Navigating the transcriptional roadmap regulating plant secondary cell wall deposition

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.The Mandela Rhodes Foundation and National Research Foundation of South Africahttp://www.frontiersin.orgam201

Induced somatic sector analysis of cellulose synthase (CesA) promoter regions in woody stem tissues

The increasing focus on plantation forestry as a renewable source of cellulosic biomass has emphasized the need for tools to study the unique biology of woody genera such as Eucalyptus, Populus and Pinus. The domestication of these woody crops is hampered by long generation times, and breeders are now looking to molecular approaches such as marker-assisted breeding and genetic modification to accelerate tree improvement. Much of what is known about genes involved in the growth and development of plants has come from studies of herbaceous models such as Arabidopsis and rice. However, transferring this information to woody plants often proves difficult, especially for genes expressed in woody stems. Here we report the use of induced somatic sector analysis (ISSA) for characterization of promoter expression patterns directly in the stems of Populus and Eucalyptus trees. As a case study, we used previously characterized primary and secondary cell wall-related cellulose synthase (CesA) promoters cloned from Eucalyptus grandis. We show that ISSA can be used to elucidate the phloem and xylem expression patterns of the CesA genes in Eucalyptus and Populus stems and also show that the staining patterns differ in Eucalyptus and Populus stems. These findings show that ISSA is an efficient approach to investigate promoter function in the developmental context of woody plant tissues and raise questions about the suitability of heterologous promoters for genetic manipulation in plant species.This work was supported through funding provided by Mondi and Sappi to the Forest Molecular Genetics (FMG) Programme, the Technology and Human Resources for Industry Programme (THRIP) and the National Research Foundation (NRF) of South Africa as well as a Linkage Grant from the Australian Research Council (LP0776563) to GB, AAM and AVS.http://link.springer.com/journal/425hb201

Diversity and cis-element architecture of the promoter regions of cellulose synthase genes in Eucalyptus

Lignocellulosic biomass from fast-growing plantation trees is composed of carbohydrate-rich materials deposited into plant cell walls in a coordinated manner during wood formation. The diversity and evolution of the transcriptional networks regulating this process have not been studied extensively.We investigated patterns of species-level nucleotide diversity in the promoters of cellulose synthase (CesA) genes from different Eucalyptus tree species and assessed the possible roles of DNA sequence polymorphism in the gain or loss of cis-elements harboured within the promoters. Promoter regions of three primary and three secondary cell wall-associated CesA genes were isolated from 13 Eucalyptus species and were analysed for nucleotide and cis-element diversity. Species-level nucleotide diversity (π) ranged from 0.014 to 0.068, and different CesA promoters exhibited distinct patterns of sequence conservation. A set of 22 putative cis-elements were mapped to the CesA promoters using in silico methods. Forty-two percent of the mapped cis-element occurrences contained singleton polymorphisms which resulted in either gain or loss of a ciselement in a particular Eucalyptus species. The promoters of Eucalyptus CesA genes contained regions that are highly conserved at the species (Eucalyptus) and genus (with Arabidopsis and Populus) level, suggesting the presence of regulatory modules imposing functional constraint on such regions. Nucleotide polymorphisms in the CesA promoters more frequently created new cis-element occurrences than disrupted existing cis-element occurrences, a process which may be important for the maintenance and evolution of cellulose gene regulation in plants.Mondi and Sappi, through the Forest Molecular Genetics (FMG) Programme, the Technology and Human Resources for Industry Programme (THRIP)and the National Research Foundation of South Africa (NRF).http://link.springer.com/journal/11295hb201

Flower orientation influences floral temperature, pollinator visits and plant fitness

