Olivier Keech sitting at the desk in his office

Keech Olivier – Metabolic regulations in response to stress and sustainable food production

Research

Olivier Keech sitting at the desk in his officePhoto: Fredrik Larsson

Our research explores several aspects of the regulation of plant metabolism in response to stress, with a particular emphasis on mitochondrial metabolism. In plants, the process of aging as well as many environmental constraints may lead to the death of leaves. This particular type of cell death is often referred to as leaf senescence and can have a profoundly negative impact on crop yields and post-harvest shelf-life.

Aim: Leaves are essential plant structures and their well-being is crucial for plant development and survival. When a stress is applied, a plant has two options: try to cope with it or induce senescence and reallocate valuable nutrients towards new, developing or storage organs. A mutual antagonistic relationship can summarize this phenomenon as shown in figure 1. Our aim is to understand how the plant validates senescence over an adaptation strategy in response to stress (Fig. 1). This work mainly covers two aspects: 1) to unveil the communication and signalling mechanisms controlling the induction of leaf senescence and 2) to determine the subsequent metabolic regulation that occurs in response to stress, and ultimately during leaf senescence.

Illustration about the relationship between adaptation and senscence including metabolic/redox balance, hormonal homeostasis, gene regulatory network and different types of stressFigure 1: Mutual antagonistic relationship between adaptation and induction of senescence in response to a stress (e.g. nutrient deficiency, light regime, temperatures, pathogene infection, etc).

1. Using dark-induced senescence as a proxy to decipher signalling pathways controlling the induction of leaf senescence

In earlier studies (Keech et al., 2007; Law et al., 2018), we have shown that a leaf from a plant entirely darkened (DP) can survive much longer than an individually-darkened leaf (IDL; Fig. 2), which suggests that upon the right signals, the induction of leaf senescence can be repressed and alternative metabolic strategies conferring extended longevity can occur.

On the left side, an individual leaf was darkened and this leaf turned yellow after 6 days of treatment. On the right side, an entire plant was darkended and stayed mostly green after 6 days of treatment. Figure 2: Experimental setup for the two darkening treatments (Weaver and Amasino, 2001; Keech et al., 2007; Law et al., 2018).

Yet, our current knowledge on the respective metabolic adjustments remains highly fragmented. In 2018, we proposed the following working models (Fig. 3).

Illustration summarising metabolic strategies in plant leaves to darkeningComic strip by Neil E. Robbins II illustrating the effects of shading in plants

Figure 3: A) Model summarising the different metabolic strategies employed by plants in response to partial or total darkening of the plant. Size and line-weight of the fonts and arrows are proportional to their implication to these metabolic processes. The large arrow behind the leaf in DP conditions depicts the conserved metabolic strategy main-tained between 3 and 6 days of darkening. Abbreviations: AAA - aromatic amino acids, BCAA - branched chain amino acids, Citr - citrate, mETC - mitochondrial electron trans-port chain, OAA - oxaloacetate, PPP - pentose phosphate pathway, Shik/Chor - shikimate/chorismate, TCA - tricarboxylic acid cycle (Law et al., 2018); B) "Are plants afraid of the dark?" Comic strip by Neil E. Robbins II explaining the content of the publication in a humoristic way. Find the full comic strip here: https://neilercomics.com/2018/05/18/are-plants-afraid-of-the-dark/

However, in order to challenge these hypotheses, we are currently investigating the metabolic regulations in a set of functional stay-green mutants issued from a genetic screen. This provides us with a much valuable tool to determine how cells can survive prolonged stress conditions.

2. Regulation of metabolism during leaf senescence

In a green leaf, the three energy organelles (peroxisome, mitochondrion and chloroplast) work in synergy to sustain an efficient assimilation of carbon while constantly maintaining the essential functions of the cell. However, when a leaf undergoes senescence (“yellowing”), whole cell-metabolism is drastically modified, and as chloroplasts are rapidly getting impaired, the remaining organelles acquire novel functions, particularly the mitochondrion. In animals, mitochondria have been shown to integrate various signals and to subsequently modulate cell death processes whereas in plants, the contribution of mitochondria in cell death regulation remains unclear, particularly during leaf senescence.

Therefore, we are currently investigating in more detail the role of mitochondria during both developmental (i.e. aging) and stress-induced leaf senescence (Fig. 4).Illustration of metabolic processes in the mitochondriumFigure 4: Production of glutamate, reducing equivalents and TCA cycle intermediates from catabolic reactions occurring in the mitochondrion during developmental leaf senescence (Chrobok et al., 2016). Transcriptomic overview of the mitochondrially localised portion of the following metabolic pathways: (I) Lysine degradation, (II) branched chain amino acid degradation, (III) D-2HG metabolism, (IV) Glycine and Alanine metabolism, (V) Urea Cycle and (VI) Proline metabolism. Specific genes of these pathways and their transcript abundance during developmental leaf senescence are illustrated here. Production of reducing equivalents is shown as an arrow with an electron (e-).

3. Towards sustainable food production

Among a few other things, we are also interested in complementary alternatives for food production systems. In particular, we are involved in several projects aiming at developing integrated aqua-agro systems in closed land-based units. The strategic implementation of numerous trophic layers within a production system is a natural way to achieve a higher sustainability while maintaining the whole production economically viable.

A concept scheme (Fig. 5), released for the PLATSEN* event end of 2016 depicts some of the interrelationships between the different trophic layers that can be implemented to for example urban farming system in order to achieve a circularity, i.e. a better use of biowaste, energy and resources.

Depiction of the nutrient cycle between the different components of the eMTE modelSchematic overview of the eMulti-Trophic Ecosystem (eMTE) concept

More information about the eMTE project and the exhibition at PLATSEN in 2016.

PLATSEN is thought as a platform where decision makers, politicians, scientists, NGOs and people from public and private sectors can meet and exchange and discuss ideas about sustainability in an urban environment. The 2016 event was initiated by the Swedish Scientific Council for Sustainability in collaboration with several other actors from the public and private sectors e.g. Umeå Municipality and Umeå University.

