Artificial metabolons – a tool for synthetic biology

Dr LJ Sweetlove

email: lee.sweetlove@plants.ox.ac.uk
Tel: +44 (0)1865 275000

Project Description

Synthetic biology aims to combine molecular parts from across the biological spectrum into synthetic organisms tailored for specific uses such as pollution clean-up, production of fuel or use for production of pharmaceuticals. In all cases, assembly of a metabolic system fit for purpose is one of the first steps that must be considered. Due to the complexity of metabolic networks, even of simple bacteria, it is unlikely that these metabolic systems will be built from scratch. Rather, it is envisaged that alteration of the capacity of existing metabolic networks will be the most efficient strategy ^1,2.

This would appear to be a simple process: identify another organism that has the desired metabolic capacity and transfer the genes encoding the relevant enzymes to the host organism. However, decades of metabolic engineering research has shown that this rarely works. This is either because the new pathway produces intermediates that are toxic and the host organism lacks the mechanisms to deal with these toxic compounds, or the new pathway produces intermediates that are common to the host metabolism and the change in intermediate concentration causes metabolic feedback inhibition.

What is needed is the ability to introduce a set of enzymes that function as a ‘plug and play’ metabolic module in which new metabolic intermediates are isolated from the host’s metabolic system. We are interested in designing such a modular system. We wish to test the hypothesis that anchoring introduced enzymes onto the inward-facing surface of the cell membrane will generate a synthetic ‘metabolon’ in which intermediates are directly handed off between sequential enzymes without diffusing into the bulk aqueous phase of the cytoplasm (a phenomenon known as metabolite channelling). This hypothesis is based on our observations that certain enzymes cluster onto the surface of intracellular membranes and when organised like this form a metabolon ³.

Currently, it is not understood why co-localisation of enzymes to a membrane surface leads to metabolite channelling and we are therefore seeking a D.Phil student to investigate this. A number of questions need to be addressed. For example, does co-localisation of enzymes promote protein-protein interactions between them? Is metabolite channelling a consequence of the biophysical properties of the membrane or the biochemical micro-environment at the membrane surface? Do the enzymes need to be spatially organised in a certain way, or is a random distribution on the membrane surface sufficient? We propose to investigate these questions using an in vitro system consisting of artificial liposomes with enzymes modified to have hydrophobic tails anchored to the liposome surface.

¹Boyle PM, Silver PA. 2011. Parts plus pipes: Synthetic biology approaches to metabolic engineering. Metab Eng 14:223-32
²Lee H, DeLoache WC, Dueber JE. 2012. Spatial organization of enzymes for metabolic engineering. Metab Eng 14:242-51
³Graham JW, Williams TC, Morgan M, Fernie AR, Ratcliffe RG, Sweetlove LJ. 2007. Glycolytic enzymes associate dynamically with mitochondria in response to respiratory demand and support substrate channeling. Plant Cell 19:3723-38

Recent Publications

Schwärzlander, M., Logan, D.C., Johnston, I.G., Jones, N.S. Fricker, M.D. and Sweetlove, L.J. (2012) Pulsing of membrane potential in individual mitochondria: a stress induced mechanism to regulate respiratory bioenergetics in Arabidopsis. Plant Cell, 24: 1188-1201

Graham, J.W., Williams, T.C., Morgan, M., Fernie, A.R., Ratcliffe, R.G., & Sweetlove, L.J. (2007). Glycolytic enzymes associate dynamically with mitochondria in response to respiratory demand and support substrate channeling. Plant Cell 19, 3723-3738.

Claridge JK, Schnell JR. 2012. Bacterial production and solution NMR studies of a viral membrane ion channel. Methods Mol Biol 831:165-79

Pielak RM, Schnell JR, Chou JJ. 2009. Mechanism of drug inhibition and drug resistance of influenza A M2 channel. Proc Natl Acad Sci U S A 106:7379-84

Funding Status

BBSRC via the Oxford Interdisciplinary Bioscience Doctoral Training Partnership (DTP) programme. See here for further details: http://www.biodtp.ox.ac.uk or Clarendon Fund - see here: http://www.clarendon.ox.ac.uk/about or via independent funding.

