In our lab we have two major research interests:
I) Evolutionary Functional Genomics:
We study the evolution of complex physiological traits. As model organisms we use several species of the Brassicaceae family related to the genetic model plant Arabidopsis thaliana, the genome sequence of which was completed in 2000... continue
II) Metal Homeostasis Networks:
The survival of all organisms depends on an adequate supply of essential transition metals such as iron, zinc, and copper. These metals possess specific and uniquely powerful chemical properties required for indispensable biochemical functions accomplished by a host of metalloproteins... continue
We study the evolution of complex physiological traits. As model organisms we use several species of the Brassicaceae family related to the genetic model plant Arabidopsis thaliana, the genome sequence of which was completed in 2000. In particular, we are interested in a specific group of extremophiles – highly heavy metal-tolerant so-called metal hyperaccumulator plants, which we study in comparison to closely related non-accumulator plants such as Arabidopsis thaliana. Our aim is to understand the molecular mechanisms underlying extreme traits, and to identify the genes governing them. This knowledge then allows us to ask the question of how these traits evolved in nature. The results from our research can be applied in the development of technologies for the plant-based clean-up of metal-contaminated soils, termed phytoremediation. This research is also relevant for the bio-fortification of crops to improve their nutritional value.
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A typical heavy metal contaminated site around a former lead/zinc mine near Littfeld/Germany.
Arabidopsis halleri growing naturally at a heavy metal contaminated site around a former lead/zinc mine near Littfeld/Germany.
Arabidopsis halleri (left) and Arabidopsis thaliana (right) grown in hydroponic culture. Photo by Josef Bergstein, Max Planck Institute of Molecular Plant Physiology.
- Evolutionary Plant Solutions to Ecological Challenges: Molecular mechanisms underlying adaptive traits in the Brassicaceae s.l. (DFG SPP 1529)
- Comparative Molecular Analysis of Metal Homeostasis in the Arabidopsis Model Species A. halleri, a Metal Hyperaccumulator, and A. thaliana (DFG)
- Evolutionary Functional Genomics – Essential Computational Tools (University of Heidelberg Excellence Initiative Programme FRONTIERS)
- GABI-ADVANCIS: Development and Combination of In Silico and Novel Experimental Tools for the Advanced Genome-wide Identification of Cis-regulatory Elements (BMBF)
- PHIME: Public health impact of long-term, low-level mixed element exposure in susceptible population strata (EU FP6 InP)
- Comparative analysis of genotype-dependent nutrient fluxes and transcriptome dynamics for the modelling of plant metal homeostasis networks (in collaboration)
- Genetic Analysis of metal hyperaccumulation
Contributing group members:
- Dr. Heike Holländer Czytko
- Iris Sandorf
- Prof. Ute Krämer
The survival of all organisms depends on an adequate supply of essential transition metals such as iron, zinc, and copper. These metals possess specific and uniquely powerful chemical properties required for indispensable biochemical functions accomplished by a host of metalloproteins. Metal homeostasis networks ensure an adequate supply and the targeted delivery of transition metals to the sites of demand and protect sensitive sites from the fatal effects of an accumulation of excess or non-essential transition metals. In higher plants, metal homeostasis networks must operate over a particularly wide range of available concentrations encountered in the soil. Plant metal homeostasis is of major importance for global photosynthetic carbon fixation and crop yields, and – positioned at the beginning of the food chain – governs human intake of essential micronutrients and chemically similar toxic contaminants such as cadmium.
Our aim is a fundamental molecular understanding of the function and regulation of plant metal homeostasis networks. In order to maintain metal homeostasis, plants sense their metal status and modify metabolism, growth and development accordingly. Moreover, developmental and metabolic transitions require an adjustment of metal homeostasis. We aim to identify the major processes and pathways affected by this cross-talk between the metal homeostasis network and plant metabolism, growth and development, and to analyze the regulatory basis of this cross-talk.
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The plant metal homeostasis network. From: Clemens S, Palmgren MG, Krämer U (2002) A long way ahead: understanding and engineering plant metal accumulation. Trends Plant Sci. 7(7): 309-315
Plant copper status affects development. Shown are Arabidopsis thaliana grown in hydroponic culture under Cu deficiency (left, no added Cu), Cu toxicity (middle, 1.5 µM Cu) and Cu sufficient conditions (right, 0.5 µM Cu).
