Article

A Standardized Synthetic Eucalyptus Transcription Factor and Promoter Panel for Re-engineering Secondary Cell Wall Regulation in Biomass and Bioenergy Crops

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Abstract

Re-engineering transcriptional networks regulating secondary cell wall formation may allow the improvement of plant biomass in widely grown plantation crops such as Eucalyptus. However, there is currently a scarcity of freely available standardized biological parts (e.g. Phytobricks) compatible with Type IIS assembly approaches from forest trees, and there is a need to accelerate transcriptional network inference in non-model biomass crops. Here we describe the design and synthesis of a versatile three-panel biological parts collection of 221 secondary cell wall-related Eucalyptus grandis transcription factor coding sequences and 65 promoters that are compatible with GATEWAY, Golden Gate, MoClo and GoldenBraid DNA assembly methods and generally conform to accepted Phytobrick syntaxes. This freely available resource is intended to accelerate synthetic biology applications in multiple plant biomass crops and enable reconstruction of secondary cell wall transcriptional networks using high-throughput assays such as DNA Affinity Purification sequencing (DAP-seq) and enhanced yeast one-hybrid (eY1H) screening.

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... These TFs feature a zinc finger domain (ZF), where six cysteine residues are paired with two zinc atoms to stabilize a scaffold for recognize and bind a specific regulatory sequence in the promoter region (Miyamoto et al. 2020). Recent advances have made it possible to purposefully reengineer individual TF or its ZF domain to significantly alter the transcriptional expression levels of downstream genes (Khalil et al. 2012;Hussey et al. 2019;Gertz et al. 2013). Therefore, the strategy of engineering the clusterspecific TF for a global perturbation of the SM biosynthesis In this study, we identified the DNA-binding residues of the MABGC-specific TF MrPigB and constructed a multi-site saturation mutagenesis library of MrPigB. ...
... In our previous work, a wide range of gene expression levels of MABGC genes was found to be strictly controlled by the MABGC-specific TF MrPigB . It was therefore possible to tune the expression profile of these MA biosynthetic genes by engineering of MrPigB (Hussey et al. 2019;Moses 2009). However, the MrPigBmediated regulation of MABGC genes is poorly understood at present. ...
... TFs with engineered ZF domain could serve as a powerful tool to regulate the target genes (Hussey et al. 2019); thus, MABGC-specific TF MrPigB was engineered to alter the expression profile of MABGC genes. A multi-sitesaturation mutagenesis library of MrPigB was constructed by simultaneous saturation mutagenesis at the predicted DNA-binding residues. ...
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Monascus azaphilones (MAs) have been extensively applied as natural food coloring agents. MAs are classified into three categories: yellow MAs (YMAs), orange MAs (OMAs), and red MAs with various biological activities. However, the exact biosynthetic mechanism of OMAs and YMAs are not thoroughly elucidated. Firstly, we identified four DNA-binding residues of transcription factor MrPigB and constructed a multi-site saturation mutagenesis library of MrPigB. Then, comparative metabolite and gene expression of the mutants revealed that two oxidoreductases MrPigE and MrPigF were responsible for the formation of YMAs and OMAs. Finally, the in vitro and in vivo assays demonstrated the opposite roles of MrPigE and MrPigF in conversion of OMAs to YMAs. To our knowledge, this is the first report of a binary oxidoreductase system for dynamic regulation of fungal secondary metabolite biosynthesis. Broadly, our work also demonstrates the transcription factor engineering strategy for elucidating the biosynthetic pathway of secondary metabolite. Key points • MrPigE converts orange Monascus azaphilones to yellow Monascus azaphilones • MrPigF oxidizes intermediates to afford orange Monascus azaphilones • MrPigE and MrPigF constitute a binary system in Monascus azaphilones biosynthesis Graphical abstract
... Coding sequences of Eucalyptus EgrMYB20 (Eucgr.F02864), EgrMYB46 (Eucgr.B03684), EgrMYB52 (Eucgr.F02756), EgrMYB83 (Eucgr.G03385), EgrMYB85 (Eucgr.D02014) and EgrMYB103 (Eucgr.D01819) from a synthetic gene panel (Hussey et al., 2019) were cloned into pCR8©/GW/TOPO TM (Invitrogen) and then transferred to the pIX-HALO expression vector (The Arabidopsis Interactome Mapping Consortium, 2011). HaloTag-transcription factor (TF) fusion proteins were expressed in vitro (TNT SP6 Coupled Wheat Germ Extract System; Promega), confirmed via Western blot and quantified via semi-quantitative dot blot analysis. ...