Effective insect pollination requires appropriate responses to internal and external environmental cues in both the plant and the pollinator. Helianthus annuus, a highly outcrossing species, is marked for its uniform eastward orientation of mature pseudanthia, or capitula. Here we investigate how this orientation affects floral microclimate and the consequent effects on plant and pollinator interactions and reproductive fitness. We artificially manipulated sunflower capitulum orientation and temperature in both field and controlled conditions and assessed flower physiology, pollinator visits, seed traits and siring success. East-facing capitula were found to have earlier style elongation, pollen presentation and pollinator visits compared with capitula manipulated to face west. East-facing capitula also sired more offspring than west-facing capitula and under some conditions produced heavier and better-filled seeds. Local ambient temperature change on the capitulum was found to be a key factor regulating the timing of style elongation, pollen emergence and pollinator visits. These results indicate that eastward capitulum orientation helps to control daily rhythms in floral temperature, with direct consequences on the timing of style elongation and pollen emergence, pollinator visitation, and plant fitness.University of Virginia; Division of Integrative Organismal Systems; University of California Berkeley; US Department of Agriculture-National Institute of Food and Agriculture.http://www.newphytologist.comhj2022Plant Production and Soil Scienc

Isolation and characterization of the cellulose synthase promoters of Eucalyptus trees

Cellulose is one of the most abundant biopolymers on earth and is an important commodity for industries such as the pulp and paper industry. Cellulose is deposited into the plant cell walls by a complex of membrane bound enzymes known as cellulose synthases. A number of cellulose synthase (CesA) genes, which encode for different cellulose synthase proteins, have been identified from plant species such as Eucalyptus, Populus and Arabidopsis. Mutant and expression profile analysis of the CesA genes indicated that a set of three CesA genes are associated with secondary cell wall formation, while a different set of CesA genes are associated with primary cell wall formation. The aim of this study was to investigate the transcriptional regulation of the different members of the CesA gene family in Eucalyptus. The promoter regions were comparatively analysed with the orthologous regions in Arabidopsis and Populus using bioinformatics tools to identify putative regulatory motifs that playa role in CesA genes regulation. Six Eucalyptus CesA gene promoters were isolated using genome walking. The Eucalyptus promoter regions and the orthologous promoter regions from Populus and Arabidopsis were analysed using TSSP (Transcriptional start site plant promoter prediction) and NNPP (Neural network promoter prediction) software packages. The software packages predicted the transcriptional start sites of the genes and the core regulatory elements such as the TATA-box and initiator elements. The in silico results were compared among species and it was found that the predicted transcriptional start sites and the core elements of the CesA gene promoters showed substantial structural conservation. The promoter regions were used in a comparative in silico analysis with the orthologous promoter regions from Arabidopsis and Populus to identify putative regulatory motifs. This is the first study in which the promoters of the CesA gene family are characterized in Arabidopsis, Populus and Eucalyptus. Three software packages (Weeder, POCO and MotifSampler) were used to analyse the promoter regions and identify over-represented motif sequences. A number of key stem-specific and xylem-specific motifs such as the AC-motif and G-box motif were identified as well as a number of novel motifs. Although all of the predicted motifs identified here will have to be functionally tested, the results of this study provide a good map for directed deletion studies and functional testing of the CesA promoters.Dissertation (MSc (Genetics))--University of Pretoria, 2009.Geneticsunrestricte

Comparative analysis of orthologous cellulose synthase promoters from Arabidopsis, Populus and Eucalyptus : evidence of conserved regulatory elements in angiosperms

The cellulose synthase (CesA) gene family encodes the catalytic subunits of a large protein complex responsible for the deposition of cellulose into plant cell walls. Early in vascular plant evolution, the gene family diverged into distinct members with conserved structures and functions (e.g. primary or secondary cell wall biosynthesis). Although the functions and expression domains of CesA genes have been extensively studied in plants, little is known about transcriptional regulation and promoter evolution in this gene family. Here, comparative sequence analysis of orthologous CesA promoters from three angiosperm genera, Arabidopsis, Populus and Eucalyptus, was performed to identify putative cis-regulatory sequences. The promoter sequences of groups of Arabidopsis genes that are co-expressed with the primary or secondary cell wall-related CesA genes were also analyzed. Reporter gene analysis of newly isolated promoter regions of six E. grandis CesA genes in Arabidopsis revealed the conserved functionality of the promoter sequences. Comparative sequence analysis identified 71 conserved sequence motifs, of which 66 were significantly over-represented in either primary or secondary wall-associated promoters. The presence of conserved cis-regulatory elements in the evolutionary distant CesA promoters of Arabidopsis, Populus and Eucalyptus suggests an ancient transcriptional network regulating cellulose biosynthesis in vascular plants