignocellulosic bioethanol production process, pressure-driven flat sheet microfiltration iMBRs were integrated into different processing stages. The application of a continuous iMBR led to a high ethanol productivity and yield (83% of theoretical yield) at high suspended solid content (up to 20% w/v) of wheat straw hydrolysate, and successful bacterial contamination separation from yeast (up to 93% removal). Moreover, using double-staged iMBRs for continuous hydrolysis-filtration and co-fermentation-filtration led to an effective separation of lignin-rich solids (up to 70% lignin) and sugar streams from the hydrolysate, and yeast cells from the fermentation product stream, stable long-term filtration performance (up to 264 h) at filtration flux of 21.9 l.m-2.h-1. In this thesis, filtration performance was thoroughly investigated, and effective physical fouling preventive approaches were applied to guarantee continuous bioprocessing. In addition, in order to remediate issues related to high content of inhibitors and presence of sequentially-fermented hexose and pentose saccharides in lignocellulosic fermentation, the cell-confinement approach of reverse membrane bioreactor (rMBR), which merges the benefits of iMBRs and cell encapsulation techniques, was introduced and applied in this thesis. It was observed that the high local cell density and diffusion-based mass transfer in the rMBR promoted co-utilization of sugars, and boosted cell furfural detoxification at concentrations of up to 16 g.l-1. Moreover, considering the needs of rMBR processes for cell recirculation, membrane envelope degassing, and media conditioning, a novel membrane module was designed, developed, and patented in this thesis work

Removal of Bacterial Contamination from Bioethanol Fermentation System Using Membrane Bioreactor

A major issue hindering efficient industrial ethanol fermentation from sugar-based feedstock is excessive unwanted bacterial contamination. In industrial scale fermentation, reaching complete sterility is costly, laborious, and difficult to sustain in long-term operation. A physical selective separation of a co-culture of Saccharomyces cerevisiae and an Enterobacter cloacae complex from a buffer solution and fermentation media at dilution rates of 0.1–1 1/h were examined using an immersed membrane bioreactor (iMBR). The effect of the presence of yeast, inoculum size, membrane pore size, and surface area, backwashing and dilution rate on bacteria removal were assessed by evaluating changes in the filtration conditions, medium turbidity, and concentration of compounds and cell biomass. The results showed that using the iMBR with dilution rate of 0.5 1/h results in successful removal of 93% of contaminating bacteria in the single culture and nearly complete bacteria decontamination in yeast-bacteria co-culture. During continuous fermentation, application of lower permeate fluxes provided a stable filtration of the mixed culture with enhanced bacteria washout. This physical selective separation of bacteria from yeast can enhance final ethanol quality and yields, process profitability, yeast metabolic activity, and decrease downstream processing costs