Integrated fish and plant production workshop 2021

"Towards sustainable urban food production with multi-trophic systems", talk starts at 59 min: Link to the recorded workshop on SLU Play

Kelly Swarts standing outside next to a pine tree with snow in the back; she has long brown hair and a black sweater on

Swarts, Kelly - Tree Ring Genomics

Research

Kelly Swarts standing outside next to a pine tree with snow in the back; she has long brown hair and a black sweater onPhoto: Johan Gunséus

How do conifers adapt under climate change?

Conifers are ecologically dominant and economically important, but are globally succumbing to drought, disease, early-budding and other challenges. One of the oldest lineages on earth, conifers have the genetic diversity to adapt to a broad range of environments. However, long generation times combined with the greatly increased rate of climate change globally challenges trees’ ability to adapt, resulting in weakened individuals and eventually stand loss.

We use quantitative, computational and population genetic approaches in forests across Europe to understand climate adaptation in conifers. To understand how trees adapt to their environment – and how they might perform under changing climate – we first need to understand how much of growth is due to genetics and how much can be attributed to the experienced environment. By focusing on annual growth measured from tree-rings, which form when growth stops during the winter months in temperate regions, we can observe individuals growing across their lifespan. For the years with available environmental data from historical records, weather stations or satellites, we can model individual environmental responses, which are often highly heritable.

Close-up of a cross-section of a tree stem showing tree rings; a red jagged line is placed on top the tree ring photo illustrating the avarage growth during different years starting from before 1950 to 2020Many trees today are struggling in the face of changing climate. This increment core from a Norway Spruce (Picea abies) tree growing in Berchtesgaden, Germany has seen a decline in growth over the last few decades.

Once we have estimates of adaptation for individuals, we can parse the genome to understand the genetics underlying adaptation, and how this may differ across species and populations. We can also generate predictive models to estimate how well adapted an individual might be to an environment that it hasn’t experienced. Because we are working with tree-rings, we are not limited to controlled populations. This means that we can evaluate any tree’s performance in any modeled environment. As environments shift under climate change, this approach provides a powerful tool to select parents for healthy, resilient forests.

Schematic overview about the work flow for doing tree ring genomics

Our work is funded by generous support from:

KAW Logotype enSciLifeLab Logotype Green POSFWF Logo Zusatz Dunkelblau RGB ENUSNSF Logo Lockup stacked RGB 1200ppiLOGO ERC FLAG FP



Vaughan Hurry standing infront of a tree.

Hurry, Vaughan - Plant Adaptation to Sub-Optimal Environments

Research

Vaughan Hurry standing infront of a tree.

Our primary research goal is to identify the key adaptive mechanisms that result in short- and long-term acquisition of abiotic stress tolerance. To address this, our research currently has two main themes: 1) how are environmental "signals" sensed and, in turn, converted into a genetic response, and 2) how is primary metabolism modulated in response to fluctuations in growth temperature. The outcomes from this research are being applied to developing new tools for increased stress tolerance in herbaceous crops and forest plantation species and to studies of how we can incorporate understanding of acclimation of primary metabolism into global circulation models.

One of the key questions on the international research agenda today is how various biotope, natural and cultivated, will respond to the changes to the environment resulting from human activities. Plant carbon metabolism plays a crucial role in determining the functioning of terrestrial ecosystems, the concentration of CO2 in the atmosphere and the mean annual temperature of the earth's surface. Each year, photosynthetic carbon assimilation removes ca. 120 gigatonnes (Gt) of carbon from the atmosphere, with much of this carbon being used by heterotrophic organisms (i.e. animals, fungi, and bacteria).

Two octagonal transparent plastic tents set up in a forest.Scaling up from laboratory based experiments to ecosystem – level responses can be facilitated by studies in intact systems. The experiment shown is from the CANIFLEX project where the fate of carbon taken up by the forest was tracked through the trees and the soil bioa and back to the atmosphere using stableisotopes. The impacts of environmental changes, such as altering nitrogen availability, could then be studied at different trophic levels within the intact forest stand. This large scale, multiyear study was carried out together with colleagues from UPSC (T. Näsholm) and SLU (P. Högberg and S. Linder).

In addition, plants return ca. 60 Gt carbon per year to the atmosphere via respiration when producing the energy and carbon intermediates necessary for biosynthesis and cellular maintenance. This is a very large flux compared with the ca. 8 Gt carbon per year released from the burning of fossil fuels. Thus, fundamental metabolic processes such as photosynthesis and respiration play a critical role in determining a wide range of ecological phenomena, from the productivity of individual plants, species fitness, particular environments, and the resulting species composition of particular biotopes. Understanding such processes, and how they respond to environmental perturbations, provides insight into the underlying mechanisms that will drive future phenotypic replacements in response to climate change. Growth temperature is one of the most important climate parameters that impacts on the global fluxes through these C-assimilatory and C-emission pathways.

For example, as part of the thermal acclimation process (i.e. adjustment in the rate of metabolism to compensate for a change in growth temperature), cold-grown leaves exhibit higher transcript and activity levels of photosynthetic and sucrose synthesis enzymes, accompanied by increased capacity of mitochondrial electron transport than their warm-grown counterparts. As a result, sustained exposure to low growth temperatures typically results in an increase in the rate of assimilation and respiration at low temperatures. Given the predicted increase in the annual mean temperature of the Earth's surface, a major challenge for plant ecology and climate-vegetation modelling is identifying whether sustained changes in growth temperature will systematically alter the leaf-trait scaling relationships linking assimilation and respiration to leaf mass to area ratio and nitrogen concentrations.

To answer this challenge, a far better understanding of the responses of organellar functions to fluctuations in environmental inputs (e.g. temperature, water and nutrients) is required. We have shown that incorporating acclimation into the predictive models results in significant regional effects on the prevalence of different functional groups in different biomes. For example, it alters the predictions of the abundance of needle trees in the boreal forest zone relative to broad-leafed trees. Such changes will have very significant consequences for major industries such as Sweden's forest industry and consequently for the national economy. Our future research will develop additional data sets to incorporate acclimation to temperature,variations in response to altered soil nutritional status, rainfall, etc. to improve the predictive capacity of climate models.

