Teaching activities:

My main teaching activities relate to the theoretical and practical understanding of plant cell biology and plant development - mainly focusing on stem-cell dynamics, cell differentiation, cytolosolic components (membrane trafficking, cytoskeleton,..) and high-throughput analyses (transcriptomics, proteomics, metabolomics). My teaching is made at all levels from Bachelor to PhD students.

(1) Developing novel advanced methods for imaging biomolecules

The "Cell differentiation and coordination in tissues" group is establishing and optimizing novel methods to directly assess changes in amount and chemical composition of plant biomass from biopsies with sub µm resolution. These techniques called “in situ quantitative chemical imaging” thus enable to directly evaluate changes in plant biomass levels and composition without having to grid and destroy the cellular and tissular organization of plants.       

Recent breakthroughs in in situ quantitative chemical imaging from our research group include:

• Solving the Wiesner test, a 150-year old mystery for one of the most used histochemical staining in Plant Biology [https://doi.org/10.3389/fpls.2020.00109]
• Using UV-excited autofluorescence to define changes in distribution and composition of plant biomass [https://doi.org/10.1007/978-1-4939-6722-3_17]
• - Establishing Raman spectroscopy for quantitative analysis of changes in the chemistry of plant biomass [https://doi.org/10.1021/acssuschemeng.0c00194]

(2) Understanding cell type specific formation and function

The "Cell differentiation and coordination in tissues" group investigates the molecular and cellular processes enabling specific cell types to form in plant tissues. We have thus developed unique technology using inducible pluripotent cell suspension cultures (iPSCs) to produce on-demand the plant cell type that we want to study and can thus follow its formation during time.               

Recent breakthroughs in inducible pluripotent cell suspension cultures (iPSCs) from our research group include:

• Establishing iPSCs technology for plants [https://doi.org/10.1007/978-1-4939-6722-3_4]
• Technology available using plant cell cultures [https://doi.org/10.1016/j.copbio.2019.02.001]    

Recent breakthroughs in understanding wood cell type formation from our research group include:

• How is biomass composition linked to cellular function for wood cells [https://doi.org/10.1093/plcell/koac284]
• How is cell wall organization linked to cellular function for wood cells [https://doi.org/10.1016/j.cub.2010.02.057]

(3) Understanding plant biomass formation

The "Cell differentiation and coordination in tissues" group investigates the formation and organization of the plant carbon biomass that is accumulated in plant cell walls. We mainly focus on the molecular mechanisms controlling changes in cell wall composition (mostly lignin) and organization (mostly through microtubule guidance).

Recent breakthroughs in identifying the molecular mechanism controlling cell wall composition from our research group include:

• Modelling laccase paralogs for differential activity in plants and other organisms  [https://doi.org/10.3389/fpls.2021.754601]
• Functional studies of laccase paralog combinations in controlling cell wall composition between cell types [https://doi.org/10.1093/plcell/koac344]

Recent breakthroughs in identifying the molecular mechanism controlling cell wall organization from our research group include:

• Understanding of the quantitative changes in proteins restructuring inner cellular skeleton to guide cell wall organization [https://doi.org/10.1105/tpc.15.00314]
• Functional studies of specific proteins restructuring inner cellular skeleton to guide cell wall organization [https://doi.org/10.1016/j.cub.2010.02.057]

I urval från Stockholms universitets publikationsdatabas

• Dynamic incorporation of specific lignin residues controls the biomechanics of the plant vasculature and its resilience to environmental changes

  Delphine Ménard (et al.).

   The accumulation of the cell wall polymer lignin in vascular cells enables long-distance water conduction and structural support in plants. Independently of the plant species, each different vascular cell type accumulates specific lignin amount and composition affecting both aromatic and aliphatic substitutions of its residues. However, the biological role of this conserved and specific lignin chemistry for each cell type remains unclear. Herein, we performed single cell analyses on plant vascular cell morphotypes to investigate the role of specific lignin composition for cellular function. We showed that distinct amounts and compositions of lignin accumulated in the different morphotypes of the sap conducting vascular cells. We discovered that lignin accumulates dynamically, increasing in quantity and changing composition, to fine-tune the cell wall mechanical properties of each conducting cell morphotype. Modification this lignin specificity impaired specifically the cell wall mechanical properties of each morphotype and consequently their capacity to optimally conduct water in normal but also to recover from drought conditions. Altogether, our findings provide the biological role of specific lignin chemistry in sap conducting cells, to dynamically adjust the hydraulic properties of each conducting cell during developmental and environmental constraints.