Evolution of metabolic regulation in plants

Dr NJ Kruger

email: nick.kruger@plants.ox.ac.uk
Tel: +44 (0)1865 275000

Project Description

An organism’s survival depends on coordination of its metabolic activities, which must be strictly regulated to integrate the operation of potentially competing processes. The regulatory mechanisms needed to allow metabolite levels and fluxes to respond to fluctuating environmental and developmental demands are likely to be adapted to meet the specific physiological needs of the system. Understanding the evolution of these mechanisms is key to explaining the adaptive significance of the regulatory processes and this underpins attempts to modify metabolism for specific biotechnological purposes.
The aim of this project is to examine how regulation of the pathways of carbohydrate utilisation has evolved in the green plant lineage. These processes are likely to have been subjected to strong selective pressure since carbohydrates are the major immediate product of photosynthesis, the dominant form in which carbon is stored and the principal respiratory substrate, making them essential for plant growth and survival. The research will focus on the signal metabolite fructose 2,6-bisphosphate (Fru-2,6-P₂), a potent regulator of carbohydrate metabolism in almost all eukaryotes. It is synthesised and degraded by the activities of a bifunctional enzyme (F2KP; fructose-6-phosphate 2-kinase, fructose-2,6-bisphosphatase). All higher plants have one or more F2KP isozymes in which the catalytic core is preceded by a conserved amino terminal domain of unknown function and absent from other eukaryotes. Moreover, although Fru-2,6-P₂ has analogous regulatory roles in different organisms the target enzymes for this effector differ - notably, plants contain an unusual pyrophosphate-dependent phosphofructokinase (PFP) that is uniquely activated by Fru-2,6-P₂. This project will use a combination of biochemical, biophysical, molecular genetic and bioinformatic approaches to investigate (i) the role of different F2KP isozymes in Fru-2,6-P₂ metabolism, (ii) the significance of the amino terminal domain in F2KP function, and (iii) the role played by this regulatory metabolite in plants through its novel interaction with PFP.

Recent Publications

N.J. Kruger and R.G. Ratcliffe (2009) Insights into plant metabolic networks from steady-state metabolic flux analysis. Biochimie 91, 697-702.
S.K. Masakapalli et al. (2010) Subcellular flux analysis of central metabolism in a heterotrophic Arabidopsis thaliana cell suspension using steady-state stable isotope labeling. Plant Physiology 152, 602-619.
T.P. Howard et al. (2011) Antisense suppression of the small chloroplast protein CP12 in tobacco alters carbon partitioning and severely restricts growth. Plant Physiology 157, 620-631.
D.J.V. Beste et al. (2011) 13C metabolic flux analysis identifies an unusual route for pyruvate dissimilation in mycobacteria which requires isocitrate lyase and carbon dioxide fixation. PLoS Pathogens 7, e1002091.
N.J. Kruger, S.K. Masakapalli and R.G. Ratcliffe (2012) Strategies for investigating the plant metabolic network with steady-state metabolic flux analysis: lessons from an Arabidopsis cell culture and other systems. Journal of Experimental Botany 63, 2309-2323.

Funding Status

BBSRC via the Oxford Interdisciplinary Bioscience Doctoral Training Partnership (DTP) programme. See here for further details: http://www.biodtp.ox.ac.uk or Clarendon Fund - see here: http://www.clarendon.ox.ac.uk/about or via independent funding.

Evolution of sex chromosomes in plants

Dr DA Filatov

email: dmitry.filatov@plants.ox.ac.uk
Tel: +44 (0)1865 275000

Project Description

Despite their independent evolution, sex chromosomes in different organismal groups have similar properties: recombination is restricted between the X and Y chromosomes, and the male-specific non-recombining Y chromosome exhibits genetic degeneration (loss of functional genes & accumulation of repetitive DNA). The X chromosome, on the other hand, continues to recombine in females and does not degenerate. One unresolved issue in sex chromosome evolution is how and why do Y-specific non-recombining regions form and expand in size? The dominant model proposes that this process is driven by sexually antagonistic (advantageous to males, but detrimental in females) genes, as moving such genes to a male-specific part of the genome (Y-chromosome) is advantageous. However, this mechanism is not expected to work in species with no or few differences between genders, such as plants. Yet plants did evolve sex chromosomes on separate independent occasions. Another interesting question is why do Y-chromosomes degenerate. All these questions are not easy to address in animals, where sex chromosomes are usually quite old. Sex chromosomes in such plants as Silene latifolia, Mercurialis annua and Carica papaya are of relatively recent origin and are more suitable to study the most informative early stages of sex chromosome evolution. Extensive genomic resources are becoming available for plants with sex chromosomes. These resources developed in our and other laboratories will be used in course of this project to study the evolutionary genetic processes involved in sex chromosome evolution in plants.

Recent Publications

1. Chibalina M.V. & Filatov D.A. (2011) Plant Y-chromosome degeneration is retarded by haploid purifying selection. Current Biology, 21: 1475-1479.