Key Publications

  • Näsholm, T., Högberg, P., Franklin, O., Metcalfe, D., Keel, S.G., Campbell, C., Hurry, V., Linder, S., Högberg, M.N. (2013). Are ectomycorrhizal fungi alleviating or aggravating the N limitation to tree growth in boreal forests? New Phytologist. 198: 214-221.
  • Ruelland, E., Vaultier, M.N., Zachowski, A., Hurry, V. (2009). Cold signalling and cold acclimation in plants. Advances in Botanical Research, 49: 36-149.
  • Atkin OK, Atkinson LJ, Fisher RA, Campbell CD, Zaragoza-Castells J, Pitchford JW, Woodward FI, Hurry V (2008). Using temperature-dependent changes in leaf scaling relationships to quantitatively account for thermal acclimation of respiration in a coupled global climate-vegetation model. Global Change Biology, 14: 2709-2726.
  • Högberg P, Högberg MN, Göttlicher SG, Betson NR, Keel SG, Metcalfe DB, Campbell C, Schindlbacher A, Hurry V, Lundmark T, Linder S, Näsholm T (2008). High temporal resolution tracing of photosynthate carbon from the tree canopy to forest soil micro- organisms. New Phytologist, 177: 220-228.
  • Campbell C, Atkinson L, Zaragoza-Castells J, Lundmark M, Atkin O, Hurry V (2007). Acclimation of photosynthesis and respira- tion in response to changes in growth temperature is asynchronous across plant functional groups. New Phytologist, 176: 375–389
Stefan Jansson in front of his favourite aspen tree

Jansson, Stefan - How do trees survive the winter?

Research

Stefan Jansson in front of his favourite aspen treePhoto: Fredrik Larsson

Trees have evolved to survive the harsh winters of the boreal forests, but deciduous trees and conifers have chosen different strategies; either to shed their leaves or to stay green over the winter. We are trying to understand the molecular details behind these strategies.

How do aspens know it is autumn?

We are learning how to identify the genetic differences that make trees different from each other. Forest trees are, in general, more genetically diverse than most other organisms and aspens are, in this respect, extreme. We have built many genomic resources for Populus in general and aspen (P. tremula) in particular, for example clone collections (the SwAsp and UmAsp collections), ESTs, genome sequences as well as bioinformatic tools and databases.

Stefan Jansson's “favourite" aspen tree in front of Umeå University.Our “favourite" aspen tree, growing on the University campus in Umeå.

Using these tools, we study how aspens acclimate and adapt to the environment. Particular attention is paid to the process of phenology, in particular autumn senescence, trying to answer the question: How do trees know it is autumn? We are studying gene expression, photosynthesis and metabolism of the leaves during autumn senescence. In these studies, we use transgenic plants but, more importantly, natural variation. There is a steep cline in autumn senescence; trees from northern latitudes enter senescence much earlier than those from southern latitudes, and by using the aspen genome sequence, the collection of aspen clones and genetic tools like association mapping, we hope to understand the genetic basis of this important trait.

Yellow and red coloured aspen leafHow does the tree know that it is autumn?

How can conifers stay green in the winter?

In the photosynthetic apparatus of green plants, the light- harvesting chlorophyll a/b-binding (LHC) proteins serve as antennae for photosystems I and II. Members of the LHC protein family have three membrane-spanning regions and bind the majority of the photosynthetic pigments (chlorophyll and carotenoids), make the photosynthetic light reaction efficient and regulate the photosynthetic light reaction, for example by dissipating excess light and adjusting the excitation balance between the photosystems. There are a group of proteins that are more distant members of this protein family. This group includes PsbS and ELIPs. PsbS is necessary for a light dissipation process – the qE type of non-photochemical quenching (NPQ) - that operates when the plants are exposed to "excess light".

We are now focusing on how the photosynthetic apparatus of conifers have been adapted to make it possible for conifers to keep their leaves (needles) green over the winter. We are using molecular biology, biochemistry, biophysics etc. to study conifers grown in the field, over the season. Have the conifers evolved specific molecular mechanisms that allow them to stay green in the winter, or do they employ the same mechanisms as other plants, but to a higher extent?

Key Publications

  • Nystedt B, Street NR et al. (2013). The Norway spruce genome sequence gives insights into conifer genome evolution.Nature 497:579-584
  • Tuskan GA, DiFazio S, Jansson S, et al. (2006). The genome of black cottonwood, Populus trichocarpa (Torr. & Gray) Science 313:1596-1604
  • Sterky F, Bhalerao RR, Unneberg P, Segerman B, Nilsson P, Brunner AM, Campaa L, Jonsson Lindvall J, Tandre K, Strauss SH, Sundberg B, Gustafsson P, Uhlén M, Bhalerao RP, Nilsson O, Sandberg G, Karlsson J, Lundeberg J, Jansson S (2004). A Populus EST resource for plant functional genomics. PNAS 101 13951–13956
  • Külheim C, Ågren J, Jansson S (2002). Rapid regulation of light harvesting is crucial for plant fitness in the field. Science 297:91-93
  • Li, X-P, Björkman, O, Shih C, Grossman, AR, Rosenquist, M, Jansson, S, Niyogi, KK (2000). A pigment binding protein essential for regulation of photosynthetic light harvesting. Nature 40: 391-395
Portrait photo of Maria Rosario Garcia-Gil

García Gil, María Rosario - Forest Tree Genetics and Breeding

Research

Portrait photo of Maria Rosario Garcia-Gil Photo: Juha Niemi

Norway spruce and Scots pine are the two most economically important species in Fennoscandia. Despite several decades of breeding activity both species remain mostly undomesticated and thus still retain great potential for improvement. The traditional phenotype-based selection of superior trees is now undergoing a fast revolution where genomic-based selection (Genomic Selection) can substitute costly field tests. 

Genomic selection full potential requires of accurate and high-throughput phenotyping tools which will allow incorporating into the breeding programs a number of properties which otherwise were too expensive or tedious to measure, such as wood physical and chemical properties.

