Maths paper and very easy question, Cheat Sheet of Biology

Download Maths paper and very easy question and more Cheat Sheet Biology in PDF only on Docsity! Xx http: //www.diva-portal. org This is the published version of a paper published in Frontiers in Plant Science. Citation for the original published paper (version of record): Myburg, A A., Hussey, S G., Street, N., Street, N R., Mizrachi, E. (2019) Systems and Synthetic Biology of Forest Trees: A Bioengineering Paradigm for Woody Biomass Feedstocks Frontiers in Plant Science, 10: 775 https: //doi.org/10.3389/fpls.2019.00775 Access to the published version may require subscription. N.B. When citing this work, cite the original published paper. Permanent link to this version http: //urn.kb.se/resolve?urn=urn: nbn:se:umu: diva-161590 + frontiers in Plant Science OPEN ACCESS Edited by: Wout Boerjan, Flanders Institute for Biotechnology, Belgium Reviewed by: Kelly Mayrink, University of Florida, United States Daniel Conde, University of Florida, United States *Correspondence: Alexander A. Myburg zandermyburg@up.ae.2a; Zander myburg@fabi.up.ac.za Specialty section: This article was submitted to Plant Biotechnology, a section of the journal Frontiers in Plant Science Received: 22 August 2018 Accepted: 28 May 2019 Published: 20 June 2019 Citation: Myburg AA, Hussey SG, Wang JP Street NR and Mizrachi E (2019) Systems and Synthetic Biology of Forest Trees: A Bioengineering Paradigm for Woody Biomass Feedstocks. Front. Plant Sci. 10:775. doi: 10.3389/fpls.2019.00775 Systems and Synthetic Biology of Forest Trees: A Bioengineering Paradigm for Woody Biomass Feedstocks Alexander A. Myburg™, Steven G. Hussey’, Jack P. Wang’, Nathaniel R. Street? and Eshchar Mizrachi’ ‘Department of Biochemistry, Genetics and Microbiology, Forestry and Agricultural Biotechnology Institute (FAB), University of Pretoria, Hatfiakd, South Africa, “Forest Biotechnology Group, Department of Forestry and Environmental Resources, North Caroina State University, Raleigh, NC, United States, Umea Plant Science Center, Department of Plant Physiology, Umea University, Umea, Sweden Fast-growing forest plantations are sustainable feedstocks of plant biomass that can serve as alternatives to fossil carbon resources for materials, chemicals, and energy. Their ability to efficiently harvest light energy and carbon from the atmosphere and sequester this into metabolic precursors for lignocellulosic biopolymers and a wide range of plant specialized metabolites make them excellent biochemical production platforms and living biorefineries. Their large sizes have facilitated multi-omics analyses and systems modeling of key biological processes such as lignin biosynthesis in trees. High-throughput ‘omics’ approaches have also been applied in segregating tree populations where genetic variation creates abundant genetic perturbations of system components allowing construction of systems genetics models linking genes and pathways to complex trait variation. With this information in hand, itis now possible to start using synthetic biology and genome editing techniques in a bioengineering approach based on a deeper understanding and rational design of biological parts, devices, and integrated systems. However, the complexity of the biology and interacting components will require investment in big data informatics, machine learning, and intuitive visualization to fully explore multi-dimensional patterns and identify emergent properties of biological systems. Predictive systems models could be tested rapidly through high-throughput synthetic biology approaches and multigene editing. Such a bioengineering paradigm, together with accelerated genomic breeding, will be crucial for the development of a new generation of woody biorefinery crops. Keywords: synthetic biology (synbio), systems biology, systems genetics, woody biomass, biorefinery, bioeconomy, lignin biosynthesis, wood formation INTRODUCTION Compared to herbaceous plants, forest trees afford numerous of advantages to plant biologists interested in studying growth and development. Most obviously, trees produce vast quantities of wood comprising multiple cells types produced from the meristematic cambial initials of the vascular cambium. In the context of developmental studies of secondary growth, greater size 1 June 2 Myburg et al is and Synthetic Biology o woop + Lignin engineering + Polysaccharide engineering + xylem patterning engineering PHLOEM + Underetudied + Madifying photoascimilates and transport sugars? + Rapping’ fine chemicals? (e.g engineering phloem specific pathwvaye in trees far continuous production} Developmentfcondition CORK and BARK + Understudied + Suberintamnin praduction and engineering + Cork morphology and corkcxylern ratio + Bark morphology engineering for ecology (e.g. rougness) of processing FIGURE 1 | Systems approaches to study