- Analysis of zinc transport systems of plant membranes (Japanese Society for the Advancement of Science)
- CENTZIN: Signalling, regulation and mechanisms of central components of zinc homeostasis (EU, Incoming International Fellowship)
- Regulatory Integration of Plant Metal Homeostasis (DFG)
GABI-ADVANCIS: Development and combination of in silico and novel experimental tools for the advanced genome-wide identification of cis-regulatory elements
Gene expression is mainly regulated by the binding of transcription factors to cis-regulatory sequence elements. These binding events thus govern plant development and architecture as well as the maintenance of critical cell type- and organ-specific functions, and enable plants to respond to endogenous and environmental cues by modifying growth, metabolism and developmental processes. All of these output processes are potential targets for crop improvement and plant biotechnology. For most agronomically important traits analyzed to date in crop plants, variation between cultivars has been genetically mapped to polymorphic regions upstream of genes rather than in their coding sequences. However, our current knowledge of the cis-regulatory sequence elements within these upstream regions and our ability to identify them reliably are still insufficient. The major goal of this project is to newly develop and combine existing and novel complementary in silico and experimental tools for an enhanced identification of cis-regulatory elements in plants (Fig 7). Of the known cis-regulatory sequence elements, many are conserved among different species of plants. The identification of novel cis-regulatory elements will allow both the design of novel effective markers for crop breeding and an improved control and specificity of transgene expression in plants.
Fig 7: Project strategy
A given transcription factor acts on a set of co-regulated genes, which are characterized by common cis-regulatory sequence elements. In this project we are generating comprehensive cross-cutting data access and bioinformatics tools for the genome-wide in silico identification of large numbers of putative endogenous cis-regulatory elements by employing the large, mostly untapped, resource of publicly available expression profiling data, open-access genome sequence data and complementary datasets newly generated in this project (Fig. 7, upper left). The identification of endogenous cis-regulatory elements in this project is based on a combination of (a) co-regulation analysis within large datasets of genome-wide expression data from several developmental stages, plant cell types and response-inducing conditions, (b) motif discovery in promoter sequences of co-regulated genes, (c) integration of transcript profiles and genomic sequence data from related plant species in a cross-species microarray hybridization approach to enhance the precision and reliability of predicted putative cis-regulatory elements (“phylogenetic footprinting”). In a complementary approach, we are additionally (d) developing and using tools for the experimental selection and evolution of synthetic cis-regulatory elements (Fig. 7, upper right, project part MPI). For experimental validation, (e) minimal promoters will be constructed using putative cis-regulatory elements to verify their responsiveness in cell culture and in intact plants, thereby generating proof-of-concept for the construction of synthetic specifically inducible promoters and providing validated cis-regulatory elements for testing in heterologous crop plant species (Fig. 7, lower right). For implementation, we will (f) integrate our results into the PathoPlant database and an expanded version of the AthaMap database (Fig. 7, lower left, project part TU Braunschweig).
The prerequisites for our combined approach are uniquely available in a set of Brassicaceae species, including Arabidopsis thaliana, A. lyrata and A. halleri. When genome sequence data and expression profiling platforms are available for additional sets of closely related plant species, it will be possible to also transfer the approach and the tools developed in this project to other sets of closely related model and agronomic plants, such as, possibly, cereals, Medicago truncatula and alfalfa, or Solanaceae species.
PHIME: Public health impact of long-term, low-level mixed element exposure in susceptible population strata
Malnutrition has been ranked second after HIV among the top five global challenges for the future. Among malnutrition-related health problems, the consequences of micronutrient deficiencies are particularly severe. Both iron and zinc deficiency have each been estimated to affect more than a third of the World's population. Toxic heavy metals, such as cadmium ions, can enter the human organism via micronutrient uptake pathways, and the contamination of soils with these heavy metals has been increasing dramatically since the beginning of the industrial revolution. Ironically, nutritional iron deficiency dramatically increases the uptake of cadmium from food in humans. Both essential and chemically similar non-essential metals enter the food chain through plants used, for example, as food or animal feed. Yet, to date, we do not have sufficient knowledge or plant cultivars to effectively improve metal contents in foods for optimum health benefits. By contrast, it appears that micronutrient contents have been decreasing in modern cereal varieties as an inadvertent consequence of breeding for other desired traits (Uauy et al., 2006).