Genomic Selection (GS) for wood properties

Genomic prediction using genome-wide dense markers or genomic selection (GS) was first introduced by Theo Meuwissen. The method builds on the estimation of the effect of large numbers of DNA markers covering the entire genome and subsequently predict the genomic value of individuals that have been genotyped, but not phenotyped. As compared to the phenotypic mass selection based on a pedigree-based relationship matrix (A matrix), genomic prediction relies on constructing a marker-based relationship matrix (G matrix). The superiority of the G-matrix is the result of a more precise estimation of genetic similarity based on Mendelian segregation that not only captures recently pedigree but also the historical pedigree, and corrects possible errors in the pedigree. At MRGG´s lab we develop GS models for wood physical (density, MFA, MOE…) and chemical (cellulose, hemicellulose, lignin…) properties in Norway spruce. Aspects such as phenotyping efficient protocols for wood properties, single nucleotide polymorphisms (SNP) coding, environmental factors, progeny size, consanguinity are part of the model development for its implementation into operational breeding.

Collage of four graphs with a bundel of lowering black lines overlayed by one red line. Radial trends for MFA of Picea abies at two trials (Höreda and Erikstorp) from cambial age 1 to 20 and for Pinus contorta at two trials (Övra and Lagfors) from cambial age 1 to 30. The black lines represent the actual observations from all individual trees and the red line is the mean radial variation of MFA against the cambial age.

Inbreeding depression in Scots pine

In Sweden, conifer breeders are aware about the possible negative effect of increased inbreeding, so called inbreeding depression (ID), after successive cycles of selection. Moreover, inbreeding has been suggested, although not empirically evaluated, as a potential breeding tool through the exploitation of within-species heterosis. With no doubt inbreeding management is a central question in conifer breeding that has mostly been address through theoretical models, whereas empirical data has not been generated to probe any of those models right. At MRGG´s lab we conduct genomic studies on inbreed crosses of Scots pine to dissect the genetic control of inbreeding and its potential exploitation in conifer breeding.

Collage of five graphs showing the ring on the x-axis in relation to ring width (top left), stem diameter (top right), wood density (bottem left), microfibril angle (MFA; bottom middle) and modulus of elasticity (MOE; bottom right)Mean values generated with SilviScan data from the open-pollinated progenies and from the clonal archive.

Local adaptation in response to light quality

As a consequence of climate change, it is expected mean temperature to increase in 1.5 oC to 2oC, which is seen by the breeders as an opportunity for assisted migration northwards of the southern genotypes to benefit from their genetically governed extended growth. This would result in a substantial gain in volume. However, in addition to potential limitations in the extention of the growth season such as early spring frost, there are certain environmental cues that are not expected to change following the climate change, but to which trees are also adapted, such as light quality, day length and light intensity. This could limit the value of assisted migration. At MRGG´s lab we are investigating the genetic basis of local adaptation to light intensity and quality. During the growth season the trees in the north are exposed to higher proportions of far-red (FR) to red (R) light, which has resulted in a steep local adaptation to the response to R/FR ratio. These studies have involved greenhouse, cabinet experiments in combination with microarray and transcriptomic (RNAseq) data.

Shade avoidance response in conifers

Plants sense and respond to light quality and intensity, which is reflected at all different stages of seedling development. Response to shade differs among species in both angiosperms and gymnosperms depending on their level of tolerance to shade. Shade avoidance syndrome (SAS) is well-characterized in the shade intolerant model species A. thaliana whereas much less is known about the shade tolerance response (STR). At MRGG´s we conduct comparative studies in two conifer species with contrasting responses to shade; Scots pine, (shade-intolerant) and Norway spruce (shade-tolerant), with the aim to understand mechanisms behind SAS and STR in conifers.  

Key Publications

  • Ranade SS & García-Gil MR (2013). Adaptive cline to light spectra in Scots pine (Pinus sylvestris L). Tree physiology. 4: 479-493
  • Ranade SS, Abrahamsson S, Niemi J, and García-Gil MR (2013). Comparison of global expression profile under red light and far- red light in a conifer species. American Journal of Plant Science 4:479-493
  • Abrahamsson S, Hallander J, Waldmann P and García-Gil MR (2013). Heterozygosity-fitness correlation (HFC) in an inbreed Scots pine population. Genetica, DOI10.1007/s10709-013-9704-y
  • Nystedt B et al (2013). The draft sequence of the 20 GBp Norway spruce (Picea abies) genome (shed light on conifer genome evolution). Nature doi:10.1038/nature12211
  • Sillanpää MJ, Pikkuhookana P, Abrahamsson S, Fries A and García-Gil MR (2012). Simultaneous estimation of multiple quantitative trait loci and growth curve parameters through hierarchical Bayesian modeling. Heredity 108(2): 134-146
  • Abrahamsson S, Nilsson JE, Wu H, García-Gil MR, Andersson B (2012). Inheritance of height growth and autumn cold hardiness based on two generations of full-sib and half-sib families of Pinus sylvestris. Scandinavian Journal of Forest Research. 27:415-413
Johannes Hanson standing in an in vitro plant growth room with a flask of plant cell culture in the hand

Hanson, Johannes - Stress adaptation in plants

Research

Johannes Hanson standing in an in vitro plant growth room with a flask of plant cell culture in the handPhoto: Mattias Petterson Plants need highly efficient responses to adverse environmental conditions as they are bound to a single location. By totally changing their physiology plant can adapt to new environmental situations. These processes are in natural environments discriminative for plant fitness and in agricultural systems determining yield. Reprogrammed metabolism and changed translational patterns are important elements of stress adaption. The goal of the group is to understand how plants adjust their metabolism and translation in response to a changing environment. On the longer term we want to use this knowledge to design better trees and crops.

Adverse environmental conditions often cause limited energy availability and plant cells respond to this by reprograming their metabolism to better fit the new situation. This dramatic change involves hundreds of gene products and metabolites; we call this the Low Energy Syndrome, LES. The change is mastered by the SnRK1 kinase complex, which is able to react to low levels of metabolizable sugars. This parallels the manner in which all eukaryotes regulate starvation responses. In plants the SnRK1 kinases regulate gene expression of genes encoding key metabolic enzymes by activating certain bZIP transcription factors. One of our projects focuses on these transcription factors. We are interested in their mode of action and how their activity is regulated. Technically we are using high throughput expression analysis (massive sequencing) and metabolic profiling as central analysis tools combined with genetics and transgene based methods.