... New Phytologist which are homologous to those regulating secondary cell wall (SCW) deposition in Arabidopsis (Hussey et al., 2019). The significant enrichment for accessible chromatin among in vitro binding sites for all six EgrMYB TFs suggests that many of them are functional and can be prioritized within the context of the immature xylem chromatin structure. ...
Article
•Accessible chromatin changes dynamically during development and harbours functional regulatory regions which are poorly understood in the context of wood development. We explored the importance of accessible chromatin in Eucalyptus grandis in immature xylem generally, and MYB transcription factor‐mediated transcriptional programmes specifically. •We identified biologically reproducible DNase I Hypersensitive Site (DHSs) and assessed their functional significance in immature xylem through their associations with gene expression, epigenomic data and DNA sequence conservation. We identified in vitro DNA binding sites for six secondary cell wall‐associated Eucalyptus MYB (EgrMYB) transcription factors using DAP‐seq, reconstructed protein‐DNA networks of predicted targets based on binding sites within or outside DHSs and assessed biological enrichment of these networks with published datasets. •25,319 identified immature xylem DHSs were associated with increased transcription and significantly enriched for various epigenetic signatures (H3K4me3, H3K27me3, RNA pol II), conserved noncoding sequences and depleted single nucleotide variants. Predicted networks built from EgrMYB binding sites located in accessible chromatin were significantly enriched for systems biology datasets relevant to wood formation, while those occurring in inaccessible chromatin were not. •Our study demonstrates that DHSs in E. grandis immature xylem, most of which are intergenic, are of functional significance to gene regulation in this tissue. This article is protected by copyright. All rights reserved.
... Coding sequences of Eucalyptus EgrMYB20 (Eucgr.F02864), EgrMYB46 (Eucgr.B03684), EgrMYB52 (Eucgr.F02756), EgrMYB83 (Eucgr.G03385), EgrMYB85 (Eucgr.D02014) and EgrMYB103 (Eucgr.D01819) from a synthetic gene panel (Hussey et al., 2019) were cloned into pCR8©/GW/TOPO TM (Invitrogen) and then transferred to the pIX-HALO expression vector (The Arabidopsis Interactome Mapping Consortium, 2011). HaloTag-transcription factor (TF) fusion proteins were expressed in vitro (TNT SP6 Coupled Wheat Germ Extract System; Promega), confirmed via Western blot and quantified via semi-quantitative dot blot analysis. ...
... New Phytologist which are homologous to those regulating secondary cell wall (SCW) deposition in Arabidopsis (Hussey et al., 2019). The significant enrichment for accessible chromatin among in vitro binding sites for all six EgrMYB TFs suggests that many of them are functional and can be prioritized within the context of the immature xylem chromatin structure. ...
Conference Paper
Next-generation DNA sequencing technologies along with high-throughput SNP genotyping platforms offer new opportunities for genetic dissection in forest trees. Here we report a high-density SNP genetic linkage map constructed for an F2 interspecific backcross of Eucalyptus grandis x E. urophylla. The backcross family consists of 295 individuals derived from backcrossing an F1 hybrid (E. grandis x E. urophylla) clone as pollen parent with an unrelated E. urophylla individual (seed parent). The parents and backcross progeny were genotyped using the Illumina Infinium II platform using a 60K SNP array (Geneseek, Lincoln, NE). Of the 64,639 SNPs assayed, 19,182 (29.7%) were polymorphic and informative in the backcross. A framework of 3,454 non-redundant markers was selected for even genome coverage and used for linkage map construction. Parental linkage maps were constructed using the “cross-pollinator” mapping strategy in JoinMap 4.1. The E. urophylla map contains 1109 markers spread over 11 linkage groups spanning 1124.7 cM with an average interval of 1.0 cM, while the F1 hybrid map spanning 1021 cM contain 1316 markers at 0.8 cM spacing. Comparison of the genetic distances and marker orders to the corresponding locations in the E. grandis genome (Phytozome) revealed several possible inconsistencies in the predicted physical positions of SNP markers. Besides their value for refining the Eucalyptus reference genome assembly and for comparative genomics in Eucalyptus, the linkage maps reported here will be useful for genetic dissection of growth and wood property traits segregating in the F1 hybrid of E. grandis x E. urophylla.
