Transcriptional and post-transcriptional regulation of leaf development in Arabidopsis thaliana

Plant growth follows a strict developmental program but needs to incorporate also environmental cues to adapt to the encountered conditions. This requires a complex regulatory network to ensure an appropriate response to changing conditions. We used the first leaf pair of Arabidopsis thaliana as a model system to study the regulation of organ development. Leaf growth can be divided in subsequent phases according to the major process driving it. In a young leaf primordium cells divide continuously and cell size homeostasis is ensured by matching rates of cell expansion. Next, cell division ceases and cell expansion becomes the driving force for growth. When the leaf has attained its final size, maturity is reached. In this thesis, I studied the regulation of leaf development at two regulatory levels. At the gene level, we analyzed the function of the CYCA2 core cell cycle regulatory gene family. We also studied the function of two new proliferation specific gene families putatively involved in cell cycle regulation. On the other hand, we profiled small RNA sequences during development and linked this with the occurrence of DNA methylation. The core machinery of the cell cycle in plants has been thoroughly studied, but our knowledge on how developmental and environmental signals impinge on cell division is still limited. CYCA2s are known core cell cycle regulators, involved in G2-to-M transition. Here, we studied the functional requirement of this gene family and showed that transcriptional repression is required for specific differentiation processes. Members of the CYCA2 protein family function in vascular development and differentiation of guard cells. For the latter process, we demonstrated that FOUR LIPS and MYB88, two transcription factors involved in stomatal development, directly repress CYCA2;3 expression, thus ensuring correct guard cell differentiation. Next to known ‘core’ cell cycle regulating genes, we also selected proliferation specific genes with unknown function, assuming them to be involved in the cell division process. We focused on two small gene families: three genes with four transmembrane domains (4TMs) and two genes containing three High Mobility Group (HMG) domains (3xHMG-box). Expression analysis and localization of transcriptional fusions with a fluorescent marker confirmed for both gene families the highly proliferation-specific expression pattern. Moreover, both families are highly induced in the M-phase of the cell cycle in synchronized cell cultures. The 4TMs localize to the cell plate during mitosis and we observed defects in cell plate formax tion upon overexpression and depletion of these genes. Therefore, we hypothesize that the 4TM genes are involved in formation of the cell plate. Profiling of small RNAs (sRNAs) in plants has thusfar mainly been focused on inflorescence tissue or whole seedlings. Here, we studied sRNAs during the different phases of development. Early in development, microRNAs implicated in nutrient stress response are upregulated, suggesting that at this phase nutrient availability is limiting for growth. We showed that specifically 24-nt sRNAs increase in expression during development. This class of sRNAs is known to be involved in RNA-dependent DNA methylation (RdDM) and can thus silence both transposons and genes. In general, the expression of sRNAs matching the coding sequences of protein-coding genes is positively correlated to the mRNA expression of this gene. We specifically selected genes that do not show this correlation, which were highly enriched in two categories: targets of microRNAs and trans-acting siRNAs, which generate phased sRNAs upon cleavage, and genes for which the sRNA profile is enriched for 24-nt sRNAs. This latter category is likely regulated through RdDM as this subset of genes shows increased DNA methylation in the gene body. This suggests that sRNA regulation could play an important role in regulating the leaf developmental process not only by preserving genome integrity by repressing transposon activity but also through silencing of protein-coding genes

Developmental change in motor competence : a latent growth curve analysis

Background: The development of childhood motor competence demonstrates a high degree of inter-individual variation. Some children's competence levels increase whilst others' competence levels remain unchanged or even decrease over time. However, few studies have examined this developmental change in motor competence across childhood and little is known on influencing factors. Aim: Using latent growth curve modeling (LGCM), the present longitudinal study aimed to investigate children's change in motor competence across a 2-year timespan and to examine the potential influence of baseline weight status and physical fitness on their trajectory of change in motor competence. Methods: 558 children (52.5% boys) aged between 6 and 9 years participated in this study. Baseline measurements included weight status, motor competence (i.e., Korperkoordinationstest fur Kinder; KTK) and physical fitness (i.e., sit and reach, standing long jump and the 20 m shuttle run test). Motor competence assessment took place three times across a 2-year timespan. LGCM was conducted to examine change in motor competence over time. Results: The analyses showed a positive linear change in motor competence across 2 years (beta = 28.48, p < 0.001) with significant variability in children's individual trajectories (p < 0.001). Girls made less progress than boys (beta = -2.12, p = 0.01). Children who were older at baseline demonstrated less change in motor competence (beta = -0.33, p < 0.001). Weight status at baseline was negatively associated with change in motor competence over time (beta = -1.418, p = 0.002). None of the physical fitness components, measured at baseline, were significantly associated with change in motor competence over time. Conclusion and Implications: This longitudinal study reveals that weight status significantly influences children's motor competence trajectories whilst physical fitness demonstrated no significant influence on motor competence trajectories. Future studies should further explore children's differential trajectories over time and potential factors influencing that change