biological processes in trees. Axes represent different dimensions or types of perturbation that can be used to dissect biological processes. Network components (colored nodes and shapes) represent the study of different biological layers (DNA, RNA, protein, metabolite, etc.) using integrative approaches that take into account development/condition (Systems biology), genetic diversity (Systems genetics), and evolution (Evo-devo). Yellow nodes in “systems genetics” indicate the connection to complex traits that can be established in population-wide studies. Networks on the left represent intermediate integration ‘steps of components that vary along these axes. Networks on the right represent integrated networks that can be constructed from each type of perturbation. Boxes show target traits and processes associated with different tissues or organs where systems approaches may provide new opportunities for research. Systems biology LEAVES + Improved pphotosynthesisicartion fixation + Architecture + Specialized secondary metaboltesioilsivaxes (akenes, tannins, phenolic, terpenes) +_ Novel defence compounds ase e © we? Systems genetics FRUITSNUTS/SEEDS. + Novel pectins ar engineering of added nutrition (e.g. MLG in dicate?) + Oils tespecially in nuts like Sheaimacadamia‘ail palm etc.) “ts e eo ©, + oe © ROOTS ke © | + Optimizing architecture and relative e investment compared to rest of tree + Nitragen and phosphorus acquiston Eve devo thraugh synthetic biology + Droughtiflaod resistanesitolerance + Biotic stress resistanceltolerance There is now extensive knowledge of alternative splicing and transcript usage, which can vary among tissues, during development or among genotypes (Bao et al., 2013; Xu et al., 2014; Zhao et al. 2014). Mapping expression at the transcript level will provide greater resolution to the link between genome and phenotype. However, fine-scale expression analysis at the transcript level remains challenging, mainly because most tissue samples contain a complex mix of transcripts and splicing variants from different cell types or cells at different stages of development. There is equally a need for improvement in network inference methods, with an understanding that no single current method is adequate to capture the range of interactions present in biological networks (Marbach et al. 2012) and with few tools available to facilitate aggregate inference approaches (Schiffthaler et al., 2018). Similarly, no studies have yet integrated genome-wide assays of DNA modifications or accessibility despite increasing evidence of the additional insight such information brings. There is also a paucity of large-scale transcription factor binding or protein-protein interaction data for plants in general, which further limits comprehensive understanding. SYNTHETIC BIOLOGY: A NEW BIOENGINEERING PARADIGM FOR FOREST TREES SynBio has made its greatest advancements in prokaryotes and single-celled eukaryotes such as yeast, but plant synthetic biology is catching up (Patron et al Schaumberg et al, 2016; Frontiers in Plant de Lange et al., 2018; Hanson and Jez, 2018; Pouvreau et al., 2018). In addition to a large number of modifications made by conventional transgenic approaches (Chang et al., 2018), there have been some notable successes in the synthetic modification of trees using single-gene strategies such as the introduction of chemically labile ester linkages into the lignin backbone of poplar trees through the xylem-specific expression of an exogenous feruloyl-coenzyme A monolignol transferase (FMT) from Angelica sinensis (Wilkerson et al., 2014). Future strategies will attempt to evaluate far more complex designs, relying to a large extent on the ability to assemble DNA fragments idempotently (that is, the flexibility to assemble basic parts with increasing complexity using a universal method and without having to re-modify each intermediate). There is currently a scarcity of freely available standardized biological parts suitable for plant biology akin to the International Genetically Engineered Machines (iGEM) BioBrick parts collection’. Encouragingly, the Phytobrick synthetic biology standard with a universal lexicon for plant gene elements coupled to powerful Type IIS idempotent assembly methods (Engler et al., 2014; Patron et al., 2015) is fast gaining traction, with an increasing number of Phytobricks now included in the iGEM Standard Registry”. Tree biologists and biotechnologists should adapt to this conceptual framework as soon as possible to keep up with synthetic biology development in annual crops. We recently developed an open access synthetic panel of 221 ‘http:/parts.igem.org *http://parts.igem.org/Collections/Plants Myburg et al Syste! s and Syntheti Machine DS i. Design | abrcation and tes ation and testing v. Application Elite parent 1 Early flowering DvAsynthesig | Construct Simple Mulkicellular