... Promoters are gene switches, located upstream of gene coding regions, and contain specific cis-acting elements that play a crucial role in gene transcription and expression [52,53]. The main active position of cis-acting elements with a biological function is 50 bp upstream of the transcription start sites (TSS), and most transcription factor binding sites (TFBS) are in the region of −1000 bp to +200 bp relative to the TSS [54]. ...
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The MYB (v-myb avian myeloblastosis viral oncogene homolog) transcription factor family plays an important role in plant growth, development, and response to biotic and abiotic stresses. However, the gene functions of MYB transcription factors in sweet potato (Ipomoea batatas (L.) Lam) have not been elucidated. In this study, an MYB transcription factor gene, IbMYB308, was identified and isolated from sweet potato. Multiple sequence alignment showed that IbMYB308 is a typical R2R3-MYB transcription factor. Further, quantitative real-time PCR (qRT-PCR) analysis revealed that IbMYB308 was expressed in root, stem, and, especially, leaf tissues. Moreover, it showed that IbMYB308 had a tissue-specific profile. The experiment also showed that the expression of IbMYB308 was induced by different abiotic stresses (20% PEG-6000, 200 mM NaCl, and 20% H2O2). After a 200 mM NaCl treatment, the expression of several stress-related genes (SOD, POD, APX, and P5CS) was upregulation in transgenic plants, and the CAT activity, POD activity, proline content, and protein content in transgenic tobacco had increased, while MDA content had decreased. In conclusion, this study demonstrated that IbMYB308 could improve salt stress tolerance in transgenic tobacco. These findings lay a foundation for future studies on the R2R3-MYB gene family of sweet potato and suggest that IbMYB308 could potentially be used as an important positive factor in transgenic plant breeding to improve salt stress tolerance in sweet potato plants.
... The synthetic, domesticated Phytobrick version of the EgrIAA13 coding DNA sequence (CDS) (Hussey et al. 2019) was cloned into overexpression (pCAMBIA 1305.1 GW +) and RNAi knockdown (pCAMBIA 1305.1 GW-) Gateway enabled cassettes (driven by Cauliflower mosaic virus 35S promoter (35S)) using Gateway LR Clonase™ II Enzyme mix (Invitrogen) according to the manufacturer's instructions (Spokevicius et al. 2007). The empty (without CDS) pCAMBIA 1305.1 GW + (EVC +) and empty (without CDS) pCAMBIA 1305.1 GW-(EVC-) vectors were used as controls. ...
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Key message Our Induced Somatic Sector Analysis and protein–protein interaction experiments demonstrate that Eucalyptus grandis IAA13 regulates xylem fibre and vessel development, potentially via EgrIAA13 modules involving ARF2, ARF5, ARF6 and ARF19. Abstract Auxin is a crucial phytohormone regulating multiple aspects of plant growth and differentiation, including regulation of vascular cambium activity, xylogenesis and its responsiveness towards gravitropic stress. Although the regulation of these biological processes greatly depends on auxin and regulators of the auxin signalling pathway, many of their specific functions remain unclear. Therefore, the present study aims to functionally characterise Eucalyptus grandis AUX/INDOLE-3-ACETIC ACID 13 ( EgrIAA13 ), a member of the auxin signalling pathway. In Eucalyptus and Populus , Egr IAA13 and its orthologs are preferentially expressed in the xylogenic tissues and downregulated in tension wood. Therefore, to further investigate EgrIAA13 and its function during xylogenesis, we conducted subcellular localisation and Induced Somatic Sector Analysis experiments using overexpression and RNAi knockdown constructs of EgrIAA13 to create transgenic tissue sectors on growing stems of Eucalyptus and Populus . Since Aux/IAAs interact with Auxin Responsive Factors (ARFs), in silico predictions of IAA13-ARF interactions were explored and experimentally validated via yeast-2-hybrid experiments. Our results demonstrate that EgrIAA13 localises to the nucleus and that downregulation of EgrIAA13 impedes Eucalyptus xylem fibre and vessel development. We also observed that EgrIAA13 interacts with Eucalyptus ARF2, ARF5, ARF6 and ARF19A. Based on these results, we conclude that EgrIAA13 is a regulator of Eucalyptus xylogenesis and postulate that the observed phenotypes are likely to result from alterations in the auxin-responsive transcriptome via IAA13-ARF modules such as EgrIAA13-EgrARF5. Our results provide the first insights into the regulatory role of EgrIAA13 during xylogenesis.