Reactivity of Gypsum-Based Materials Subjected to Thermal Load: Investigation of Reaction Mechanisms

The thermal stability of gypsum-based materials, and in this context, especially their long-term behavior, is the background of our current research activities. A comprehensive investigation program was compiled in which detailed examinations of various model materials exposed to thermal loads were carried out. The understanding of the partly not entirely consistent state of knowledge shall be sharpened especially by in situ observations of the thermally induced conversion reaction of gypsum into hemihydrate. The temporal course of the reaction was investigated non-destructively by in situ investigations in a high-resolution X-ray computed tomography setup, and the experiment was accompanied by detailed characterizations of the microstructure and composition. In this contribution, selected results of experiments with a high-purity natural gypsum rock as the model substance are presented. Studying the influence of temperature on the reaction showed that, even under supposedly dry conditions, the reaction could take place at much lower temperatures than usually reported in the literature. It was demonstrated that the transformation of gypsum into hemihydrate could take place at a temperature of already 50 °C. The results indicated that even under “classical” heating conditions in a conventional oven, the dissolution and crystallization processes in water films on the mineral surfaces could be suggested to be a driving force for the reaction. A corresponding reaction model, which took these aspects into account, was proposed in this work

The TriForC database : a comprehensive up-to-date resource of plant triterpene biosynthesis

Triterpenes constitute a large and important class of plant natural products with diverse structures and functions. Their biological roles range from membrane structural components over plant hormones to specialized plant defence compounds. Furthermore, triterpenes have great potential for a variety of commercial applications such as vaccine adjuvants, anti-cancer drugs, food supplements and agronomic agents. Their biosynthesis is carried out through complicated, branched pathways bymultiple enzyme types that include oxidosqualene cyclases, cytochrome P450s, and UDP-glycosyltransferases. Given that the number of characterized triterpene biosynthesis enzymes has been growing fast recently, the need for a database specifically focusing on triterpene enzymology became eminent. Here, we present the TriForC database (http://bioinformatics. psb. ugent. be/triforc/), encompassing a comprehensive catalogue of triterpene biosynthesis enzymes. This highly interlinked database serves as a user-friendly access point to versatile data sets of enzyme and compound features, enabling the scanning of a complete catalogue of experimentally validated triterpene enzymes, their substrates and products, as well as the pathways they constitute in various plant species. The database can be accessed by direct browsing or through convenient search tools including keyword, BLAST, plant species and substructure options. This database will facilitate gene mining and creating genetic toolboxes for triterpene synthetic biology

PLAZA 4.0 : an integrative resource for functional, evolutionary and comparative plant genomics

PLAZA (https://bioinformatics.psb.ugent.be/plaza) is a plant-oriented online resource for comparative, evolutionary and functional genomics. The PLAZA platform consists of multiple independent instances focusing on different plant clades, while also providing access to a consistent set of reference species. Each PLAZA instance contains structural and functional gene annotations, gene family data and phylogenetic trees and detailed gene colinearity information. A user-friendly web interface makes the necessary tools and visualizations accessible, specific for each data type. Here we present PLAZA 4.0, the latest iteration of the PLAZA framework. This version consists of two new instances (Dicots 4.0 and Monocots 4.0) providing a large increase in newly available species, and offers access to updated and newly implemented tools and visualizations, helping users with the ever-increasing demands for complex and in-depth analyzes. The total number of species across both instances nearly doubles from 37 species in PLAZA 3.0 to 71 species in PLAZA 4.0, with a much broader coverage of crop species (e.g. wheat, palm oil) and species of evolutionary interest (e.g. spruce, Marchantia). The new PLAZA instances can also be accessed by a programming interface through a RESTful web service, thus allowing bioinformaticians to optimally leverage the power of the PLAZA platform

Correlation analysis of the transcriptome of growing leaves with mature leaf parameters in a maize RIL population