gt assembly chassis : — cotrses a ch Elite parent 2 systems Model * “i ge », a bite porent 1 ia eqgous “a fas FIGURE 2 | A proposed bioengineering paradigm based on forest tree synthetic biology. (i) Systems biology models inform the initial design of synthetic multigene constructs. Individual components (DNA parts) are sourced from existing or novel genetic resources (e.g., bioprospecting), synthesized, and submitted to biological parts collections as standardized Phytobricks (li). The fabrication and testing phase (ii) involves high-throughput idempotent construct assembly folowed by transformation and testing in a simplified chassis such as a cell culture derived from the target tree species (iv). Construct expression and phenotypic data are then integrated into a machine learning model to optimize the construct design, an iterative process that produces a reduced number of semi-optimized constructs for validation in a genotype of the target species selected to perform favorably in laboratory conditions and conducive to transformation ("ab rat"). Such validation may include greenhouse trials involving juvenile trees or mature trees in field trials. (v) Successful constructs may be introduced into a preferred elite parental genotype for intraspecific or interspecific hybrid breeding. A possible avenue to rapidly mobilize synthetic gene constructs into more diverse genetic backgrounds would be to introduce an early flowering construct (such as overexpression of the FLOWERING LOCUS T gene; FT-OX) into a number of elite parental genotypes. Such genotypes can then be transformed with the optimized synthetic gene constructs and used as female parents in crosses with non-transgenic (wid-type, WT) parents to produce F1 progeny segregating for both constructs. If the two constructs are on different chromosomes, approximately 25% of the progeny should be WT for growth and flowering but contain the synthetic gene construct. Early flowering parental genotypes can be propagated in vitro to be transformed with vatious synthetic constructs and, if unrelated, different parental genotypes could be crossed for transgene stacking. oe Sea’egat on | F1 hybrids 09 Standardized biological Parts collection Fy a 2 rox Target species 25% 25% ii. Standardization iv. Chassis development secondary cell wall-related Eucalyptus grandis transcription factors and 65 promoter sequences in partnership with the Department of Energy Joint Genome Institute, most of which were designed as Phytobricks (Hussey et al., in preparation). Accessibility of high-throughput DNA synthesis services will ensure that a growing number of standardized parts become available under open material transfer agreements. One considerable challenge to tree synthetic biology is precise spatiotemporal control of complex multigene constructs, especially in woody tissues where inducing gene expression with external agents is impractical. Such constructs must function optimally in a tissue of interest, be resistant to eukaryotic silencing mechanisms such as RNAi or epigenetic silencing, be somatically stable such that somatic mutations that disrupt a synthetic construct should not be selectively favored, take into account compartmentalized plant cell biology, and have built-in biosafety mechanisms preventing transgene escape. Furthermore, synthetic gene circuits should consist of composable parts (de Lange et al., 2018) that individually encode defined and transferrable functions (a property known as modularity) and that function independently of endogenous processes to avoid unwanted interference, a property known as orthogonality. Designer transcription factors based on zinc finger, TALE, and dCas9 technologies targeting endogenous or synthetic promoters (Liu and Stewart, 2016) are ideal orthogonal synthetic tools Frontiers in Plant ntiersin.org 5 that allow considerable transgene regulation flexibility but may require extensive testing and optimization. Currently, thousands of iterations of multigene constructs can be produced by robotics-assisted DNA Foundry services. However, it is not feasible to transform and phenotype thousands of transgenic trees for iterative design-build-test-learn cycles envisioned for plant synthetic biology (Pouvreau et al., 2018). Early synthetic designs will therefore have to be tested ina simplified system (or “chassis” in synthetic biology terminology) until optimized constructs can be evaluated in target tree species. This will necessitate the development of experimental chassis derived from the target tree species that are easier to transform and phenotype en masse, such as protoplasts, cell cultures, or agroinfiltrated leaves, or “lab rat” genotypes that perform well in tissue culture and have high transformation rates (Figure 2). In vitro tracheary element transdifferentiation approaches relying on hormonal induction (Fukuda and Komamine, 