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• Laccase paralogs non-redundantly direct the lignin amount and composition of specific cell wall layers and cell types in Arabidopsis

  Leonard Blaschek (et al.).

  Läs mer om Laccase paralogs non-redundantly direct the lignin amount and composition of specific cell wall layers and cell types in Arabidopsis

• Overexpression of EgrIAA20 from Eucalyptus grandis, a Non-Canonical Aux/IAA Gene, Specifically Decouples Lignification of the Different Cell-Types in Arabidopsis Secondary Xylem

  2022. Hong Yu (et al.). International Journal of Molecular Sciences 23 (9)

  Artikel

  Wood (secondary xylem) formation is regulated by auxin, which plays a pivotal role as an integrator of developmental and environmental cues. However, our current knowledge of auxin-signaling during wood formation is incomplete. Our previous genome-wide analysis of Aux/IAAs in Eucalyptus grandis showed the presence of the non-canonical paralog member EgrIAA20 that is preferentially expressed in cambium. We analyzed its cellular localization using a GFP fusion protein and its transcriptional activity using transactivation assays, and demonstrated its nuclear localization and strong auxin response repressor activity. In addition, we functionally tested the role of EgrIAA20 by constitutive overexpression in Arabidopsis to investigate for phenotypic changes in secondary xylem formation. Transgenic Arabidopsis plants overexpressing EgrIAA20 were smaller and displayed impaired development of secondary fibers, but not of other wood cell types. The inhibition in fiber development specifically affected their cell wall lignification. We performed yeast-two-hybrid assays to identify EgrIAA20 protein partners during wood formation in Eucalyptus, and identified EgrIAA9A, whose ortholog PtoIAA9 in poplar is also known to be involved in wood formation. Altogether, we showed that EgrIAA20 is an important auxin signaling component specifically involved in controlling the lignification of wood fibers.

  Läs mer om Overexpression of EgrIAA20 from Eucalyptus grandis, a Non-Canonical Aux/IAA Gene, Specifically Decouples Lignification of the Different Cell-Types in Arabidopsis Secondary Xylem

• Phenoloxidases in Plants-How Structural Diversity Enables Functional Specificity

  2021. Leonard Blaschek, Edouard Pesquet. Frontiers in Plant Science 12

  Artikel

  The metabolism of polyphenolic polymers is essential to the development and response to environmental changes of organisms from all kingdoms of life, but shows particular diversity in plants. In contrast to other biopolymers, whose polymerisation is catalysed by homologous gene families, polyphenolic metabolism depends on phenoloxidases, a group of heterogeneous oxidases that share little beyond the eponymous common substrate. In this review, we provide an overview of the differences and similarities between phenoloxidases in their protein structure, reaction mechanism, substrate specificity, and functional roles. Using the example of laccases (LACs), we also performed a meta-analysis of enzyme kinetics, a comprehensive phylogenetic analysis and machine-learning based protein structure modelling to link functions, evolution, and structures in this group of phenoloxidases. With these approaches, we generated a framework to explain the reported functional differences between paralogs, while also hinting at the likely diversity of yet undescribed LAC functions. Altogether, this review provides a basis to better understand the functional overlaps and specificities between and within the three major families of phenoloxidases, their evolutionary trajectories, and their importance for plant primary and secondary metabolism.

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• Cellular and Genetic Regulation of Coniferaldehyde Incorporation in Lignin of Herbaceous and Woody Plants by Quantitative Wiesner Staining

  2020. Leonard Blaschek (et al.). Frontiers in Plant Science 11

  Artikel

  Lignin accumulates in the cell walls of specialized cell types to enable plants to stand upright and conduct water and minerals, withstand abiotic stresses, and defend themselves against pathogens. These functions depend on specific lignin concentrations and subunit composition in different cell types and cell wall layers. However, the mechanisms controlling the accumulation of specific lignin subunits, such as coniferaldehyde, during the development of these different cell types are still poorly understood. We herein validated the Wiesner test (phloroglucinol/HCl) for the restrictive quantitative in situ analysis of coniferaldehyde incorporation in lignin. Using this optimized tool, we investigated the genetic control of coniferaldehyde incorporation in the different cell types of genetically-engineered herbaceous and woody plants with modified lignin content and/or composition. Our results demonstrate that the incorporation of coniferaldehyde in lignified cells is controlled by (a) autonomous biosynthetic routes for each cell type, combined with (b) distinct cell-to-cell cooperation between specific cell types, and (c) cell wall layer-specific accumulation capacity. This process tightly regulates coniferaldehyde residue accumulation in specific cell types to adapt their property and/or function to developmental and/or environmental changes.

