Project description:Analysis of the subunits composition of the thylakoids protein complexes in Picea abies (Norway spruce) by means of two-dimensional large-pore Blue-Native/sodium dodecyl sulfate-polyacrylamide gel electrophoresis (2D lpBN/SDS-PAGE) and in-gel tryptic digestion of single spots.
Project description:We investigated root associated fungi in young Norway spruce (Picea abies) cuttings rooted from slow- and fast-growing trees showing variable growth rate in long-term field experiments and compared their roots’ gene expression patterns five and 18 months after adventitious root initiation. Gene expression patterns of adventitious roots could not be systematically linked with the growth phenotype at the initiation of root formation, and thus fundamental differences in the receptiveness of fungal symbionts could not be assumed.
Project description:Coping of evergreen conifers of boreal forests with freezing temperatures on bright winter days puts the photosynthetic machinery in great risk of oxidative damage. To survive harsh winter conditions, conifers have evolved a unique but poorly characterised photoprotection mechanism, a sustained form of non-photochemical quenching (sustained NPQ). Here we focused on functional properties and underlying molecular mechanisms related to the development of sustained NPQ in Norway spruce (Picea abies). Data was collected during four consecutive years (2016-19) from trees growing in sun and shade habitats. When day temperatures dropped below -4°C, specific N-terminally triply phosphorylated LHCB1 isoform (3p-LHCII) and phosphorylated PSBS (p-PSBS) were detected in the thylakoid membrane. Development of sustained NPQ coincided with the highest level of 3p-LHCII and p-PSBS, occurring after prolonged combination of bright winter days and temperature close to -10°C. Artificial induction of both the sustained NPQ and recovery from naturally induced sustained NPQ provided information on differential dynamics and light-dependence of 3p-LHCII and p-PSBS accumulation and dephosphorylation as essential prerequisites of sustained NPQ. Data obtained collectively suggest three novel components related to sustained NPQ in spruce. (i) Freezing temperatures induce 3p-LHCII accumulation independently of light, which is suggested to initiate de-stacking of appressed thylakoid membranes due to increased electrostatic repulsion of adjacent membranes. (ii) p-PSBS accumulation is both light- and temperature-dependent and closely linked to the initiation of sustained NPQ, which (iii) in concert with PSII photoinhibition is likely to trigger sustained NPQ in spruce.
Project description:The goal is to look at changes in the pattern of expression of the xylem transcriptome through the growth season in two spruces (Picea glauca and Picea abies). One-color comparison of active xylem collected in June, July, August and September, in two spruce species. Six biological repetitions per time point and specie, for a total of 48 slides.
Project description:The goal is to look at changes in the pattern of expression of the xylem transcriptome through the growth season in two spruces (Picea glauca and Picea abies).
Project description:Photosystem II (PSII) complexes are organized into large supercomplexes with variable amounts of light-harvesting proteins (Lhcb). A typical PSII supercomplex in plants is formed by four trimers of Lhcb proteins (LHCII trimers), which are bound to the PSII core dimer via monomeric antenna proteins. However, the architecture of PSII supercomplexes in Norway spruce (Picea abies (L.) Karst.) is different, most likely due to a lack of two Lhcb proteins, Lhcb6 and Lhcb3. Interestingly, the spruce PSII supercomplex shares similar structural features with its counterpart in the green alga Chlamydomonas reinhardtii (Kouřil et al.: New Phytol. 210, 808-814, 2016). Here we present a single-particle electron microscopy study of isolated PSII supercomplexes from Norway spruce that revealed binding of a variable amount of LHCII trimers to the PSII core dimer at positions that have never been observed in any other plant species so far. The largest spruce PSII supercomplex, which was found to bind 8 LHCII trimers, is even larger than the current largest known PSII supercomplex from Chlamydomonas reinhardtii. We have also shown that the spruce PSII supercomplexes can form various types of PSII megacomplexes, which were also identified in intact grana membranes. Some of these large PSII supercomplexes and megacomplexes were identified also in Pinus sylvestris, another representative of Pinaceae family. The structural variability and complexity of LHCII organization in Pinaceae seems to be related to the absence of Lhcb6 and Lhcb3 in this family and may be beneficial for the optimization of light-harvesting under varying environmental conditions.