1980; Kubo et al., 2005; Saito et al., 2017), or the VND7 inducible system (Yamaguchi et al., 2010; Goué et al., 2013), for example, could be used to induce secondary cell wall formation in suspension culture cells or explant tissues of a target tree species and thus evaluate the phenotypes of many cell wall-modifying constructs before semi-optimized constructs are tested in a multicellular tree model. Successful constructs may either be directly introduced into an elite June ume 1 Myburg et al Syste! s and Syntheti genotype of the target species or rapidly crossed into elite breeding material after co-transformation of parental genotypes with an early flowering construct (Figure 2). FUTURE PERSPECTIVE Most fundamental discoveries, including proof-of-concept cell wall and growth modifications (and even extrapolations on biomass processing efficiency), are still derived from the analysis of Arabidopsis inflorescence stems, which remains a poor representation of large tree stems comprised mainly of wood. Of priority in the short term is testing genetic perturbations as much as possible in a woody model such as Populus and, if possible, directly in target species of interest. Several priorities must be met here, such as enhancing the transformation efficiency of commercial species or genotypes, capacity for large-scale transformation experiments, as well as (crucially) field trials confirming greenhouse phenotypes in mature trees. Large consortia and industry collaborations, as well as engagement and an improvement in the regulatory landscape, must be met for this to be truly realized. Also in the short to medium term, the convergence of high-resolution technologies that capture genomics, epigenomics, and other cell ‘omics, phenomics, and environment (including microbiome) data, as well as computational modeling of the interactions of these, requires transdisciplinary innovations and probably the application of artificial intelligence methodology. Combined with genome editing (with broader scale synthetic biology applications), this makes the field of forest biotechnology ripe for a new wave of creativity, especially in thinking of the tree itself as a living biorefinery and as a stable and continuous producer of specialized high-value compounds or polymers in sustainably harvestable tissues and organs such as leaves, secondary phloem, and bark. Higher resolution knowledge of metabolite precursors, tissue-specific pathway engineering, and knowledge of novel high-value derivatives that can be discovered using bioprospecting methods and produced in trees has the REFERENCES Bao, H., Li, E., Mansfield, S. D., Cronk, Q. C., El-Kassaby, Y. A., and Douglas, C. J. (2013). The developing xylem transcriptome and genome-wide analysis of alternative splicing in Populus trichocarpa (black cottonwood) populations. 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M., Frey, B. L., and Smith, L. M. (2016). Human proteomic variation revealed by combining RNA-Seq Frontiers in Plant ntiersin.org potential for a new generation of relatively low volume, but high value, forest products. How far does the application of these technologies go? Given the long rotation times of forest trees as harvestable biomass crops, it is unlikely (and indeed not essential) that we will see movement toward a “bottom up” approach that builds on a synthetic minimal tree genome. It is much more important to optimize the precise introduction of complex regulatory circuits and metabolic pathways that remain stable through breeding generations, a nascent field of research in itself. Such a bioengineering paradigm, combined with advanced genomic breeding approaches and accelerated flowering technologies, may empower rapid development of woody biomass crops tailored for diverse biorefinery, biomaterials, and timber construction products. In many forest-growing countries, an advanced forest products industry will be one of the cornerstones of the bioeconomy and key to achieving global sustainable development goals. AUTHOR CONTRIBUTIONS All authors contributed to the drafting and editing of the manuscript. FUNDING This study was supported by National Research Foundation of South Africa (Research Funding, NRF Bioinformatics and Functional Genomics Programme grant UID 97911), Department of Science and Technology of South Africa, (Research Funding) Technology Innovation Agency (TIA) of South Africa (Research Support), Sappi and Mondi (Forestry Industry Partners and Research Support), and University of Pretoria (Research Facilities and Support). NRS is supported by the Trees for the Future (T4F) project (Sweden). proteogenomics and global post-translational modification (G-PTM) search strategy. J. Proteome Res, 15, 800-808. doi: 10,1021 /acs.jproteome.Sb00817 Chang, S., Mahon, E. L., Mackay, H. A. Rottmann, W. 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