Background: To sustain the global requirements for food and renewable resources, unraveling the molecular networks underlying plant growth is becoming pivotal. Although several approaches to identify genes and networks involved in final organ size have been proven successful, our understanding remains fragmentary. Results: Here, we assessed variation in 103 lines of the Zea mays B73xH99 RIL population for a set of final leaf size and whole shoot traits at the seedling stage, complemented with measurements capturing growth dynamics, and cellular measurements. Most traits correlated well with the size of the division zone, implying that the molecular basis of final leaf size is already defined in dividing cells of growing leaves. Therefore, we searched for association between the transcriptional variation in dividing cells of the growing leaf and final leaf size and seedling biomass, allowing us to identify genes and processes correlated with the specific traits. A number of these genes have a known function in leaf development. Additionally, we illustrated that two independent mechanisms contribute to final leaf size, maximal growth rate and the duration of growth. Conclusions: Untangling complex traits such as leaf size by applying in-depth phenotyping allows us to define the relative contributions of the components and their mutual associations, facilitating dissection of the biological processes and regulatory networks underneath

Galaxy: A Decade of Realising CWFR Concepts

Despite recent encouragement to follow the FAIR principles, the day-to-day research practices have not changed substantially. Due to new developments and the increasing pressure to apply best practices, initiatives to improve the efficiency and reproducibility of scientific workflows are becoming more prevalent. In this article, we discuss the importance of well-annotated tools and the specific requirements to ensure reproducible research with FAIR outputs. We detail how Galaxy, an open-source workflow management system with a web-based interface, has implemented the concepts that are put forward by the Canonical Workflow Framework for Research (CWFR), whilst minimising changes to the practices of scientific communities. Although we showcase concrete applications from two different domains, this approach is generalisable to any domain and particularly useful in interdisciplinary research and science-based applications.publishedVersio

From Invisibility to Readability: Recovering the Ink of Herculaneum

The noninvasive digital restoration of ancient texts written in carbon black ink and hidden inside artifacts has proven elusive, even with advanced imaging techniques like x-ray-based micro-computed tomography (micro-CT). This paper identifies a crucial mistaken assumption: that micro-CT data fails to capture any information representing the presence of carbon ink. Instead, we show new experiments indicating a subtle but detectable signature from carbon ink in micro-CT. We demonstrate a new computational approach that captures, enhances, and makes visible the characteristic signature created by carbon ink in micro-CT. This previously unseen evidence of carbon inks, which can now successfully be made visible, is a discovery that can lead directly to the noninvasive digital recovery of the lost texts of Herculaneum

Proteome Profiling of Wheat Shoots from Different Cultivars

Wheat is a cereal grain and one of the world's major food crops. Recent advances in wheat genome sequencing are by now facilitating its genomic and proteomic analyses. However, little is known about possible differences in total protein levels of hexaploid versus tetraploid wheat cultivars, and also knowledge of phosphorylated wheat proteins is still limited. Here, we performed a detailed analysis of the proteome of seedling leaves from two hexaploid wheat cultivars (Triticum aestivum L. Pavon 76 and USU-Apogee) and one tetraploid wheat (T. turgidum ssp. durum cv. Senatore Cappelli). Our shotgun proteomics data revealed that, whereas we observed some significant differences, overall a high similarity between hexaploid and tetraploid varieties with respect to protein abundance was observed. In addition, already at the seedling stage, a small set of proteins was differential between the small (USU-Apogee) and larger hexaploid wheat cultivars (Pavon 76), which could potentially act as growth predictors. Finally, the phosphosites identified in this study can be retrieved from the in-house developed plant PTM-Viewer (bioinformatics.psb.ugent.be/webtools/ptm_viewer/), making this the first searchable repository for phosphorylated wheat proteins. This paves the way for further in depth, quantitative (phospho) proteome-wide differential analyses upon a specific trigger or environmental change

A SARS-CoV-2 sequence submission tool for the European Nucleotide Archive

Abstract Summary Many aspects of the global response to the COVID-19 pandemic are enabled by the fast and open publication of SARS-CoV-2 genetic sequence data. The European Nucleotide Archive (ENA) is the European recommended open repository for genetic sequences. In this work, we present a tool for submitting raw sequencing reads of SARS-CoV-2 to ENA. The tool features a single-step submission process, a graphical user interface, tabular-formatted metadata and the possibility to remove human reads prior to submission. A Galaxy wrap of the tool allows users with little or no bioinformatic knowledge to do bulk sequencing read submissions. The tool is also packed in a Docker container to ease deployment. Availability CLI ENA upload tool is available at github.com/usegalaxy-eu/ena-upload-cli (DOI 10.5281/zenodo.4537621); Galaxy ENA upload tool at toolshed.g2.bx.psu.edu/view/iuc/ena_upload/382518f24d6d and https://github.com/galaxyproject/tools-iuc/tree/master/tools/ena_upload (development) and; ENA upload Galaxy container at github.com/ELIXIR-Belgium/ena-upload-container (DOI 10.5281/zenodo.4730785) </jats:sec