Illustration of the signalling pathway activated by stress Low energy availability and stress activate signaling cascades in the plant, initiated by activation of the SnRK1 kinase and resulting in changed metabolism and growth – The Low Energy Syndrome (LES). The aspects of interests for us are indicated.

When conditions are favorable for plant growth the SnRK1 complex is deactivated and a second major signaling system takes over mastered by another kinase - The Target of rapamycin, TOR that is positively regulates growth in all eukaryotes. TOR does so partly by regulating translation, which is a very energy consuming process and is therefore tightly regulated. The second major project in the laboratory deals with the regulatory mechanism of translational control by focusing on the activity of the ribosome. We currently are identifying novel components involved in translational changes using transcriptomics, translatomics, proteomics and genetic methodology.

The growing population of this planet will change our society. It is clear that food, feed and other plant-based resources will be limiting in the future. The grand challenge is to increase plant production a sustainable way. The transition to less fossil fuel dependent production will challenge our agricultural systems even further. Consequently, there is a basic need to optimize plant growth. This can be done by changed growth practices and reducing post-harvest losses, etc. However, we must use crop improvement to reach increased productivity similarly the green revolution half a decade ago. This is not limited to classical crops. We will need novel corps for biomass, bioenergy and biorefinery needs. By understanding the underlying mechanisms of growth-control we hope to find new ways to improve plant based production.

The figure illustrates how translation activity is assayedTranslation is assayed using density gradients where polysomes (P, translating ribosomes) are separated from monosomes (M, non-translating ribosomes). Translation varies dramatically depending on experimental condition or developmental changes A) Translation is inhibited by 6h extended night and increased by sucrose treatments (6 h treatment of 100 mM sucrose), as indicated by increased relative levels of polysomes. Sucrose treatments compensate for the extended night treatment and allow continued translation although low energy input from the light. B) Ribosomal preparation from germinating seeds showing primarily monosomes in dry seeds (0h) and more translation (polysomes) during germination (5 to 72 hours) (Bai et al., 2017). C) In poplar buds, with primary monosomes present in the dormant winter buds and increased translation as the bud growth is initiated during the spring as evident from increased polysome levels (André and Mahboubi, unpublished). D) By Using RiboSeq we can map the translational activity of single ribosomes to mRNAs Image indicate the ribosomes bound to mRNA and after degrading the parts of the mRNA that is not bound by ribosome we can sequence the protected fragments. E) Resulting patterns of mapped reads (blue bars) representing fragments translated by active ribosomes on a mRNA sequence (red bars, thick parts representing Open reading frames). Distance between the ticks on the scale is 5 kbp.
Ulrika Egertsdotter in a growth room with plants propagated by somatic embryogenesis

Egertsdotter, Ulrika - Somatic Embryogenesis (SE) in Conifers

Research

Somatic embryogenesis (SE) in conifers: a powerful research tool and a method to capture genetic gains from the breeding programs

Ulrika Egertsdotter in a growth room with plants propagated by somatic embryogenesis

Somatic embryogenesis (SE) is an in vitro based clonal propagation method that can be used as a model system for research, or for multiplication of valuable seeds of commercial value or for conservation of threatened species.

In order to meet the demands on future forests for higher productivity and also for higher adaptability to climate change, it is necessary to capture the genetic gains from the breeding programs. This can only be done by large-scale clonal propagation of elite trees selected from the breeding programs.

For spruce and other conifers, somatic embryogenesis (SE) is the only method that has the potential for clonal propagation to sufficiently large numbers of elite trees for commercial planting.

The process steps of somatic embryogenesis (SE) in conifers: it starts with the selection of elite seeds, followed by SE initiation, multiplication and maturation phases; mature embryos are selected for germination and the germinants are developing further into SE plants that on the long-term perspective produce new seedsThe process steps of somatic embryogenesis (SE) in conifers

Cost effective SE-plant production requires automated methods. Such methods have been demonstrated on a pilot scale for Swedish forestry operations.

In the UPSC SE lab, we have an automated instrument based on the same key technology utilized in the pilot system for SE plant production (the SE Fluidics System). The instrument is a valuable tool in fundamental research projects to study embryo development. It can perform dispersion, separation/singulation, image analysis and selection of plant propagules such as somatic embryos.

A collage of several photos illustrating the R&D SE Fluidics System and the different steps to singulate and harvest mature somatic embryos The R&D SE Fluidics System is mainly used for singulation and harvest of mature somatic embryos (system overview in A). Mature somatic embryos are produced in liquid medium in a temporary immersion bioreactor system (B) where hundreds to thousands of mature embryos are produced in each bioreactor (C: top view into bioreactor), or on solid medium in a petri plate (E). After processing in the Fluidics system, singulated mature embryos are analyzed by image analysis (D) and selected based on the pre-programmed selection criteria for embryos with good chances to form a plant (F).

Our research interest is to understand and explain the different processes that regulate development of conifer embryos into early-stage plants. We utilize somatic embryogenesis as a model system to study how metabolic processes are required and regulated during embryo development. By studying the nutritional requirements of the embryo during development and the correlated active cellular processes, we have found that nitrogen utilization appears to be regulated over the course of embryo development with the earlier embryo stages benefiting from a supply of organic nitrogen such as glutamine (Dahrendorf et al. 2018). During later stages of embryo development, our results indicate the importance of desiccation tolerance and suggest key functions for different types of carbohydrates (Businge et al. 2013). Key metabolic events during shoot and root apical meristem formation are associated with morphological events during early plant formation (Dobrowolska et al. 2016)

Key publications

  • Dahrendorf, J., Clapham, D., Egertsdotter, U. 2018. Analysis of Nitrogen Utilization Capability during the Proliferation and Maturation Phases of Norway Spruce (Picea abies (L.) H.Karst.) Somatic Embryogenesis. Forests 9(6): 288. https://doi.org/10.3390/f9060288
  • Dobrowolska, I., Businge, E., Abreu, I.N., Moritz, T., Egertsdotter, U. 2017. Metabolome and transcriptome profiling reveal new insights into somatic embryo germination in Norway spruce (Picea abies). Tree Physiology 00, 1–15. https://doi.org/10.1093/treephys/tpx078
  • Businge E, Bygdell J, Wingsle G, Moritz T, Egertsdotter U. (2013). The effect of carbohydrates and osmoticum on storage reserve accumulation and germination of Norway spruce somatic embryos. Physiologia plantarum 149(2): 273–285. https://doi.org/10.1111/ppl.12039
Black and white image of Maria Eriksson leaning against a tree trunk

