| 1. |
- Andersson, Mariette, et al.
(author)
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An alternative pathway to plant cold tolerance in the absence of vacuolar invertase activity
- 2023
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In: Plant Journal. - : Wiley. - 0960-7412 .- 1365-313X. ; 113, s. 327-341
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Journal article (peer-reviewed)abstract
- To cope with cold stress, plants have developed antioxidation strategies combined with osmoprotection by sugars. In potato (Solanum tuberosum) tubers, which are swollen stems, exposure to cold stress induces starch degradation and sucrose synthesis. Vacuolar acid invertase (VInv) activity is a significant part of the cold-induced sweetening (CIS) response, by rapidly cleaving sucrose into hexoses and increasing osmoprotection. To discover alternative plant tissue pathways for coping with cold stress, we produced VInv-knockout lines in two cultivars. Genome editing of VInv in 'Desiree' and 'Brooke' was done using stable and transient expression of CRISPR/Cas9 components, respectively. After storage at 4 degrees C, sugar analysis indicated that the knockout lines showed low levels of CIS and maintained low acid invertase activity in storage. Surprisingly, the tuber parenchyma of vinv lines exhibited significantly reduced lipid peroxidation and reduced H2O2 levels. Furthermore, whole plants of vinv lines exposed to cold stress without irrigation showed normal vigor, in contrast to WT plants, which wilted. Transcriptome analysis of vinv lines revealed upregulation of an osmoprotectant pathway and ethylene-related genes during cold temperature exposure. Accordingly, higher expression of antioxidant-related genes was detected after exposure to short and long cold storage. Sugar measurements showed an elevation of an alternative pathway in the absence of VInv activity, raising the raffinose pathway with increasing levels of myo-inositol content as a cold tolerance response.
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| 2. |
- Antoniadi, Ioanna, et al.
(author)
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Fluorescence-activated multi-organelle mapping of subcellular plant hormone distribution
- 2023
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In: Plant Journal. - 0960-7412 .- 1365-313X. ; 116, s. 1825-1841
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Journal article (peer-reviewed)abstract
- Auxins and cytokinins are two major families of phytohormones that control most aspects of plant growth, development and plasticity. Their distribution in plants has been described, but the importance of cell- and subcellular-type specific phytohormone homeostasis remains undefined. Herein, we revealed auxin and cytokinin distribution maps showing their different organelle-specific allocations within the Arabidopsis plant cell. To do so, we have developed Fluorescence-Activated multi-Organelle Sorting (FAmOS), an innovative subcellular fractionation technique based on flow cytometric principles. FAmOS allows the simultaneous sorting of four differently labelled organelles based on their individual light scatter and fluorescence parameters while ensuring hormone metabolic stability. Our data showed different subcellular distribution of auxin and cytokinins, revealing the formation of phytohormone gradients that have been suggested by the subcellular localization of auxin and cytokinin transporters, receptors and metabolic enzymes. Both hormones showed enrichment in vacuoles, while cytokinins were also accumulated in the endoplasmic reticulum.
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| 3. |
- Bag, Pushan, 1993-, et al.
(author)
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An atlas of the Norway spruce needle seasonal transcriptome
- 2021
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In: The Plant Journal. - : John Wiley & Sons. - 0960-7412 .- 1365-313X. ; 108:6, s. 1815-1829
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Journal article (peer-reviewed)abstract
- Boreal conifers possess a tremendous ability to survive and remain evergreen during harsh winter conditions and resume growth during summer. This is enabled by coordinated regulation of major cellular functions at the level of gene expression, metabolism, and physiology. Here we present a comprehensive characterization of the annual changes in the global transcriptome of Norway spruce (Picea abies) needles as a resource to understand needle development and acclimation processes throughout the year. In young, growing needles (May 15 until June 30), cell walls, organelles, etc., were formed, and this developmental program heavily influenced the transcriptome, explained by over-represented Gene Ontology (GO) categories. Later changes in gene expression were smaller but four phases were recognized: summer (July–August), autumn (September–October), winter (November–February), and spring (March–April), where over-represented GO categories demonstrated how the needles acclimated to the various seasons. Changes in the seasonal global transcriptome profile were accompanied by differential expression of members of the major transcription factor families. We present a tentative model of how cellular activities are regulated over the year in needles of Norway spruce, which demonstrates the value of mining this dataset, accessible in ConGenIE together with advanced visualization tools.