2. Muir G., Bergero R., Charlesworth D. & Filatov D.A. (2011) Does local adaptation cause high population differentiation of Silene latifolia Y chromosomes? Evolution. 65:3368-3380.

3. Howell E.C., Armstrong S.J. & Filatov D.A. (2011) Dynamic gene order on the Silene latifolia Y chromosome. Chromosoma 120: 287-296.

4. Howell EC, Armstrong SJ, Filatov DA.(2009) Evolution of neo-sex chromosomes in Silene diclinis. Genetics. 182:1109-1115.

5. Filatov D.A. (2005) Evolutionary history of Silene latifolia sex chromosomes revealed by genetic mapping of four genes. Genetics, 170: 975-979.

Funding Status

BBSRC via the Oxford Interdisciplinary Bioscience Doctoral Training Partnership (DTP) programme. See here for further details: http://www.biodtp.ox.ac.uk or Clarendon Fund - see here: http://www.clarendon.ox.ac.uk/about or via independent funding.

Metabolic flux analysis of plant pathogens

Dr GM Preston

email: gail.preston@plants.ox.ac.uk
Tel: +44 (0)1865 275000

Dr NJ Kruger

email: nick.kruger@plants.ox.ac.uk
Tel: +44 (0)1865 275000

Prof. RG Ratcliffe

email: george.ratcliffe@plants.ox.ac.uk
Tel: +44 (0)1865 275000

Project Description

Bacterial pathogens cause economically important diseases in many plant species and the molecular interactions that occur between the pathogen and its host are central to the disease process. Changes in gene expression in both partners are pivotal, but these changes occur against a background of metabolic cross-talk. Indeed there is increasing evidence that a pathogen such as Pseudomonas syringae manipulates the metabolism of its host for nutritional and other reasons. While substantial progress in delineating these metabolic processes has been made using bacterial cultures, complete understanding of the metabolic phenotype of a pathogen, and the metabolic response of the host, requires methods for measuring metabolic activity in situ.

To address this question, we propose to develop a new method for measuring metabolic fluxes in the host and bacterial cells of an infected plant tissue. The fluxes supported by a metabolic network are crucial to its function and they can be measured with stable isotope and radioisotope labelling methods. Here we propose to use the labelling of GFP, expressed in either the host or the pathogen, as a reporter protein for the metabolic activity of the corresponding cell type in the infected tissue.

The project will begin with an analysis of the metabolic activity of P. syringae in its host tissue, building on the extensive knowledge of this system in Oxford, but the proposed methodology could be equally applicable to other diseases, and to probing the metabolic interactions that occur in pathogen interactions with disease-resistant plants.

Recent Publications

S.K. Masakapalli, P. Le Lay, J.E. Huddleston, N.L. Pollock, N.J. Kruger and R.G. Ratcliffe (2010) Subcellular flux analysis of central metabolism in a heterotrophic Arabidopsis thaliana cell suspension using steady-state stable isotope labeling. Plant Physiology 152, 602-619.

A. Rico, S.L. McCraw and G.M. Preston (2011) The metabolic interface between Pseudomonas syringae and plant cells. Current Opinion in Microbiology 14, 31-38.

D.J.V. Beste, B. Bonde, N. Hawkins, J.L. Ward, M.H. Beale, S. Noack, K. Nöh, N.J. Kruger, R.G. Ratcliffe and J. McFadden (2011) 13C metabolic flux analysis identifies an unusual route for pyruvate dissimilation in mycobacteria which requires isocitrate lyase and carbon dioxide fixation. PLoS Pathogens 7, e1002091.

A. Mithani, J. Heinand G.M. Preston (2011) Comparative analysis of metabolic networks provides insight into the evolution of plant pathogenic and non-pathogenic lifestyles in Pseudomonas. Molecular Biology and Evolution 28, 483-499.

N.J. Kruger, S.K. Masakapalli and R.G. Ratcliffe (2012) Strategies for investigating the plant metabolic network with steady-state metabolic flux analysis: lessons from an Arabidopsis cell culture and other systems. Journal of Experimental Botany 63, 2309-2323.

Funding Status

BBSRC via the Oxford Interdisciplinary Bioscience Doctoral Training Partnership (DTP) programme. See here for further details: http://www.biodtp.ox.ac.uk or Clarendon Fund - see here: http://www.clarendon.ox.ac.uk/about or via independent funding.