... The SCW provides rigidity and strength for plants to support their body weight and ensure water and nutrient transport (Oda and Fukuda, 2012;Wang and Dixon, 2012). It is mainly composed of lignin, cellulose, and hemicellulose, and their biosynthesis was highly related to the transformation and production of biofuels and biological products (Mohnen, 2008;Hussey et al., 2019). It is regulated by microRNA, MYB, NAC, and WRKY in the SCW complex regulatory network (Zhang et al., 2018b). ...
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Poplar is an important bioenergy tree species. lncRNAs play important roles in various biological regulatory processes, and their expression pattern is more tissue-specific than mRNAs. In this study, P. deltoides “Danhong” (Pd) and P. simonii “Tongliao1” (Ps) with different growth rates and wood quality were used as experimental materials, and the transcriptomes of their shoot apical meristem, xylem, and phloem were sequenced. Furthermore, high-throughput RNA sequencing analysis revealed that the expression patterns of genes and lncRNAs are different between the two genotypes. 6,355 lncRNAs were identified. Based on target prediction, lncRNAs and target genes were involved in ADP binding, oxidoreductase activity, phenylpropanoid biosynthesis, and cyanoamino acid metabolism. The DElncRNAs in two poplars were co-expressed with transcription factors and structural genes of lignin and flavonoid pathways. In addition, we found the potential target lncRNAs of miRNA. This result provides basic evidence for a better understanding of the regulatory role of lncRNAs in regulating phenylalanine molecular pathways and wood formation.
... The released assembly of E. grandis had totally 55,643 genes belonging to various classes like protein coding, non-coding, pseudogenes and genes with variants [30]. Recent studies in eucalypts highlighted the role of TFs and PKs associated with secondary cell wall development [31], biotic resistance [32] and abiotic tolerance [33,34]. Accordingly, results of this study offer a comprehensive view of regulatory sequences associated with almost all essential cellular functions and provides a foundation for further characterization. ...
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Eucalyptus is one of the major plantation species with wide variety of industrial uses. Polymorphic and informative simple sequence repeats (SSRs) have broad range of applications in genetic analysis. In this study, two individuals of Eucalyptus tereticornis (ET217 and ET86), one individual each from E. camaldulensis (EC17) and E. grandis (EG9) were subjected to whole genome resequencing. Low coverage (10×) genome sequencing was used to find polymorphic SSRs between the individuals. Average number of SSR loci identified was 95,513 and the density of SSRs per Mb was from 157.39 in EG9 to 155.08 in EC17. Among all the SSRs detected, the most abundant repeat motifs were di-nucleotide (59.6%-62.5%), followed by tri- (23.7%-27.2%), tetra- (5.2%-5.6%), penta- (5.0%-5.3%), and hexa-nucleotide (2.7%-2.9%). The predominant SSR motif units were AG/CT and AAG/TTC. Computational genome analysis predicted the SSR length variations between the individuals and identified the gene functions of SSR containing sequences. Selected subset of polymorphic markers was validated in a full-sib family of eucalypts. Additionally, genome-wide characterization of single nucleotide polymorphisms, InDels and transcriptional regulators were carried out. These variations will find their utility in genome-wide association studies as well as understanding of molecular mechanisms involved in key economic traits. The genomic resources generated in this study would provide an impetus to integrate genomics in marker-trait associations and breeding of tropical eucalypts.
... Despite these advancements, the conversion of natural DNA sequences into Phytobricks can be laborious, and the removal of "illegal" restriction sites in these sequences to enable Type IIS assembly can introduce unintended alterations to the part's function. In the recent production of a standardized parts library of 221 Eucalyptus transcription factors and 65 promoters [98], the risk of altering promoter function in their conversion to Phytobricks was minimized by using known single nucleotide polymorphism data in Eucalyptus populations to mutate undesirable restriction sites. ...