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| 4. |
- Barcala, Maximiliano Estravis, et al.
(author)
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Whole-genome resequencing facilitates the development of a 50K single nucleotide polymorphism genotyping array for Scots pine (Pinus sylvestris L.) and its transferability to other pine species
- 2024
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In: The Plant Journal. - 0960-7412 .- 1365-313X. ; 117:3, s. 944-955
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Journal article (peer-reviewed)abstract
- Scots pine (Pinus sylvestris L.) is one of the most widespread and economically important conifer species in the world. Applications like genomic selection and association studies, which could help accelerate breeding cycles, are challenging in Scots pine because of its large and repetitive genome. For this reason, genotyping tools for conifer species, and in particular for Scots pine, are commonly based on transcribed regions of the genome. In this article, we present the Axiom Psyl50K array, the first single nucleotide polymorphism (SNP) genotyping array for Scots pine based on whole-genome resequencing, that represents both genic and intergenic regions. This array was designed following a two-step procedure: first, 192 trees were sequenced, and a 430K SNP screening array was constructed. Then, 480 samples, including haploid megagametophytes, full-sib family trios, breeding population, and range-wide individuals from across Eurasia were genotyped with the screening array. The best 50K SNPs were selected based on quality, replicability, distribution across the draft genome assembly, balance between genic and intergenic regions, and genotype–environment and genotype–phenotype associations. Of the final 49 877 probes tiled in the array, 20 372 (40.84%) occur inside gene models, while the rest lie in intergenic regions. We also show that the Psyl50K array can yield enough high-confidence SNPs for genetic studies in pine species from North America and Eurasia. This new genotyping tool will be a valuable resource for high-throughput fundamental and applied research of Scots pine and other pine species.
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| 5. |
- Bourras, Salim
(author)
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Single residues in the LRR domain of the wheat PM3A immune receptor can control the strength and the spectrum of the immune response
- 2020
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In: Plant Journal. - : Wiley. - 0960-7412 .- 1365-313X. ; 104, s. 200-214
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Journal article (peer-reviewed)abstract
- The development of improved plant nucleotide-binding, leucine-rich repeat (LRR) immune receptors (NLRs) has mostly been based on random mutagenesis or on structural information available for specific receptors complexed with the recognized pathogen effector. Here, we use a targeted mutagenesis approach based on the natural diversity of thePm3powdery mildew resistance alleles present in different wheat (Triticum aestivum) genotypes. In order to understand the functional importance of the amino acid polymorphisms between the active immune receptor PM3A and the inactive ancestral variant PM3CS, we exchanged polymorphic regions and residues in the LRR domain of PM3A with the corresponding segments of PM3CS. These novel variants were functionally tested for recognition of the corresponding AVRPM3(A2/F2)avirulence protein inNicotiana benthamiana. We identified polymorphic residues in four regions of PM3A that enhance the immune response, but also residues that reduce it or result in complete loss of function. We found that the identified critical residues in PM3A modify its activation threshold towards different protein variants of AVRPM3(A2/F2). PM3A variants with a lowered threshold gave a stronger overall response and gained an extended recognition spectrum. One of these variant proteins with a single amino acid change was stably transformed into wheat, where it conferred race-specific resistance to mildew. This is a proof of concept that improved PM3A variants with an enlarged recognition spectrum can be engineered based on natural diversity by exchanging single or multiple residues that modulate resistance function.
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| 6. |
- Boussardon, Clément, et al.