Metabolic flux analysis of Rhizobium leguminosarum

Dr NJ Kruger

email: nick.kruger@plants.ox.ac.uk
Tel: +44 (0)1865 275000

Prof. RG Ratcliffe

email: george.ratcliffe@plants.ox.ac.uk
Tel: +44 (0)1865 275000

Project Description

Nitrogen fixation by legumes plays is essential for sustainable agriculture and the global nitrogen cycle. The process depends on establishing a symbiosis between nitrogen-fixing bacteria, such as Rhizobium spp, and the roots of a host plant, such as pea, leading to the formation of root nodules. The metabolic integration of the bacteroid into the host plant cell is essential for nitrogen fixation, and surprisingly amino acid import into the bacteroid from the host cell is a prerequisite for the net provision of fixed nitrogen to the host cell in the rhizobial symbiosis.

A complete understanding of the metabolic phenotype of the bacteroid and its host cell requires methods for measuring cell-specific metabolic activity, including measurements of the metabolic fluxes supported by the bacterial and plant cell metabolic networks. To address this problem, we intend to apply a novel strategy for analyzing cell-specific metabolism based on stable isotope labelling of cell-specific marker proteins.

The project builds on our expertise in steady-state metabolic flux analysis (MFA) and a technique that uses GFP as a marker protein to interrogate the metabolic state of specific cell types following incubation with ¹³ C-labelled substrates. We shall apply this method to Rhizobium leguminosarum, with the aim of understanding the changes in metabolic phenotype that occur between the free-living organism and the symbiotic state. Working with wild type and mutant strains of the bacterium, we shall deduce flux maps of primary metabolism for the bacteroid within its host for the first time.

Recent Publications

S.K. Masakapalli, P. Le Lay, J.E. Huddleston, N.L. Pollock, N.J. Kruger and R.G. Ratcliffe (2010) Subcellular flux analysis of central metabolism in a heterotrophic Arabidopsis thaliana cell suspension using steady-state stable isotope labeling. Plant Physiology 152, 602-619.

T.C.R. Williams, M.G. Poolman, A.J.M. Howden, M. Schwarzländer, D.A. Fell, R.G. Ratcliffe and L.J. Sweetlove (2010) A genome-scale metabolic model accurately predicts fluxes in central carbon metabolism under stress conditions. Plant Physiology 154, 311-323.

D.J.V. Beste, B. Bonde, N. Hawkins, J.L. Ward, M.H. Beale, S. Noack, K. Nöh, N.J. Kruger, R.G. Ratcliffe and J. McFadden (2011) 13C metabolic flux analysis identifies an unusual route for pyruvate dissimilation in mycobacteria which requires isocitrate lyase and carbon dioxide fixation. PLoS Pathogens 7, e1002091.

T.C.R. Williams, L.J. Sweetlove and R.G. Ratcliffe (2011) Capturing metabolite channeling in metabolic flux phenotypes. Plant Physiology 157, 981-984.

N.J. Kruger, S.K. Masakapalli and R.G. Ratcliffe (2012) Strategies for investigating the plant metabolic network with steady-state metabolic flux analysis: lessons from an Arabidopsis cell culture and other systems. Journal of Experimental Botany 63, 2309-2323.

Funding Status

BBSRC via the Oxford Interdisciplinary Bioscience Doctoral Training Partnership (DTP) programme. See here for further details: http://www.biodtp.ox.ac.uk or Clarendon Fund - see here: http://www.clarendon.ox.ac.uk/about or via independent funding.

Niche construction by plant pathogenic pseudomonads

Dr GM Preston

email: gail.preston@plants.ox.ac.uk
Tel: +44 (0)1865 275000

Project Description

Plant pathogens such as the bacterial pathogen Pseudomonas syringae infect plant tissues by injecting secreted proteins into plant cells to suppress plant defences, and by producing toxins and hormones that alter plant physiology to promote bacterial growth. The ability of P. syringae to manipulate its plant hosts can be regarded as a form of niche construction or ecosystem engineering, processes in which the activities of an organism or group of organisms alter the environment occupied by their descendants and by other organisms within the same ecosystem.
We have shown that the presence of P. syringae in plant tissues alters the chemical composition of the apoplastic fluid that surrounds plant cells, in which P. syringae grows. These changes may promote or restrict P. syringae virulence and growth, as well as altering plant fitness and interactions with other organisms. The aim of this project will be to investigate whether the chemical changes we have observed in the apoplast of infected plants represent positive or negative niche construction, i.e. whether they promote or inhibit the growth of subsequent generations of bacteria. We will also consider whether niche construction on an evolutionary time-scale has shaped the metabolic and regulatory features of P. syringae. In the long term we hope to use knowledge of niche construction by plant pathogens to develop novel strategies to enhance disease control.