Project description:The widespread use of plant grafting has enabled different plants to join and combine their best properties to improve stress tolerance, growth and yields. Grafting is commonly performed between closely related eudicots or gymnosperms where mechanisms permit tissue fusion yet limit success as plants become unrelated. To investigate these aspects, we developed a micrografting method using young conifer tree seedlings that enabled divergent conifer members to successfully graft. Conifer grafts showed rapid connection of phloem and xylem at the junction, while a genome-wide transcriptome analysis of the Picea abies (Norway spruce) healing junction revealed differential expression of thousands of genes including those related to auxin response and cell wall biogenesis. We compared these genes to those induced during Arabidopsis thaliana graft healing and found a common activation of cambium, cell division, phloem and xylem-related genes. A gene regulatory network analysis revealed that PHYTOCHROME A SIGNAL TRANSDUCTION 1 (PAT1) acted as a central hub during Picea grafting and was also induced during Arabidopsis grafting. Arabidopsis mutants lacking PATs failed to attach tissues or successfully graft, while complementing Arabidopsis PAT mutants with the Picea abies PAT1 homolog could rescue tissue attachment and enhance callus formation. Together, our data demonstrate a competency for young tissues to graft to distantly related species and identifies the PAT gene family as conserved regulators of graft healing and tissue regeneration in eudicots and gymnosperms.
Project description:Monolignol transport during lignification is a partially solved puzzle: both the mechanism(s) and the transported form of monolignols are unknown in developing xylem of trees. We tested a hypothesis of an active, plasma membrane (PM)-localized transport of monolignol monomers, dimers, and/or glucosidic forms with membrane vesicles prepared of developing xylem, phloem, and lignin-forming, tissue-cultured cells of Norway spruce (Picea abies L. Karst.), as well as of a control material, non-lignifying tobacco (Nicotiana tabacum L.) BY-2 cells. Xylem and BY-2 vesicles transported both coniferin and p-coumaryl alcohol glucoside, but inhibitor assays suggested this transport being over the tonoplast. Based on similar inhibitor assays, lignin-forming, tissue42 cultured cells of spruce had coniferin transport putatively localizing on PM. Liquid chromatography-mass spectrometry (LC-MS/MS) analysis of membrane proteins isolated from spruce developing xylem, phloem and tissue-cultured cells revealed multiple transporters. These were compared to a transporter gene set that was gained by a correlation analysis with a selected set of spruce monolignol biosynthesis genes. Biochemical membrane vesicle assays showed no support for the ABC-transporter-mediated monolignol transport but point to secondary active transporters (such as MFS- or MATE-transporters). In contrast, proteomic and co-expression analyses suggest a role for ABC-transporters and MFS-transporters.
Project description:The widespread use of plant grafting has enabled different plants to join and combine their best properties to improve stress tolerance, growth and yields. Grafting is commonly performed between closely related eudicots or gymnosperms where mechanisms permit tissue fusion yet limit success as plants become unrelated. To investigate these aspects, we developed a micrografting method using young conifer tree seedlings that enabled divergent conifer members to successfully graft. Conifer grafts showed rapid connection of phloem and xylem at the junction, while a genome-wide transcriptome analysis of the Picea abies (Norway spruce) healing junction revealed differential expression of thousands of genes including those related to auxin response and cell wall biogenesis. We compared these genes to those induced during Arabidopsis thaliana graft healing and found a common activation of cambium, cell division, phloem and xylem-related genes. A gene regulatory network analysis revealed that PHYTOCHROME A SIGNAL TRANSDUCTION 1 (PAT1) acted as a central hub during Picea grafting and was also induced during Arabidopsis grafting. Arabidopsis mutants lacking PATs failed to attach tissues or successfully graft, while complementing Arabidopsis PAT mutants with the Picea abies PAT1 homolog could rescue tissue attachment and enhance callus formation. Together, our data demonstrate a competency for young tissues to graft to distantly related species and identifies the PAT gene family as conserved regulators of graft healing and tissue regeneration in eudicots and gymnosperms.