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In pillar IV of the project PHIME, barley (Hordeum vulgare) is used as a cereal model to improve our knowledge on the molecular factors determining grain metal contents. Pillar IV includes a number of other European research groups: Michael G. Palmgren (Copenhagen, Denmark; coordinator), Prof. Maria Antosiewicz (Warsaw, Poland), Prof. Stephan Clemens (Bayreuth, Germany), Prof. Preben B. Holm (Flakkeberg, Copenhagen), Prof. Rafal Kucharski (Katowice, Poland), Prof. Enrico Martinoia (Zurich, Switzerland), Prof. Dale Sanders (York, UK), Prof. Lorraine Williams (Southampton, UK).
Our project part in PHIME has two research aims:
- The functional characterization of candidate genes that may have important roles in determining grain metal contents in barley, and of the encoded proteins.
- The exploration of natural diversity in grain metal contents in barley and closely related species, in collaboration with Prof. Stephan Clemens (University of Bayreuth, Germany) and Dr. Klaus Pillen (Max Planck Institute for Plant Breeding Research, Cologne, Germany), and the determination of physiological determinants of grain metal contents. Our long-term objective is to identify alleles that improve grain metal content for marker-assisted or molecular breeding.
Click here to go to the central website of the EU Integrated Project PHIME.
- Uauy C, Distelfeld A, Fahima T, Blechl A, Dubcovsky J (2006) A NAC Gene regulating senescence improves grain protein, zinc, and iron content in wheat. Science 314:1298-301
Comparative analysis of genotype-dependent nutrient fluxes and transcriptome dynamics for the modelling of plant metal homeostasis networks
We are using a combination of genomic, molecular and physiological techniques to generate datasets for the integrative modelling of metal fluxes and of the metal homeostasis network of plants. The modelling is being designed to explore and examine existing models of the plant metal homeostasis network (Clemens et al., 2002) using available knowledge on metal homeostasis protein biochemistry, transcriptional regulation and metal fluxes. Two processes will be considered. Firstly, we are developing data and computational approaches to model the pathway of a metal through the root and into the xylem, which is key in controlling metal accumulation in above-ground plant parts, such as leaves, fruits and seeds, which are often harvested. Secondly, we are analyzing the regulation of the metal homeostasis network at the transcript level (a) in response to alterations in metal supply, (b) following the targeted disruption by reverse genetics of key molecular elements of metal homeostasis (e.g. the Zn transporter controlling root-to-shoot transport of Zn), and both (c) using extremes of plant natural phenotypic diversity in metal homeostasis. We expect that the use of these datasets for modelling will reveal key functional and regulatory properties of the metal homeostasis network and will thus enable us to generate experimentally testable hypotheses concerning metal fluxes and their regulation in plants at the molecular level.
This project is a collaboration with Andrés Chavarría Krauser (BIOMS), who is in charge of the computational modelling.
- Clemens S, Palmgren MG, Krämer U (2002) A long way ahead: understanding and engineering plant metal accumulation. Trends Plant Sci. 7: 309-315
Earlier work has suggested that MTP1 (metal tolerance protein 1)-like transporters could play a key role in naturally selected as well as basal Zn tolerance and Zn partitioning in plants (Dräger et al., 2004; Desbrosses-Fonrouge et al., 2005; Willems et al., 2007). The aim of this project is to further elucidate the biological function of MTP1 proteins, which is still poorly understood, in the metal-tolerant model hyperaccumulator Arabidopsis halleri, compared to Arabidopsis lyrata and Arabidopsis thaliana as closely related non-accumulator model plants with only basal metal tolerance. This will be done using three approaches: First, RNA interference in A. halleri; second, the localization of MTP1 within plant organs and cells; and third, the isolation and analysis of the promoter sequences of the three MTP1 gene copies of A. halleri. In addition to providing novel insights into the molecular basis of naturally selected metal tolerance in A. halleri, an improved understanding of the role of MTP1 proteins in planta may have applications in phytoremediation or biofortification.