Eriksson, Maria E - Circadian Clock Function and its Importance for the Regulation of Growth

Research

Black and white image of Maria Eriksson leaning against a tree trunkPhoto: Happy Wilder

The focus of the research group is to understand the functional aspects of the circadian clockwork in Arabidopsis and trees (Populus and other species), and how this timing machinery regulates growth. To anticipate the diurnal cycle of light and dark during a day and to anticipate the seasonal changes, most organisms have developed a molecular time measuring system called a circadian (from "circa diem" which in Latin means "about a day") oscillator or clock.

Light and temperature can be received by multiple photoreceptors in the red, far-red and blue spectra and mediates re-setting of this clock. In Arabidopsis, there are five red/far-red light photoreceptors called phytochromes (phy). The best characterized are phyA (far-red) and phyB (red). In the blue wavelengths, receptors like the cryptochromes (cry1 and cry2) are important, but also the ZEITLUPE (ZTL) gene family of F-box, Kelch-, and LOV/PAS domain containing proteins are capable of receiving blue light directly to regulate the circadian clock and seasonal timing. A central loop includes the morning expressed CIRCADIAN CLOCK ASSOCIATED1 (CCA1), and LATE ELONGATED HYPOCOTYL (LHY) which are MYB transcription factors that negatively regulate the gene expression of TIMING OF CAB2 EXPRESSION 1 (TOC1) so that it is expressed in the evening when CCA1 and LHY are turned over. TOC1 in turn mitigate expression of CCA1 and LHY. In addition, this negative feedback loop is intertwined with at least two additional interlocked feedback loops.

Populus orthologues of core clock genes LATE ELONGATED 1 (LHY1), LHY2 and TOC1 were targeted by RNA interference (RNAi) and allowed us to experimentally test their clock function and effect on growth. These studies showed that the circadian clock of Populus sp. trees contain a negative feedback loop of LHY1, LHY2 with TOC1 – similar to the situation in Arabidopsis. Our Populus ‘clock mutant’ RNAi trees also helped us to show that these proteins control seasonal timing of growth, cold response and freezing tolerance of trees.

Collage of four photos of the top of a poplar tree showing different growth stages Figure 1: Signs of season. An apex of Populus in active growth (upper left), at bud set (upper right), during dormancy (lower right) and at bud burst (lower left)


In the daily context, we found that a functional clock and the expression of the morning clock genes LHY1 and LHY2 are needed for growth. A key aspect of their regulation is obtained through regulation of CYCLIN D3 expression and thereby the G1 to S-phase transition of the cell cycle. Their functions are also needed to maintain cytokinin levels required for cell proliferation and growth, promoting biomass of plants.

Our very recent work places the photoreceptor and circadian clock protein ZTL (introduced above) as a critical integrator of light and circadian clock function with abscisic acid (ABA) signalling. ZTL promotes ABA-induced stomatal closure. It acts upstream of the PSEUDO-RESPONSE REGULATOR 5 (PRR5) to mitigate its function – but in addition ZTL also promotes ABA-induced gene expression and partner up with OPEN STOMATA 1 (OST1) to induce closing of stomata in response to ABA under drought stress. While timely expression of PRRs from dawn till dusk help keep stomata open, ZTL can short-cut and promote closure at the right time of day and in time of stress. Further, the role of ZTL is conserved between Arabidopsis and Populus trees. This picture (below) summarises our recent findings by Jurca et al., (2022).


Schematic overview on how ZEITLUPE promotes ABA regulated stomata closureFigure 2: Wild type (WT) and zeitlupe (ztl) mutants in Arabidopsis and Populus sp. trees show different responses to applied stress hormone abscisic acid (ABA) or drought stress in midday. The difference is for instance manifested by the inability of ztl mutants to close stomata to maintain water status in leaves that are detached. Leaves were weighted at regular intervals to track the loss of water vapor through stomata and those experiments showed a much larger water loss from the ztl mutant (shown by large water droplets in the picture) compared to the WT (smaller water droplets) in our recently published study by Jurca et al., 2022 in Frontiers in Plant Science. We also tested another clock mutant with a deficiency in PSUEDO-RESPONSE REGULATOR 5 (PRR5) (the prr5-1 mutant) which showed that PRR5 mitigates closure of stomata. The latter was elucidated using a triple mutant of ztl-3, prr5-1 and open stomata 1-3 (ost1-3). Our results suggested that ZTL could act to inhibit PRR5 (plain T-formed bar shows inhibition of activity, dotted bars indicate loss of this function) as well as independently to promote (plain arrow shows positive action, dotted arrows show loss-of-function) stomatal closure at the right time, in response to ABA and stress to protect the plant from losing precious water. (Illustration made by DC SciArt)

Hence, as we learn more about temporal regulation, there is a great potential for biotechnological application in adapting new plants or re-adapting (in case of climate warming) local plants to rapidly evolving "new" local conditions. Such adaptation may involve a means to increase the length of critical daylength requirements of plants to match a novel growth season, while keeping winter hardiness, as well as increasing biomass production.

To experimentally explore clock function and its role in growth, we use Arabidopsis thaliana for gene discovery. As tree model systems, we mainly use the deciduous tree hybrid aspen (Populus tremula x P. tremuloides) and the gymnosperm Norway spruce (Picea abies) to address the clock’s role in wood regulation and growth. We apply forward and reverse genetic approaches as well as assays of natural variation, as appropriate.

In the laboratory, we also use a combination of bioinformatics, genetic and molecular tools with in vitro/in vivo studies to study clock and protein function. Such tools for studying the clockwork and its adaptive value include plant cells or plants with altered levels of clock gene expression, molecular tools such as RNAseq, promoter:LUCIFERASE expression, real time PCR and protein assays to monitor circadian clock regulated gene and protein expression. To investigate perennial growth, we monitor elongation and diameter growth as well as physiological manifestations of season such as flowering, growth cessation, bud set and bud break. Mutants with an altered timing mechanism in this way help us to build a model for clock function and its impact on daily and seasonal regulation of growth.