Recent Publications

Rico, A., McCraw, S.L. & Preston, G.M. (2011) The metabolic interface between Pseudomonas syringae and plant cells. Current Opinion in Microbiology. 14, 31-38

Fones, H., and G. M. Preston. (2012). Reactive oxygen and oxidative stress tolerance in plant pathogenic Pseudomonas. FEMS Microbiology Letters 327:1-8.

Mithani, A., Hein, J. & Preston, G.M. (2011) Comparative analysis of metabolic networks provides insight into the evolution of plant pathogenic and non-pathogenic lifestyles in Pseudomonas. Molecular Biology and Evolution 28, 483-499

Preston, G.M., and Arnold, D.L. (2010) Karma chameleons: How bacterial plant pathogens escape their fate in disease resistant plants. Microbiology Today: 164-169.

Fones, H., Davis, C.A.R., Rico, A., Fang, F., Smith, J.A.C. & Preston, G.M. (2010) Metal hyperaccumulation armors plants against disease. PLoS Pathogens 6, e1001093

Howden, A.J.M., Rico, A., Mentlak, T., Miguet, L., and Preston, G.M. (2009). Pseudomonas syringae pv. syringae B728a hydrolyses indole-3-acetonitrile to the plant hormone indole-3-acetic acid. Molecular Plant Pathology I0: 857-865.

Funding Status

BBSRC via the Oxford Interdisciplinary Bioscience Doctoral Training Partnership (DTP) programme. See here for further details: http://www.biodtp.ox.ac.uk or Clarendon Fund - see here: http://www.clarendon.ox.ac.uk/about or via independent funding.

Cell biology of cell patterning

Dr IR Moore

email: ian.moore@plants.ox.ac.uk
Tel: +44 (0)1865 275000

Project Description

Significance: Most organisms comprise cells surrounded by a cell wall. Of these, land plants are the most complex. Their uniquely structured and assembled cell walls profoundly affect cell form and function and are a major determinant of crop plant quality and utility. A major challenge in plant biology is to understand how the extracellular cell wall is positioned, assembled, and remodelled through the activities of intracellular organelles and plasma membrane domains.

Plant cells cannot migrate so the form of the whole organism is determined through control of cell size and shape. Plant cells achieve polyhedral shapes that cannot be explained by simple packing and energy minimisation. Experimental and modelling approaches in this field have concentrated either on the physics and chemistry of cell wall polymers or on supra-cellular morphogenetic mechanisms. What is crucially lacking is an understanding of how cell shape is achieved at the most important level of organisation - the individual cell.

Research Background: Work in my laboratory has identified families of regulatory Rab GTPases that have arisen during land-plant evolution and define previously unrecognised intracellular organelles that are components of the secretory pathways for cell wall assembly and plasma membrane patterning. One of these GTPases also identifies the geometric edges of young plant cells as a distinct and previously unrecognised spatial domain. Perturbing the activity of these Rab GTPases leads to defects in cell division, growth, and cell-shape establishment. The challenge now is to identify the critical regulatory interactions that have co-evolved with these novel Rab GTPase specificities (Rab proteins do not contribute directly to the cellular activity they control but, like administrators in an organisation, they are crucial for organising the activities of those that do). Several candidates are under investigation and various projects are available to study these and other aspects of Arabidopsis Rab GTPase function using biochemical, imaging, or informatics approaches. Potential projects include:

(i) 1. Proteomic identification of membrane domains defined by Rab GTPases. This would involve the use of Rab GTPases as affinity-tags to purify specific subpopulations of membrane vesicles for quantitative mass spectrometry followed by the characterisation of candidates with potentially important roles in membrane identity or function.

(ii) 2. Characterisation of Rab GTPase regulatory interactions using in vivo affinity methods and in silico interaction-databases. This would involve bioinformatic analysis plus construction and evaluation of c-terminal affinity-tagged Rab GTPases (wild-type and GTPase deficient mutants) in planta followed by molecular characterisation of interacting proteins or complexes.

(iii) 3. Genetic analysis of Rab GTPase regulatory interactions. This would involve the development of mutant screens for enhancers and suppressors of phenotypes associated with Rab GTPase dysfunction in Arabidopsis. This would be followed by identification of the mutant loci and molecular characterisation of the gene product.

Recent Publications

Au, K.K.-C, Pérez-Gómez, J., Neto, H., Müller, C., Meyer, A.J., Fricker, M.D., and Moore, I., (2012) A perturbation in glutathione biosynthesis disrupts endoplasmic reticulum morphology and secretory membrane traffic in Arabidopsis thaliana. Plant Journal 71 (6): pp881-894. doi: 10.1111/j.1365-313X.2012.05022.x

Boutte, Y, Frescatada-Rosa, M, Men, S, Chow, C.-M, Ebine, K, Gustavsson, A, Johansson, L, Ueda, T, Moore, I, Jurgens, G, Grebe, M. (2010) Endocytosis restricts Arabidopsis KNOLLE syntaxin to the cell division plane during late cytokinesis EMBO Journal. 29 (3): pp 546-558. doi:10.1038/emboj.2009.363.