Fig 10: Model of the cellular function of MTP1 (Krämer, TIPS 2005).
- Desbrosses-Fonrouge A-G, Voigt K, Schröder A, Arrivault S, Thomine S, Krämer U (2005) Arabidopsis thaliana MTP1 is a Zn transporter in the vacuolar membrane which mediates Zn detoxification and drives leaf Zn accumulation. FEBS Lett. 579: 4165-4174
- Dräger DB, Desbrosses-Fonrouge A-G, Krach C, Chardonnens AN, Meyer, RC, Saumitou-Laprade P, Krämer U (2004) Two genes encoding Arabidopsis halleri metal transport proteins 1 (MTP1) co-segregate with zinc tolerance and account for high MTP1 transcript levels. Plant J. 39: 425-439
- Willems G, Dräger DB, Courbot M, Godé C, Verbruggen N, Saumitou-Laprade P. (2007) The genetic basis of zinc tolerance in the metallophyte Arabidopsis halleri ssp. halleri (Brassicaceae): an analysis of quantitative trait loci. Genetics 176: 659-74
It is a necessity for all organisms to adjust micronutrient uptake and transfer to the organism's needs at the target sites, i.e., apo-metalloproteins. In addition, plants in particular have evolved to respond appropriately to extreme fluctuations in the external supply of micronutrient metal ion concentrations and to fluctuations in the balance between different micronutrient elements and also non-essential heavy metals in the soil. For this, metal ions have to be sensed, and signals have to be generated, propagated and perceived, both locally and over long distances. There is clear evidence for metal-specific metal-status dependent regulation in plants. However, almost none of the molecular components of metal-dependent regulation are known to date.
In a comparative approach we have begun to address the transcriptional regulation of some key metal homeostasis genes in A. thaliana and A. halleri. We are particularly interested in Zn-dependent transcriptional regulation.
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MTP3 expression pattern in roots grown under excess zinc supply (Arrivault et al., 2006).
Vacuolar membrane localization of MTP3 (green; Arrivault et al., 2006)
In the yeast Saccharomyces cerevisiae, the synthesis of proteins requiring iron or iron-containing cofactors is decreased under conditions of iron deficiency because the transcript levels of the genes encoding these proteins are decreased (Puig et al., 2005). This regulation operates at the level of transcript stability and involves AU-rich element binding proteins of the tristetraprolin or CCCH family, in particular ScCTH2. Similar mechanisms may operate in A. thaliana. We are examining the role of Arabidopsis ScCTH2-related proteins in metal homeostasis, and in the interaction of metal homeostasis with plant metabolism and development (collaboration with Dr. Sergi Puig, University of Valencia, Spain).
Different divalent transition metal cations have similar ionic radii, and it is known that some metal cations can displace others from their binding sites. For example, Zn(II) can replace Fe(II). This is largely governed by the chemical properties of the metal ions as reflected by the Irving-Williams series. The displacement of metallic cofactor ions has to be avoided because the corresponding metalloprotein is inactivated as a result. Where and how do plants generate metal specificity?
To begin to answer this question, we are studying the example of the regulation of A. thaliana MTP3 at the transcript and protein levels. MTP3 transcript levels increase in response to iron deficiency, excess zinc and excess manganese, and yet the MTP3 protein can only detoxify excess Zn and Co through transport into the vacuole (Arrivault et al., 2006). Plants transformed with a fusion of the MTP3 promoter and 5' upstream sequences followed by a chimeric gene consisting of a partial MTP3 coding sequence and the UidA reporter showed enhanced reporter activity under conditions of iron deficiency, excess zinc, but not upon exposure to excess manganese. We are investigating which components of the regulatory pathway for MTP3 are common/distinct between the different metals.
- Arrivault S, Senger T, Krämer U (2006) The Arabidopsis metal tolerance protein AtMTP3 maintains metal homeostasis by mediating Zn exclusion from the shoot under Fe deficiency and Zn oversupply. Plant J. 46: 861-879
- Puig S, Askeland E, Thiele DJ (2005) Coordinated Remodeling of Cellular Metabolism during Iron Deficiency through Targeted mRNA Degradation. Cell 120, 99-110