(author)
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Tissue-specific isolation of Arabidopsis/plant mitochondria - IMTACT (isolation of mitochondria tagged in specific cell types)
- 2020
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In: The Plant Journal. - : John Wiley & Sons. - 0960-7412 .- 1365-313X. ; 103:1, s. 459-473
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Journal article (peer-reviewed)abstract
- Plant cells contain numerous subcompartments with clearly delineated metabolic functions. Mitochondria represent a very small fraction of the total cell volume and yet are the site of respiration and thus crucial for cells throughout all developmental stages of a plant's life. As such, their isolation from the rest of the cellular components is a basic requirement for numerous biochemical and physiological experiments. Although procedures exist to isolate plant mitochondria from different organs (i.e. leaves, roots, tubers, etc.), they are often tedious and do not provide resolution at the tissue level (i.e. phloem, mesophyll or pollen). Here, we present a novel method called IMTACT (isolation of mitochondria tagged in specific cell types), developed inArabidopsis thaliana(Arabidopsis) that involves biotinylation of mitochondria in a tissue-specific manner using transgenic lines expressing a synthetic version of theOM64(Outer Membrane 64) gene combined withBLRPand theBirAbiotin ligase gene. Tissue specificity is achieved with cell-specific promoters (e.g.CAB3andSUC2). Labeled mitochondria from crude extracts are retained by magnetic beads, allowing the simple and rapid isolation of highly pure and intact organelles from organs or specific tissues. For example, we could show that the mitochondrial population from mesophyll cells was significantly larger in size than the mitochondrial population isolated from leaf companion cells. To facilitate the applicability of this method in both wild-type and mutant Arabidopsis plants we generated a set of OM64-BLRP one-shot constructs with different selection markers and tissue-specific promoters.
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| 7. |
- Brouwer, Sophie, et al.
(author)
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Visualising the ionome in resistant and susceptible plant-pathogen interactions
- 2021
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In: Plant Journal. - : Wiley. - 0960-7412 .- 1365-313X. ; 108, s. 870-885
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Journal article (peer-reviewed)abstract
- At the morphological and anatomical levels, the ionome, or the elemental composition of an organism, is an understudied area of plant biology. In particular, the ionomic responses of plant-pathogen interactions are scarcely described, and there are no studies on immune reactions. In this study we explored two X-ray fluorescence (XRF)-based ionome visualisation methods (benchtop- and synchrotron-based micro-XRF [mu XRF]), as well as the quantitative inductively coupled plasma optical emission spectroscopy (ICP-OES) method, to investigate the changes that occur in the ionome of compatible and incompatible plant-pathogen interactions. We utilised the agronomically important and comprehensively studied interaction between potato (Solanum tuberosum) and the late blight oomycete pathogen Phytophthora infestans as an example. We used one late blight-susceptible potato cultivar and two resistant transgenic plant lines (only differing from the susceptible cultivar in one or three resistance genes) both in control and P. infestans-inoculated conditions. In the lesions from the compatible interaction, we observed rearrangements of several elements, including a decrease of the mobile macronutrient potassium (K) and an increase in iron (Fe) and manganese (Mn), compared with the tissue outside the lesion. Interestingly, we observed distinctly different distribution patterns of accumulation at the site of inoculation in the resistant lines for calcium (Ca), magnesium (Mg), Mn and silicon (Si) compared to the susceptible cultivar. The results reveal different ionomes in diseased plants compared to resistant plants. Our results demonstrate a technical advance and pave the way for deeper studies of the plant-pathogen ionome in the future.
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| 8. |
- Bünder, Anne, et al.