Camacho, L, Smertenko, A.P, Perez-Gomez, J, Hussey, P.J, Moore, I. (2009) Arabidopsis Rab-E GTPases exhibit a novel interaction with a plasma-membrane phosphatidylinositol-4-phosphate 5-kinase Journal of Cell Science. 122 (23): pp 4383-4392. doi:10.1242/jcs.053488.

Pinheiro, H, Samalova, M, Geidner, N, Chory, J, Martinez, A, Moore, I. (2009) Genetic evidence that the higher plant Rab-D1 and Rab-D2 GTPases exhibit distinct but overlapping interactions in the early secretory pathway Journal of Cell Science. 122 (20): pp 3749-3758. doi:10.1242/jcs.050625.

Chow, CM, Neto, H, Foucart, C, Moore, I. (2008) Rab-A2 and Rab-A3 GTPases define a trans-golgi endosomal membrane domain in Arabidopsis that contributes substantially to the cell plate. Plant Cell. 20 (1): pp 101-23. doi:10.1105/tpc.107.052001.

Samalova, M, Fricker, M, Moore, I. (2008) Quantitative and Qualitative Analysis of Plant Membrane Traffic Using Fluorescent Proteins Methods in Cell Biology. 85: pp 353-380. doi:Sullivan K.F..

Woollard, A.A, Moore, I. (2008) The functions of Rab GTPases in plant membrane traffic Current Opinion in Plant Biology.. doi:10.1016/j.pbi.2008.09.010.

Teh, O.-K, Moore, I. (2007) An ARF-GEF acting at the Golgi and in selective endocytosis in polarized plant cells Nature. 448 (7152): pp 493-496. doi:10.1038/nature06023

Funding Status

BBSRC via the Oxford Interdisciplinary Bioscience Doctoral Training Partnership (DTP) programme. See here for further details: http://www.biodtp.ox.ac.uk or Clarendon Fund - see here: http://www.clarendon.ox.ac.uk/about or via independent funding.

Developmental genetics of plant soil interface and food security

Prof. L Dolan

email: liam.dolan@plants.ox.ac.uk
Tel: +44 (0)1865 275000

Project Description

The evolution of cells at the plant soil interface

Filamentous rooting cells form at the interface between land plants and the soil where they are responsible for nutrient and water uptake. To define the genetic mechanism controlling the development of these cells in the earliest land plants we (1) sequenced the liverwort Marchantia polymorpha genome (2) initiated a genetic screen to identify mutants with defective rhizoid development and (3) carried out a transcriptomic analysis of rhizoid development. The first aim of the studentship will be to use these resources to characterise the function of genes that control rhizoid development and function in M. polymorpha. The second aim will be to determine how the function of these genes has changed during land plant evolution. The third aim will be to asses if any of these genes can be used to enhance nutrient uptake in crops.

Recent Publications

1. Jang G, Pires N, Keke Y, Menand B, Dolan L 2011 RSL genes are sufficient for rhizoid system development in early diverging land plants Development 138, 2273-2281

2. Pernas-Ochoa M, Ryan E, Dolan L 2010 SCHIZORHIZA controls tissue system complexity in plants. Current Biology 20, 818-823

3. Keke Y, Bell E, Menand B, Dolan L 2010 A basic helix loop helix transcription factor controls cell growth and size in root hairs Nature Genetics 42, 264-267

4. Duque Pires N and Dolan L 2010 Origin and diversification of basic helix-loop helix proteins in plants. Molecular Biology and Evolution 27, 862-874

5. Takeda S, Gapper C, Kaya H, Bell E, Kuchitsu K, Dolan L 2008 Local positive feedback regulation determines cell shape in root hair cells Science 319, 1241-1243

Funding Status

BBSRC via the Oxford Interdisciplinary Bioscience Doctoral Training Partnership (DTP) programme. See here for further details: http://www.biodtp.ox.ac.uk or Clarendon Fund - see here: http://www.clarendon.ox.ac.uk/about or via independent funding.