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• Determining the Genetic Regulation and Coordination of Lignification in Stem Tissues of Arabidopsis Using Semiquantitative Raman Microspectroscopy

  2020. Leonard Blaschek (et al.). ACS Sustainable Chemistry and Engineering 8 (12), 4900-4909

  Artikel

  Lignin is a phenolic polymer accumulatig in the cell walls of specific plant cell types to confer unique properties such as hydrophobicity, mechanical strengthening, and resistance to degradation. Different cell types accumulate lignin with specific concentration and composition to support their specific roles in the different plant tissues. Yet the genetic mechanisms controlling lignin quantity and composition differently between the different lignified cell types and tissues still remain poorly understood. To investigate this tissue-specific genetic regulation, we validated both the target molecular structures as well as the linear semi-quantitative capacity of Raman microspectroscopy to characterize the total lignin amount, S/G ratio, and coniferyl alcohol content in situ directly in plant biopsies. Using the optimized method on stems of multiple lignin biosynthesis loss-of-function mutants revealed that the genetic regulation of lignin is tissue specific, with distinct genes establishing nonredundant check-points to trigger specific compensatory adjustments affecting either lignin composition and/or cell wall polymer concentrations.

  Läs mer om Determining the Genetic Regulation and Coordination of Lignification in Stem Tissues of Arabidopsis Using Semiquantitative Raman Microspectroscopy

• Importance of Lignin Coniferaldehyde Residues for Plant Properties and Sustainable Uses

  2020. Masanobu Yamamoto (et al.). ChemSusChem 13 (17), 4400-4408

  Artikel

  Increases in coniferaldehyde content, a minor lignin residue, significantly improves the sustainable use of plant biomass for feed, pulping, and biorefinery without affecting plant growth and yields. Herein, different analytical methods are compared and validated to distinguish coniferaldehyde from other lignin residues. It is shown that specific genetic pathways regulate amount, linkage, and position of coniferaldehyde within the lignin polymer for each cell type. This specific cellular regulation offers new possibilities for designing plant lignin for novel and targeted industrial uses.

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• Cell culture systems: invaluable tools to investigate lignin formation and cell wall properties

  2019. Edouard Pesquet, Armin Wagner, John H. Grabber. Current Opinion in Biotechnology 56, 215-222

  Artikel

  Although the use of cell culture systems in Plant Biology and Biotechnology has been limited compared to other areas of Life Sciences, plant cell cultures capable of lignifying on demand have proven invaluable in unravelling the lignification process and its impact on biomass utilization. Inducible cell cultures have enabled researchers to decipher multiple levels of cellular control used in and between plant cells to define the spatiotemporal deposition, composition, structure, and quantity of lignin. Artificially lignified cell cultures have also been used to determine the effects of lignin composition on the susceptibility of cell walls to chemical treatments, and digestion by rumen microflora or fungal enzymes. Plant cell cultures have enabled the fast-tracking of lignin-related research and provided insights into the lignification processes that could not have been easily obtained by using whole plants as model systems.

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• Establishment of Photosynthesis through Chloroplast Development Is Controlled by Two Distinct Regulatory Phases

  2018. Carole Dubreuil (et al.). Plant Physiology 176 (2), 1199-1214

  Artikel

  Chloroplasts develop from undifferentiated proplastids present in meristematic tissue. Thus, chloroplast biogenesis is closely connected to leaf development, which restricts our ability to study the process of chloroplast biogenesis per se. As a consequence, we know relatively little about the regulatory mechanisms behind the establishment of the photosynthetic reactions and how the activities of the two genomes involved are coordinated during chloroplast development. We developed a single cell-based experimental system from Arabidopsis (Arabidopsis thaliana) with high temporal resolution allowing for investigations of the transition from proplastids to functional chloroplasts. Using this unique cell line, we could show that the establishment of photosynthesis is dependent on a regulatory mechanism involving two distinct phases. The first phase is triggered by rapid light-induced changes in gene expression and the metabolome. The second phase is dependent on the activation of the chloroplast and generates massive changes in the nuclear gene expression required for the transition to photosynthetically functional chloroplasts. The second phase also is associated with a spatial transition of the chloroplasts from clusters around the nucleus to the final position at the cell cortex. Thus, the establishment of photosynthesis is a two-phase process with a clear checkpoint associated with the second regulatory phase allowing coordination of the activities of the nuclear and plastid genomes.