Tips of populus trees in pixalated blue-to-white or green-to-yellow colour.Figure 3: Populus trees carrying firefly LUCIFERASE under control of a circadianly controlled promoter

Together, our studies of the circadian clock have contributed to understanding the importance of the circadian clock mechanism in weeds and trees: from its crucial impact on controlling water balance and photosynthesis through the control of stomatal regulation, to metabolism and synthesis of plant hormones as well as regulation of the cell cycle. Our future studies will further clarify the circadian clock mechanism and the important aspects of daily and seasonal timing for plant growth and development.

Key Publications

  • Jurca, M., Sjölander, J., Ibáñez, C., Matrosova, A., Johansson, M., Kozarewa, I., Takata, N., Bakó, L., Webb, A. A. R., Israelsson-Nordström, M., & Eriksson, M. E. (2022) ZEITLUPE Promotes ABA-Induced Stomatal Closure in Arabidopsis and Populus. Frontiers in Plant Science https://doi.org/10.3389/fpls.2022.829121
  • Edwards KD, Takata N, Johansson M, Jurca M, Novák O, Hényková E, Liverani S, Kozarewa I, Strnad M, Millar AJ, Ljung K, Eriksson ME (2018) Circadian clock components control daily growth activities by modulating cytokinin levels and cell division-associated gene expression in Populus trees. Plant Cell & Environment: 41(6):1468-1482 https://doi.org/10.1111/pce.13185
  • Eriksson, M. E., Hoffman, D., Kaduk, M., Mauriat, M., & Moritz, T. (2015) Transgenic hybrid aspen trees with increased gibberellin (GA) concentrations suggest that GA acts in parallel with FLOWERING LOCUS T2 to control shoot elongation. New Phytologist, 205(3): 1288–1295. https://nph.onlinelibrary.wiley.com/doi/pdf/10.1111/nph.13144
  • Johansson M, McWatters HG, Bakó L, Takata N, Gyula P, Hall A, Somers DE, Millar AJ, Eriksson ME (2011). Partners in time: EARLY BIRD associates with ZEITLUPE and regulates the speed of the Arabidopsis clock. Plant Physiology: 155:2108-2122 https://doi.org/10.1104/pp.110.167155
  • Ashelford K, Eriksson ME, Allen CM, D’Amore L, Johansson M, Gould P, Kay S, Millar AJ, Hall N, Hall A (2011). Full genome re-sequencing reveals a novel circadian clock mutation in Arabidopsis. Genome Biology: 12:R28, 12 pp https://doi.org/10.1186/gb-2011-12-3-r28
  • Ibáñez C, Kozarewa I, Johansson M, Ögren E, Rohde A, Eriksson ME (2010). Circadian clock components regulate entry and affect exit of seasonal dormancy as well as winter hardiness in Populus trees. Plant Physiology: 153:1823-1833 https://doi.org/10.1104/pp.110.158220
  • Kozarewa I, Ibáñez C, Johansson M, Ögren E, Mozley D, Nylander E, Chono M, Moritz T, Eriksson ME (2010). Alteration of PHYA expression change circadian rhythms and timing of bud set in Populus. Plant Molecular Biology: 73:143-156 https://doi.org/10.1007/s11103-010-9619-2
  • Eriksson ME, Hanano S, Southern MM, Hall A, Millar AJ (2003). Response regulator homologues have complementary, light- dependent functions in the Arabidopsis circadian clock. Planta: 218:159-162 https://doi.org/10.1007/s00425-003-1106-4
  • Eriksson ME, Israelsson M, Olsson O, Moritz T (2000). Increased gibberellin biosynthesis in transgenic trees promotes growth, biomass production and xylem fiber length. Nature Biotechnology 18:784-788 https://doi.org/10.1038/77355
Rishikesh Bhalerao standing in a growth room at one of UPSC's greenhouses with aspen trees in front and behind him

Bhalerao, Rishikesh P - Environmental information processing and seasonal adaptation

Research

Rishikesh Bhalerao standing in a growth room at one of UPSC's greenhouses with aspen trees in front and behind him Photo: Andreas Palmén

Overarching goal of my research group is to understand how plants process noisy environmental information and make robust decisions that are crucial to their survival. To address this fundamental question, we are using photoperiodic and temperature regulation of the timing of growth cessation, dormancy release and bud break that are the key developmental transitions in annual growth cycle in perennial plants using hybrid aspen as a model system.

Plants use environmental cues to make highly consequential decisions that are crucial to their survival. However environmental information such as temperature, a major regulator of plant development, is inherently noisy and how plants process such noisy information to make robust decisions remains poorly understood. Answering this fundamental question is essential to uncover mechanisms that enable plants to cope with their everchanging environment. Understanding how plants process noisy information is particularly fascinating because they make robust decisions despite lacking a central organizer such as brain in animals or a nervous system. My group is using the regulation of growth cessation, dormancy release and bud break, the key developmental transitions regulated by photoperiod and temperature to uncover molecular mechanisms that underlie environmental information processing in plants.

In long-lived trees native to boreal and temperate regions, growth stops prior to winter. The activity of shoot apical meristem (SAM) and formation and growth of leaf primordia is arrested and these are enclosed within a protective apical bud. We have identified the key components of a genetic network regulating growth cessation by photoperiod signal with sensing of shortening day length inducing growth cessation and bud break. The growth arrest is then maintained by until the advent of spring by establishment of dormancy. Dormant state is characterized by the insensitivity of SAM to growth promotive signal and involves suppression of cell-cell communication by blocking of specialized structures called plasmodesmata by callose, a polysaccharide. In order to undergo bud break and reinitiate growth in the spring, dormancy has to be broken. Release of dormancy is mediated by prolonged exposure (typical several weeks) of buds to low temperature, typically 4-8 °C. Once the buds receive sufficient chilling i. e. their chilling requirement is fulfilled, dormancy is released, progressively warmer temperatures induce bud break.