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Human life intimately depends on plants for food, biomaterials, health, energy, and a sustainable environment. Various plants have been genetically improved mostly through breeding, along with limited modification via genetic engineering, yet they are still not able to meet the ever-increasing needs, in terms of both quantity and quality, resulting from the rapid increase in world population and expected standards of living. A step change that may address these challenges would be to expand the potential of plants using biosystems design approaches. This represents a shift in plant science research from relatively simple trial-and-error approaches to innovative strategies based on predictive models of biological systems. Plant biosystems design seeks to accelerate plant genetic improvement using genome editing and genetic circuit engineering or create novel plant systems through de novo synthesis of plant genomes. From this perspective, we present a comprehensive roadmap of plant biosystems design covering theories, principles, and technical methods, along with potential applications in basic and applied plant biology research. We highlight current challenges, future opportunities, and research priorities, along with a framework for international collaboration, towards rapid advancement of this emerging interdisciplinary area of research. Finally, we discuss the importance of social responsibility in utilizing plant biosystems design and suggest strategies for improving public perception, trust, and acceptance.
... Similarly, there has been significant recent progress in systems biology tools. DNA affinity purification sequencing (DAP-Seq or DAP-chip) enables low-cost and highthroughput profiling genome-wide DNA-binding motifs of individual transcription factors (TFs) [13,14] to facilitate characterization of regulatory architecture of transcriptional networks [15,16]. Likewise, high-throughput proteomic analysis capable of analyzing hundreds of samples per day [17], tools to shorten method development time [18], and automated proteomic sample preparation procedures [19] make these types of experiments routine. ...
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Plant cell walls contain the most abundant sustainable biomass resource for biofuels and biomaterials on the earth. However, lignocellulose recalcitrance generally requires a costly biomass process unacceptable for large-scale bioethanol production with the potential formation of secondary wastes. To address this bottleneck-like biomass recalcitrance issue, this review attempts to connect recent innovation progress regrading up-stream lignocellulose modification, middle-stream cellulases production and down-stream biomass processing. Particularly, the site-specific gene editing is demonstrated for precise and mild modification of plant cell walls to generate recalcitrance-much-reduced cellulose nanofibers, which not only leads to little impact on plant strength and biomass yield, but also causes remarkably enhanced enzymatic saccharification in major bioenergy crops. By selecting the size-reduced cellulose nanofibers of engineered bioenergy crops as enzyme-inducing substrate, fungal strains are then engineered to secret the optimal cellulases enzymes cocktails enabled for complete enzymatic saccharification of diverse lignocellulose residues from cost-effective biomass pretreatments. Consequently, engineered yeast strains could use both hexoses and xylose released from complete saccharification as carbon sources for maximum bioethanol production by an efficient co-fermentation. Finally, the green-like processing technology is introduced to generate biomaterials and biochemicals by using the remaining lignin-rich residues. Therefore, this work has originally proposed a novel strategy that dynamically cascades the engineered bioenergy crops and fungal strains with the advanced biomass process technology, which should be considered as next generation of integrated biotechnology for both cost-effective biofuels production and value-added bioproducts with minimum waste releases into the environment.
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The current status of lignocellulosic biomass as an invaluable resource in industry, agriculture, and health has spurred increased interest in understanding the transcriptional regulation of secondary cell wall (SCW) biosynthesis. The last decade of research has revealed an extensive network of NAC, MYB and other families of transcription factors regulating Arabidopsis SCW biosynthesis, and numerous studies have explored SCW-related transcription factors in other dicots and monocots. Whilst the general structure of the Arabidopsis network has been a topic of several reviews, they have not comprehensively represented the detailed protein-DNA and protein-protein interactions described in the literature, and an understanding of network dynamics and functionality has not yet been achieved for SCW formation. Furthermore the methodologies employed in studies of SCW transcriptional regulation have not received much attention, especially in the case of non-model organisms. In this review, we have reconstructed the most exhaustive literature-based network representations to date of SCW transcriptional regulation in Arabidopsis. We include a manipulable Cytoscape representation of the Arabidopsis SCW transcriptional network to aid in future studies, along with a list of supporting literature for each documented interaction. Amongst other topics, we discuss the various components of the network, its evolutionary conservation in plants, putative modules and dynamic mechanisms that may influence network function, and the approaches that have been employed in network inference. Future research should aim to better understand network function and its response to dynamic perturbations, whilst the development and application of genome-wide approaches such as ChIP-seq and systems genetics are in progress for the study of SCW transcriptional regulation in non-model organisms.