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• AspWood: High-Spatial-Resolution Transcriptome Profiles Reveal Uncharacterized Modularity of Wood Formation in Populus tremula

  2017. David Sundell (et al.). The Plant Cell 29 (7), 1585-1604

  Artikel

  Trees represent the largest terrestrial carbon sink and a renewable source of ligno-cellulose. There is significant scope for yield and quality improvement in these largely undomesticated species, and efforts to engineer elite varieties will benefit from improved understanding of the transcriptional network underlying cambial growth and wood formation. We generated high-spatial-resolution RNA sequencing data spanning the secondary phloem, vascular cambium, and wood-forming tissues of Populus tremula. The transcriptome comprised 28,294 expressed, annotated genes, 78 novel protein-coding genes, and 567 putative long intergenic noncoding RNAs. Most paralogs originating from the Salicaceae whole-genome duplication had diverged expression, with the exception of those highly expressed during secondary cell wall deposition. Coexpression network analyses revealed that regulation of the transcriptome underlying cambial growth and wood formation comprises numerous modules forming a continuum of active processes across the tissues. A comparative analysis revealed that a majority of these modules are conserved in Picea abies. The high spatial resolution of our data enabled identification of novel roles for characterized genes involved in xylan and cellulose biosynthesis, regulators of xylem vessel and fiber differentiation and lignification. An associated web resource (AspWood, http://aspwood.popgenie.org) provides interactive tools for exploring the expression profiles and coexpression network.

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• Two Complementary Mechanisms Underpin Cell Wall Patterning during Xylem Vessel Development

  2017. Rene Schneider (et al.). The Plant Cell 29 (10), 2433-2449

  Artikel

  The evolution of the plant vasculature was essential for the emergence of terrestrial life. Xylem vessels are solute-transporting elements in the vasculature that possess secondary wall thickenings deposited in intricate patterns. Evenly dispersed microtubule (MT) bands support the formation of these wall thickenings, but how the MTs direct cell wall synthesis during this process remains largely unknown. Cellulose is the major secondary wall constituent and is synthesized by plasma membrane-localized cellulose synthases (CesAs) whose catalytic activity propels them through the membrane. We show that the protein CELLULOSE SYNTHASE INTERACTING1 (CSI1)/POM2 is necessary to align the secondary wall CesAs and MTs during the initial phase of xylem vessel development in Arabidopsis thaliana and rice (Oryza sativa). Surprisingly, these MT-driven patterns successively become imprinted and sufficient to sustain the continued progression of wall thickening in the absence of MTs and CSI1/POM2 function. Hence, two complementary principles underpin wall patterning during xylem vessel development.

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• Chemical Genetics Uncovers Novel Inhibitors of Lignification, Including p-Iodobenzoic Acid Targeting CINNAMATE-4-HYDROXYLASE

  2016. Dorien Van de Wouwer (et al.). Plant Physiology 172 (1), 198-220

  Artikel

  Plant secondary-thickened cell walls are characterized by the presence of lignin, a recalcitrant and hydrophobic polymer that provides mechanical strength and ensures long-distance water transport. Exactly the recalcitrance and hydrophobicity of lignin put a burden on the industrial processing efficiency of lignocellulosic biomass. Both forward and reverse genetic strategies have been used intensively to unravel the molecular mechanism of lignin deposition. As an alternative strategy, we introduce here a forward chemical genetic approach to find candidate inhibitors of lignification. A high-throughput assay to assess lignification in Arabidopsis (Arabidopsis thaliana) seedlings was developed and used to screen a 10-k library of structurally diverse, synthetic molecules. Of the 73 compounds that reduced lignin deposition, 39 that had a major impact were retained and classified into five clusters based on the shift they induced in the phenolic profile of Arabidopsis seedlings. One representative compound of each cluster was selected for further lignin-specific assays, leading to the identification of an aromatic compound that is processed in the plant into two fragments, both having inhibitory activity against lignification. One fragment, p-iodobenzoic acid, was further characterized as a new inhibitor of CINNAMATE 4-HYDROXYLASE, a key enzyme of the phenylpropanoid pathway synthesizing the building blocks of the lignin polymer. As such, we provide proof of concept of this chemical biology approach to screen for inhibitors of lignification and present a broad array of putative inhibitors of lignin deposition for further characterization.

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