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4. Ectopic callose deposition into woody biomass modulates the nano-architecture of macrofibrils

Author

Bourdon, Matthieu, Lyczakowski, Jan J., Cresswell, Rosalie, Amsbury, Sam, Vilaplana, Francisco, Le Guen, Marie-Joo, Follain, Nadège, Wightman, Raymond, Su, Chang, Alatorre-Cobos, Fulgencio, Ritter, Maximilian, Liszka, Aleksandra, Terrett, Oliver M., Yadav, Shri Ram, Vatén, Anne, Nieminen, Kaisa, Eswaran, Gugan, Alonso-Serra, Juan, Müller, Karin H., Iuga, Dinu, Miskolczi, Pal Csaba, Kalmbach, Lothar, Otero, Sofia, Mähönen, Ari Pekka, Bhalerao, Rishikesh, Bulone, Vincent, Mansfield, Shawn D., Hill, Stefan, Burgert, Ingo, Beaugrand, Johnny, Benitez-Alfonso, Yoselin, Dupree, Ray, Dupree, Paul, and Helariutta, Ykä

Published
2023
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7. A biophysical model for plant cell plate maturation based on the contribution of a spreading force.

Author

Jawaid, Muhammad Zaki, Sinclair, Rosalie, Bulone, Vincent, Cox, Daniel L, and Drakakaki, Georgia

Subjects
Cytoplasm, Glucans, Biophysics, Models, Biological, Plant Physiological Phenomena, Plant Cells, Biological Sciences, Agricultural and Veterinary Sciences, Plant Biology & Botany
Abstract

Plant cytokinesis, a fundamental process of plant life, involves de novo formation of a "cell plate" partitioning the cytoplasm of dividing cells. Cell plate formation is directed by orchestrated delivery, fusion of cytokinetic vesicles, and membrane maturation to form a nascent cell wall by timely deposition of polysaccharides. During cell plate maturation, the fragile membrane network transitions to a fenestrated sheet and finally a young cell wall. Here, we approximated cell plate sub-structures with testable shapes and adopted the Helfrich-free energy model for membranes, including a stabilizing and spreading force, to understand the transition from a vesicular network to a fenestrated sheet and mature cell plate. Regular cell plate development in the model was possible, with suitable bending modulus, for a two-dimensional late stage spreading force of 2-6 pN/nm, an osmotic pressure difference of 2-10 kPa, and spontaneous curvature between 0 and 0.04 nm-1. With these conditions, stable membrane conformation sizes and morphologies emerged in concordance with stages of cell plate development. To reach a mature cell plate, our model required the late-stage onset of a spreading/stabilizing force coupled with a concurrent loss of spontaneous curvature. Absence of a spreading/stabilizing force predicts failure of maturation. The proposed model provides a framework to interrogate different players in late cytokinesis and potentially other membrane networks that undergo such transitions. Callose, is a polysaccharide that accumulates transiently during cell plate maturation. Callose-related observations were consistent with the proposed model's concept, suggesting that it is one of the factors involved in establishing the spreading force.

Published
2022

14. Glycosyl transferase GT2 genes mediate the biosynthesis of an unusual (1,3;1,4)‐β‐glucan exopolysaccharide in the bacterium Sarcina ventriculi.

Author

Lampugnani, Edwin R., Ford, Kris, Ho, Yin Ying, van de Meene, Allison, Lahnstein, Jelle, Tan, Hwei‐Ting, Burton, Rachel A., Fincher, Geoffrey B., Shafee, Thomas, Bacic, Antony, Zimmer, Jochen, Xing, Xiaohui, Bulone, Vincent, Doblin, Monika S., and Roberts, Eric M.

Subjects
MICROBIAL exopolysaccharides, BINDING sites, BIOSYNTHESIS, POLYSACCHARIDES, GRAM-positive bacteria, GLUCANS
Abstract

Linear, unbranched (1,3;1,4)‐β‐glucans (mixed‐linkage glucans or MLGs) are commonly found in the cell walls of grasses, but have also been detected in basal land plants, algae, fungi and bacteria. Here we show that two family GT2 glycosyltransferases from the Gram‐positive bacterium Sarcina ventriculi are capable of synthesizing MLGs. Immunotransmission electron microscopy demonstrates that MLG is secreted as an exopolysaccharide, where it may play a role in organizing individual cells into packets that are characteristic of Sarcina species. Heterologous expression of these two genes shows that they are capable of producing MLGs in planta, including an MLG that is chemically identical to the MLG secreted from S. ventriculi cells but which has regularly spaced (1,3)‐β‐linkages in a structure not reported previously for MLGs. The tandemly arranged, paralogous pair of genes are designated SvBmlgs1 and SvBmlgs2. The data indicate that MLG synthases have evolved different enzymic mechanisms for the incorporation of (1,3)‐β‐ and (1,4)‐β‐glucosyl residues into a single polysaccharide chain. Amino acid variants associated with the evolutionary switch from (1,4)‐β‐glucan (cellulose) to MLG synthesis have been identified in the active site regions of the enzymes. The presence of MLG synthesis in bacteria could prove valuable for large‐scale production of MLG for medical, food and beverage applications. [ABSTRACT FROM AUTHOR]

Published
2024
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17. Deciphering the uniqueness of Mucoromycotina cell walls by combining biochemical and phylogenomic approaches.