(author)
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CELLULOSE SYNTHASE INTERACTING 1 is required for wood mechanics and leaf morphology in aspen
- 2020
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In: The Plant Journal. - : John Wiley & Sons. - 0960-7412 .- 1365-313X. ; 103:5, s. 1858-1868
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Journal article (peer-reviewed)abstract
- Cellulose microfibrils synthesized by CELLULOSE SYNTHASE COMPLEXES (CSCs) are the main load‐bearing polymers in wood. CELLULOSE SYNTHASE INTERACTING1 (CSI1) connects CSCs with cortical microtubules, which align with cellulose microfibrils. Mechanical properties of wood are dependent on cellulose microfibril alignment and structure in the cell walls, but the molecular mechanism(s) defining these features is unknown. Herein, we investigated the role of CSI1 in hybrid aspen (Populus tremula × Populus tremuloides ) by characterizing transgenic lines with significantly reduced CSI1 transcript abundance. Reduction in leaves (50–80%) caused leaf twisting and misshaped pavement cells, while reduction (70–90%) in developing xylem led to impaired mechanical wood properties evident as a decrease in the elastic modulus and rupture. X‐ray diffraction measurements indicate that microfibril angle was not impacted by the altered CSI1 abundance in developing wood fibres. Instead, the augmented wood phenotype of the transgenic trees was associated with a reduced cellulose degree of polymerization. These findings establish a function for CSI1 in wood mechanics and in defining leaf cell shape. Furthermore, the results imply that the microfibril angle in wood is defined by CSI1 independent mechanism(s).
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| 9. |
- Cardoni, Simone, et al.
(author)
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5S‐IGS rDNA in wind‐pollinated trees ( Fagus L.) encapsulates 55 million years of reticulate evolution and hybrid origins of modern species
- 2021
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In: The Plant Journal. - : Wiley. - 0960-7412 .- 1365-313X. ; 109:4, s. 909-926
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Journal article (peer-reviewed)abstract
- Standard models of plant speciation assume strictly dichotomous genealogies in which a species, theancestor, is replaced by two offspring species. The reality in wind-pollinated trees with long evolutionaryhistories is more complex: species evolve from other species through isolation when genetic drift exceeds gene flow; lineage mixing can give rise to new species (hybrid taxa such as nothospecies and allopolyploids). The multi-copy, potentially multi-locus 5S rDNA is one of few gene regions conserving signal from dichotomous and reticulate evolutionary processes down to the level of intra-genomic recombination. Therefore, it can provide unique insights into the dynamic speciation processes of lineages that diversified tens of millions of years ago. Here, we provide the first high-throughput sequencing (HTS) of the 5S intergenic spacers (5S-IGS) for a lineage of wind-pollinated subtropical to temperate trees, the Fagus crenata – F.sylvatica s.l. lineage, and its distant relative F. japonica. The observed 4963 unique 5S-IGS variants reflect acomplex history of hybrid origins, lineage sorting, mixing via secondary gene flow, and intra-genomic competition between two or more paralogous-homoeologous 5S rDNA lineages. We show that modern species are genetic mosaics and represent a striking case of ongoing reticulate evolution during the past 55 million years.
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| 10. |
- Chen, Zeyuan, et al.
(author)
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Survival in the Tropics despite isolation, inbreeding and asexual reproduction : insights from the genome of the world's southernmost poplar (Populus ilicifolia)
- 2020
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In: The Plant Journal. - : Wiley. - 0960-7412 .- 1365-313X. ; 103:1, s. 430-442
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Journal article (peer-reviewed)abstract
- Species are becoming extinct at unprecedented rates as a consequence of human activity. Hence it is important to understand the evolutionary dynamics of species with already small population sizes. Populus ilicifolia is a vulnerable poplar species that is isolated from other poplar species and is uniquely adapted to the Tropics. It has a very limited size, reproduces partly clonally and is therefore an excellent case study for conservation genomics. We present here the first annotated draft genome of P. ilicifolia, characterize genome-wide patterns of polymorphisms and compare those to other poplar species with larger natural ranges. P. ilicifolia experienced a more prolonged and severe decline of effective population size (Ne) and signs of genetic erosion than any other poplar species with which it was compared. At present, the species has the lowest genome-wide genetic diversity, the highest abundance of long runs of homozygosity, high inbreeding levels as well as a high overall accumulation of deleterious variants. However, more effective purging of severely deleterious variants and adaptation to the Tropics may have contributed to its survival. Hence, in spite of its limited genetic variation, it is certainly worth pursuing the conservation efforts of this unique species.
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