Modulating root hair growth to enhance uptake of nutrients in crops

Prof. L Dolan

email: liam.dolan@plants.ox.ac.uk
Tel: +44 (0)1865 275000

Project Description

Modulating root hair growth to enhance uptake of nutrients in crops

Root hairs play critical roles in the uptake of phosphate from the soil and we discovered that the transcription factor, RSL4, is a master regulator of root hair growth. Using RSL4 we developed a technology that allows us to modulate the growth of root hairs. We hypothesised that engineering plants with longer root hairs would enhance the ability of plants to access phosphate and increase yields on soils with small amounts of available phosphate. A preliminary study demonstrates that yields on these plants are up to 100 times longer than wild type. This studentship will develop RSL4 technology and a novel non-GM technology to develop crops with enhanced nutrient uptake.

Recent Publications

1. Jang G, Pires N, Keke Y, Menand B, Dolan L 2011 RSL genes are sufficient for rhizoid system development in early diverging land plants Development 138, 2273-2281

2. Pernas-Ochoa M, Ryan E, Dolan L 2010 SCHIZORHIZA controls tissue system complexity in plants. Current Biology 20, 818-823

3. Keke Y, Bell E, Menand B, Dolan L 2010 A basic helix loop helix transcription factor controls cell growth and size in root hairs Nature Genetics 42, 264-267

4. Duque Pires N and Dolan L 2010 Origin and diversification of basic helix-loop helix proteins in plants. Molecular Biology and Evolution 27, 862-874

5. Takeda S, Gapper C, Kaya H, Bell E, Kuchitsu K, Dolan L 2008 Local positive feedback regulation determines cell shape in root hair cells Science 319, 1241-1243

Funding Status

BBSRC via the Oxford Interdisciplinary Bioscience Doctoral Training Partnership (DTP) programme. See here for further details: http://www.biodtp.ox.ac.uk or Clarendon Fund - see here: http://www.clarendon.ox.ac.uk/about or via independent funding.

The role of genome organisation in gene expression regulation

Dr S Kelly

email: steven.kelly@plants.ox.ac.uk
Tel: +44 (0)1865 275000

Project Description

This project focuses on the relationship between genes and genomes. In particular, how the structure and organisation of a plant genome controls and regulates the genes that it contains. Through the advent of DNA sequencing technologies we know much about the linear sequence of genomes from many organisms. However, we know relatively little about how these genomes are organised within the 3D volume of a cell nucleus and how that organisation influences phenotype. The aim of this project is to uncover how genome organisation within the plant cell nucleus influences the way in which plant genes are expressed. Understanding how 3D organisation influences gene expression will be critical for genetic engineering of globally important crop plants in the future.

Effective techniques for capturing the 3D genome organisation of chromosomes within plant cell nuclei have recently been developed and they have revealed, like in humans and yeast, that 3D genome organisation plays a critical role in controlling how genes are expressed. Currently technological limitations make studying large genomes such as those of globally important crop plants like maize and wheat prohibitively expensive. However, the model plant Arabidopsis thaliana has a much smaller genome (e.g. ~1/20th the size of maize or wheat) that can be used to begin to determine the rules by which genome organisation regulates gene expression in plant cells. Given the small size of the genome, the ease of cell culture and the extensive public resources of data, arabidopsis is an ideal system to investigate how dynamic 3D genome organisation effects the regulation of gene expression in plants on a genome wide scale. This project will combine both molecular and computational approaches.

Recent Publications

[1] Fransz, P., de Jong, H. 2011 From nucleosome to chromosome: a dynamic organization of genetic information. Plant J 66: 4-17.

[2] Louwers, M., Bader, R., Haring, M., van Driel, R., de Laat, W., Stam, M. 2009 Tissue- and expression level-specific chromatin looping at maize b1 epialleles. Plant Cell 21: 832-842.

[3] Louwers, M., Splinter, E., van Driel, R., de Laat, W., Stam, M. 2009 Studying physical chromatin interactions in plants using Chromosome Conformation Capture (3C). Nat Protoc 4: 1216-1229.

[4] LiebermanAiden, E., van Berkum, N. L., Williams, L., Imakaev, M., Ragoczy, T., Telling, A., Amit, I., Lajoie, B. R., Sabo, P. J., Dorschner, M. O., et al. 2009 Comprehensive mapping of long-range interactions reveals folding principles of the human genome. Science 326: 289-293.

Funding Status

BBSRC via the Oxford Interdisciplinary Bioscience Doctoral Training Partnership (DTP) programme. See here for further details: http://www.biodtp.ox.ac.uk or Clarendon Fund - see here: http://www.clarendon.ox.ac.uk/about or via independent funding.