Author

Mélida, Hugo, Sain, Divya, Stajich, Jason E, and Bulone, Vincent

Abstract

Most fungi from the Mucoromycotina lineage occur in ecosystems as saprobes, although some species are phytopathogens or may induce human mycosis. Mucoromycotina represent early diverging models that are most valuable for understanding fungal evolution. Here we reveal the uniqueness of the cell wall structure of the Mucoromycotina Rhizopus oryzae and Phycomyces blakesleeanus compared with the better characterized cell wall of the ascomycete Neurospora crassa. We have analysed the corresponding polysaccharide biosynthetic and modifying pathways, and highlight their evolutionary features and higher complexity in terms of gene copy numbers compared with species from other lineages. This work uncovers the presence in Mucoromycotina of abundant fucose-based polysaccharides similar to algal fucoidans. These unexpected polymers are associated with unusually low amounts of glucans and a higher proportion of chitin compared with N. crassa. The specific structural features are supported by the identification of genes potentially involved in the corresponding metabolic pathways. Phylogenomic analyses of genes encoding carbohydrate synthases, polysaccharide modifying enzymes and enzymes involved in nucleotide-sugar formation provide evidence for duplication events during evolution of cell wall metabolism in fungi. Altogether, the data highlight the specificity of Mucoromycotina cell walls and pave the way for a finer understanding of their metabolism.

Published
2014

20. An Investigation into Post-Secondary Students' Understanding of Combinatorial Questions

Author

Bulone, Vincent William

Abstract

The purpose of this dissertation was to study aspects of how post-secondary students understand combinatorial problems. Within this dissertation, I considered understanding through two different lenses: i) student connections to previous problems; and ii) common combinatorial distinctions such as ordered versus unordered and repetitive versus non-repetitive. Students were given 12 combinatorial problems and were asked to respond to three rounds of interview questions pertaining to the areas of interest about each problem. Students were not asked to answer each of the 12 problems, but rather were asked to describe how they understood the problems through clinical interviews. The findings indicate that students, in general, do not use connections to previous problems as a means to understanding such questions. Furthermore, students, in general, can distinguish between ordered versus unordered and repetitive and non-repetitive, however, they struggle when questions of the same type (i.e. sequences) are altered in ways that may cause some ambiguity about the problem. The findings also highlight the importance of sets of outcomes and how they play a role with regard to students' struggles with combinatorics questions. In general, I provide a summary on the research that has been conducted within the field of combinatorics education and provide implications to future educators as well as future directions for combinatorics education research. [The dissertation citations contained here are published with the permission of ProQuest LLC. Further reproduction is prohibited without permission. Copies of dissertations may be obtained by Telephone (800) 1-800-521-0600. Web page: http://www.proquest.com/en-US/products/dissertations/individuals.shtml.]

Published
2017

25. Cell Wall Carbohydrate Dynamics during the Differentiation of Infection Structures by the Apple Scab Fungus, Venturia inaequalis

Author

Rocafort, Mercedes, primary, Srivastava, Vaibhav, additional, Bowen, Joanna K., additional, Díaz-Moreno, Sara M., additional, Guo, Yanan, additional, Bulone, Vincent, additional, Plummer, Kim M., additional, Sutherland, Paul W., additional, Anderson, Marilyn A., additional, Bradshaw, Rosie E., additional, and Mesarich, Carl H., additional

Published
2023
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30. Structural Analysis and Identity Confirmation of Anthocyanins in Brassica oleracea Extracts by Direct Injection Ion Mobility-Mass Spectrometry

Author

Pachulicz, River J., Yu, Long, Jovcevski, Blagojce, Bulone, Vincent, Pukala, Tara L., Pachulicz, River J., Yu, Long, Jovcevski, Blagojce, Bulone, Vincent, and Pukala, Tara L.

Abstract

Anthocyanins are a subclass of plant-derived flavonoids that demonstrate immense structural heterogeneity which is challenging to capture in complex extracts by traditional liquid chromatography-mass spectrometry (MS)-based approaches. Here, we investigate direct injection ion mobility-MS as a rapid analytical tool to characterize anthocyanin structural features in red cabbage (Brassica oleracea) extracts. Within a 1.5 min sample run time, we observe localization of structurally similar anthocyanins and their isobars into discrete drift time regions based upon their degree of chemical modifications. Furthermore, drift time-aligned fragmentation enables simultaneous collection of MS, MS/MS, and collisional cross-section data for individual anthocyanin species down to a low picomole scale to generate structural identifiers for rapid identity confirmation. We finally identify anthocyanins in three other Brassica oleracea extracts based on red cabbage anthocyanin identifiers to demonstrate our high-throughput approach. Direct injection ion mobility-MS therefore provides wholistic structural information on structurally similar, and even isobaric, anthocyanins in complex plant extracts, which can inform the nutritional value of a plant and bolster drug discovery pipelines., QC 20231023