We have uncovered the key components of dormancy and bud break by temperature. A key feature of dormancy release is the restoration of cell-cell communication by deblocking of plasmodesmata by removal of callose, controlled by a recently uncovered genetic network. While these studies are highly successful, these have almost exclusively been performed in controlled growth conditions. Consequently, these studies do not reveal how buds perform long-term integration of variable (noisy) temperature signal. Our goal is to address this major gap in our understanding of mechanisms underlying variable temperature processing by trees in dormancy release and bud break in trees and from these studies provide a broader context of noisy environmental information processing in plants.

 Understanding the mechanisms underlying variable temperature processing by trees in dormancy release and bud break will provide broader insights into seasonal adaptation of trees to withstand increasingly unpredictable winter and spring temperatures caused by climate change.

Key Publications

  1. Nair A., Maurya JP., Pandey S., Singh R., Miskolczi P., Aryal A and Bhalerao RP (2025) ELF3 coordinates temperature and photoperiodic control of seasonal growth in hybrid aspen. Current Biology https://doi.org/10.1016/j.cub.2025.02.027
  2. Pandey S., Maurya JP., Aryal B., Drydna K., Nair A., Miskolczi P., Singh R., Wang X., Ma Y., Moraes T., Bayer E., Farcot E., Bassel G., Band L and Bhalerao RP (2024) A regulatory module mediating temperature control of cell-cell communication facilitates tree bud dormancy release. EMBO J: 43: 5793–5812 DOI: https://doi.org/10.1038/s44318-024-00256-5
  3. Ding J., Wang, K., Pandey S., Busov V and Bhalerao RP (2024) Molecular advances in bud dormancy in trees. Journal of Experimental Botany 75: 6063–6075 DOI: https://doi.org/10.1093/jxb/erae183
  4. Bhalerao RP (2024) Getting it right: Suppression and leveraging of noise in robust decision-making. Quantitative Plant Biology 5: e10 DOI: https://doi.org/10.1017/qpb.2024.10
  5. Azeez A., Zhao YC., Singh RK., Yordanov Y., Dash M., Misckolzi P., Stojkovic., Strauss S., Bhalerao RP* and Busov V (2021) The EARLY BUD-BREAK regulatory module in Populus. Nature Communications 12: 1123 DOI: https://doi.org/10.1038/s41467-021-21449-0 *co-corresponding author
  6. Maurya J., Misckolzi P., Mishra S., Singh R and Bhalerao RP (2020) A genetic framework for regulation and seasonal adaptation of shoot architecture in hybrid aspen. PNAS 117(21): 11523-11530 https://doi.org/10.1073/pnas.2004705117
  7. Maurya JP., Singh R., Misckolczi P., Prasad AN., Jonsson K., Wu F and Bhalerao RP (2020) Branching regulator BRC1 mediates photoperiodic control of seasonal growth in hybrid aspen. Current Biology 30: 122-126 https://doi.org/10.1016/j.cub.2019.11.001
  8. Misckolczi P., Singh RK., Tylewicz S., Azeez A., Maurya JP., Tarkowska D., Novak O., Jonsson K and Bhalerao RP (2019) Long-range mobile signals mediate seasonal control of shoot growth. PNAS 116: 10852-10857 https://doi.org/10.1073/pnas.1902199116
  9. Singh R., Misckolczi P., Maurya JP and Bhalerao RP (2019) A tree ortholog SHORT VEGETATIVE PHASE floral repressor mediates photoperiodic control of bud dormancy. Current Biology 29: 128-133 https://doi.org/10.1016/j.cub.2018.11.006
  10. Singh R., Misckolczi P., Maurya JP., Azeez A., Tylewicz S., Busov V and Bhalerao RP (2018) A genetic network mediating the control of bud break in hybrid aspen. Nature Communications 9: 4173 https://doi.org/10.1038/s41467-018-06696-y
  11. Tylewicz S., Petterle A., Martilla S., Misckolzi P., Singh R., Immanen J., Mähler N., Hvidsten T., Eklund D., Bowman J., Helariutta Y and Bhalerao RP (2018) Photoperiodic control of seasonal growth is mediated by ABA acting on cell-cell communication. Science 360: 212-215 https://doi.org/10.1126/science.aan8576

Current funding:

Logo of the Knut and Alice Wallenberg Foundation


Our work has been funded through the years by generous support from:

Logo of the Swedish Research CouncilLogo of the Knut and Alice Wallenberg FoundationLogo of the Human Frontier Science Program (HFSP)Logo of Kempestiftelserna - the Kempe FoundationsLogo of Marie Skłodowska-Curie ActionsLogo of Formas, a Swedish Research Council for Sustainable DevelopmentLogo of the Swedish Foundation for Strategic Research

 

Peter Marhavý at the confocal microscope

Marhavý, Peter - Short distance communication in response to wound stress

Research

Peter Marhavy at the confocal microscopePhoto: Fredrik Larsson

In my group, we are studying the mechanisms involved in short distance cell-to-cell communication in response to localized wound stress in plant roots using plant parasitic nematodes and state-of-the-art laser ablation approach.

Plants are able to recognize biotic, abiotic and physical stressors and orchestrate different signaling pathways accordingly. Although defensive stress response signals have been intensively studied and documented, the exact mechanisms by which these signals are perceived by cells and how the signal is further transmitted from one cell to another are still largely unknown. Our research will provide insights into molecular mechanisms of cell-to-cell communication to underlying early responses to wounding in plant roots.

  
Real-time monitoring of calcium wave propagation after cortex cell ablation using an R-GECO1 reporter line. Laser ablation of cortex cells leads to signal increases a few seconds after ablation at the ablated root side (indicated by white arrow). In non-ablated control roots, no changes of signal intensity were observed.
Arabidopsis root expressing SCR::SCR-YFP (green) in endodermis and stained with cell-wall fluorescence dye propidium iodide (red). Time lapse demonstrate single cell laser ablation in root meristem. White arrow indicates ablated cell.
Real-time monitoring (xyt) of cyst nematodes (Heterodera schachtii) during infection in 5-day-old roots of Arabidopsis thaliana stained with propidium iodide (red). Video demonstrate nematode progression between epidermal cells.

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