Plant species diversity and land use in agricultural landscapes

Dr SA Harris

email: stephen.harris@plants.ox.ac.uk
Tel: +44 (0)1865 275000

Project Description

Global biodiversity hotspots have been crudely and ambiguously defined using endemicity and threat. However, within non-global hotspot areas it is likely local diversity hotspots exist. To detect such areas it is necessary to use high resolution, practical approaches where data can be collected rapidly. Furthermore, methodologies must be rigorous and open to peer-review. Such a methodology, the Rapid Botanic Survey (RBS), has been developed for use in diverse, tropical ecosystems but has not been investigated extensively in temperate ecosystems. RBS is a field survey methodology designed for mapping, prioritising and conserving plant species and the vegetation they constitute, whilst integrating species and community assessments. RBS data can also be used for determining the main patterns of floristic variation in plant communities across a landscape and for measuring bioquality, the degree to which a sample represents a biodiversity hotspot.

Agriculture has transformed the lowland British landscape from an environment dominated by forests to a heterogenous mixture of habitats ranging through woodlands and hedgerows to grasslands and wetlands. During the past millennium, change has been episodic and often associated with social and political circumstances. One such episode occured over the past five decades, as agriculture intensified and management practises changed in response to social, political and economic drivers. One consequence of these changes has been as decline in the diversities and ranges of plant and animal species found in agricultural landscapes. However, changes in plant and animal diversity as a consequence of farming are not unique to Britain. Globally, farming has had a greater impact on biological diversity than any other human activity. Consequently, considerable debate has arisen around the concepts of land-sharing versus land-sparing for conservation purposes. Land-sharing, which is the basis of European agri-environment policies, aims to make existing farmland as hospitable as possible to as many species as possible. In contrast, under the land-sparing model production is concentrated into particular areas and unmodified habitat spared from future modification.

Ditchley Park, a heterogenous, agricultural landscape, with a long management history, would be an excellent model in which to test the applicability of RBS methodologies, investigate localised biodiversity hotspots and contribute to the land-sharing-land-sparing debate. The data will enable investigation of British plant biodiversity at a range of scales. From single plant communities within Ditchley Park, through Ditchley Park in the context of the Oxfordshire landscape to Oxfordshire in the context of Britain and the British flora in a global context. Importantly, these data would enable assessment of not only how many species but which species are being maintained.

Predictive modelling of biodiversity effects on ecosystem functions

Dr SA Harris

email: stephen.harris@plants.ox.ac.uk
Tel: +44 (0)1865 275000

Project Description

* Jointly supervised by: Prof A Hector, Dr S Harris & Dr L Turnbull *

NOTE: Must start by 1st March 2013.

Loss of biodiversity normally has a detrimental effect on ecosystem functions but the underlying biological mechanisms are not well understood limiting our ability for prediction. We will develop predictive models of resource uptake and plant interactions that can be parameterized with data from some of the best known biodiversity experiments in grasslands (Univ. Minnesota) and tropical forest (Sabah Biodiversity Experiment). This work will lead to a better understanding of the number of species required to maintain essential ecosystem functions and how and why species differ in their impact.

The project will involve developing and analysing quantitative models (in R). The behaviour of these models can be explored theoretically but they can also be parameterized using data from field experiments. Much of the required data is already available but will require processing while any additional data collection will involve field work at the sites of the grassland and tropical forest biodiversity experiments (www.SabahBiodiversityExperiment.org). The project provides the opportunity to work and collaborate with an outstanding international team of ecologists.

The successful candidate will be independent and self-motivated and have strong quantitative skills (both statistical and computational). Experience of practical research, particularly with plants, and knowledge of R will be an advantage.

Recent Publications

Hector, A. and R. Bagchi. 2007. Biodiversity and ecosystem multifunctionality. Nature 448:188-190.

Hector, A., C. Philipson, P. Saner, J. Chamagne, D. Dzulkifli, M. O'Brien, J. L. Snaddon, P. Ulok, M. Weilenmann, G. Reynolds, and H. C. J. Godfray. 2011. The Sabah Biodiversity Experiment: A long-term test of the role of tree diversity in restoring tropical forest structure and functioning. The Philosophical Transactions of the Royal Society B 366:3303-3315.

Isbell, F., V. Calcagno, A. Hector, J. Connolly, W. S. Harpole, P. B. Reich, M. Scherer-Lorenzen, B. Schmid, D. Tilman, J. van Ruijven, A. Weigelt, B. J. Wilsey, E. S. Zavaleta, and M. Loreau. 2011. High plant diversity is needed to maintain ecosystem services. Nature 477:199-202.

Turnbull, L. A., J. Levine, M. Loreau, and A. Hector. In review. Coexistence, niches, and the effects of biodiversity on ecosystem functioning. Ecology Letters (contact us for a preprint).

Funding Status

Funded by a NERC studentship. Must start by 1st March 2013