Published
2023
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31. Cell Wall Carbohydrate Dynamics during the Differentiation of Infection Structures by the Apple Scab Fungus, Venturia inaequalis

Author

Rocafort, Mercedes, Srivastava, Vaibhav, Bowen, Joanna K., Diaz-Moreno, Sara M, Guo, Yanan, Bulone, Vincent, Plummer, Kim M., Sutherland, Paul W., Anderson, Marilyn A., Bradshaw, Rosie E., Mesarich, Carl H., Rocafort, Mercedes, Srivastava, Vaibhav, Bowen, Joanna K., Diaz-Moreno, Sara M, Guo, Yanan, Bulone, Vincent, Plummer, Kim M., Sutherland, Paul W., Anderson, Marilyn A., Bradshaw, Rosie E., and Mesarich, Carl H.

Abstract

Scab, caused by the biotrophic fungal pathogen Venturia inaequalis, is the most economically important disease of apples. During infection, V. inaequalis colonizes the subcuticular host environment, where it develops specialized infection structures called runner hyphae and stromata. These structures are thought to be involved in nutrient acquisition and effector (virulence factor) delivery, but also give rise to conidia that further the infection cycle. Despite their importance, very little is known about how these structures are differentiated. Likewise, nothing is known about how these structures are protected from host defenses or recognition by the host immune system. To better understand these processes, we first performed a glycosidic linkage analysis of sporulating tubular hyphae from V. inaequalis developed in culture. This analysis revealed that the V. inaequalis cell wall is mostly composed of glucans (44%) and mannans (37%), whereas chitin represents a much smaller proportion (4%). Next, we used transcriptomics and confocal laser scanning microscopy to provide insights into the cell wall carbohydrate composition of runner hyphae and stromata. These analyses revealed that, during subcuticular host colonization, genes of V. inaequalis putatively associated with the biosynthesis of immunogenic carbohydrates, such as chitin and b-1,6-glucan, are downregulated relative to growth in culture, while on the surface of runner hyphae and stromata, chitin is deacetylated to the less-immunogenic carbohydrate chitosan. These changes are anticipated to enable the subcuticular differentiation of runner hyphae and stromata by V. inaequalis, as well as to protect these structures from host defenses and recognition by the host immune system. IMPORTANCE Plant-pathogenic fungi are a major threat to food security. Among these are subcuticular pathogens, which often cause latent asymptomatic infections, making them difficult to control. A key feature of these pathogens is thei, QC 20230823

Published
2023
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40. Multiple enzymatic approaches to hydrolysis of fungal beta-glucans by the soil bacterium Chitinophaga pinensis

Author

Lu, Zijia, Ramgard, Carl, Ergenlioglu, Irem, Sandin, Lova, Hammar, Hugo, Andersson, Helena, King, Katharine, Inman, Annie R., Hao, Mengshu, Bulone, Vincent, and McKee, Lauren S.

Subjects
Biochemistry and Molecular Biology
Abstract

The genome of the soil Bacteroidota Chitinophaga pinensis encodes a large number of glycoside hydrolases (GHs) with noteworthy features and potentially novel functions. Several are predicted to be active on polysaccharide components of fungal and oomycete cell walls, such as chitin, beta-1,3-glucan and beta-1,6-glucan. While several fungal beta-1,6-glucanase enzymes are known, relatively few bacterial examples have been characterised to date. We have previously demonstrated that C. pinensis shows strong growth using beta-1,6-glucan as the sole carbon source, with the efficient release of oligosaccharides from the polymer. We here characterise the capacity of the C. pinensis secretome to hydrolyse the beta-1,6-glucan pustulan and describe three distinct enzymes encoded by its genome, all of which show different levels of beta-1,6-glucanase activity and which are classified into different GH families. Our data show that C. pinensis has multiple tools to deconstruct pustulan, allowing the species' broad utility of this substrate, with potential implications for bacterial biocontrol of pathogens via cell wall disruption. Oligosaccharides derived from fungal beta-1,6-glucans are valuable in biomedical research and drug synthesis, and these enzymes could be useful tools for releasing such molecules from microbial biomass, an underexploited source of complex carbohydrates.

Published
2023

44. Cell wall carbohydrate dynamics during the differentiation of infection structures by the apple scab fungus, Venturia inaequalis

Author

Rocafort, Mercedes, primary, Srivastava, Vaibhav, additional, Bowen, Joanna K., additional, Díaz-Moreno, Sara M., additional, Bulone, Vincent, additional, Plummer, Kim M., additional, Sutherland, Paul W., additional, Anderson, Marilyn A., additional, Bradshaw, Rosie E., additional, and Mesarich, Carl H., additional

Published
2022
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