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Tension wood has distinct physical and chemical properties, including altered fibre properties, cell wall composition and ultrastructure. It serves as a good system for investigating the genetic regulation of secondary cell wall biosynthesis and wood formation. The reference genome sequence for Eucalyptus grandis allows investigation of the global transcriptional reprogramming that accompanies tension wood formation in this global wood fibre crop. We report the first comprehensive analysis of physicochemical wood property changes in tension wood of Eucalyptus measured in a hybrid (E. grandis × Eucalyptus urophylla) clone, as well as genome-wide gene expression changes in xylem tissues 3 wk post-induction using RNA sequencing. We found that Eucalyptus tension wood in field-grown trees is characterized by an increase in cellulose, a reduction in lignin, xylose and mannose, and a marked increase in galactose. Gene expression profiling in tension wood-forming tissue showed corresponding down-regulation of monolignol biosynthetic genes, and differential expression of several carbohydrate active enzymes. We conclude that alterations of cell wall traits induced by tension wood formation in Eucalyptus are a consequence of a combination of down-regulation of lignin biosynthesis and hemicellulose remodelling, rather than the often proposed up-regulation of the cellulose biosynthetic pathway.
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NAC domain transcription factors regulate many developmental processes and stress responses in plants and vary widely in number and family structure. We analysed the characteristics and evolution of the NAC gene family of Eucalyptus grandis, a fast-growing forest tree in the rosid order Myrtales.NAC domain genes identified in the E. grandis genome were subjected to amino acid sequence, phylogenetic and motif analyses. Transcript abundance in developing tissues and abiotic stress conditions in E. grandis and E. globulus was quantified using RNA-seq and reverse transcription quantitative PCR (RT-qPCR).One hundred and eighty-nine E. grandis NAC (EgrNAC) proteins, arranged into 22 subfamilies, are extensively duplicated in subfamilies associated with stress response. Most EgrNAC genes form tandem duplicate arrays that frequently carry signatures of purifying selection. Sixteen amino acid motifs were identified in EgrNAC proteins, eight of which are enriched in, or unique to, Eucalyptus. New candidates for the regulation of normal and tension wood development and cold responses were identified.This first description of a Myrtales NAC domain family reveals an unique history of tandem duplication in stress-related subfamilies that has likely contributed to the adaptation of eucalypts to the challenging Australian environment. Several new candidates for the regulation of stress, wood formation and tree-specific development are reported.
Article
Keywords: cambium; Eucalyptus grandis ; evolution; R2R3-MYB transcription factors; secondary growth; tandem duplications; wood formation Summary The R2R3-MYB family, one of the largest transcription factor families in higher plants, controls a wide variety of plant-specific processes including, notably, phenylpropanoid metabolism and secondary cell wall formation. We performed a genome-wide analysis of this superfamily in Eucalyptus, one of the most planted hardwood trees world-wide. A total of 141 predicted R2R3-MYB sequences identified in the Eucalyptus grandis genome sequence were subjected to comparative phylogenetic analyses with Arabidopsis thaliana, Oryza sativa, Populus trichocarpa and Vitis vinifera. We analysed features such as gene structure, conserved motifs and genome location. Transcript abundance patterns were assessed by RNAseq and validated by high-throughput quantitative PCR. We found some R2R3-MYB subgroups with expanded membership in E. grandis, V. vinifera and P. trichocarpa, and others preferentially found in woody species, suggesting diversification of specific functions in woody plants. By contrast, subgroups containing key genes regulating lignin biosynthesis and secondary cell wall formation are more conserved across all of the species analysed. In Eucalyptus, R2R3-MYB tandem gene duplications seem to disproportionately affect woody-preferential and woody-expanded subgroups. Interestingly, some of the genes belonging to woody-preferential subgroups show higher expression in the cambial region, suggesting a putative role in the regulation of secondary growth.
Navigating the transcriptional roadmap regulating plant secondary cell wall deposition
  • S G Hussey
  • E Mizrachi
  • N M Creux
  • A A Myburg
  • N J Patron
  • D Orzaez
  • S Marillonnet
  • H Warzecha
  • C Matthewman
  • M Youles
  • O Raitskin
  • A Leveau
  • G Farre
  • C Rogers
Hussey, S. G., Mizrachi, E., Creux, N. M., and Myburg, A. A. (2013) Navigating the transcriptional roadmap regulating plant secondary cell wall deposition. Front. Plant Sci. 4, 325. (2) Patron, N. J., Orzaez, D., Marillonnet, S., Warzecha, H., Matthewman, C., Youles, M., Raitskin, O., Leveau, A., Farre, G., Rogers, C., et al. (2015) Standards for plant synthetic biology: a common syntax for exchange of DNA parts. New Phytol. 208, 13−19.