Analysis of plant mitochondrial metabolism using NMR and mass spectrometry

Project Description

The pivotal roles of mitochondria in respiration and biosynthesis require an extensive suite of enzymes to support a network of mitochondrial reactions that is itself embedded in the wider metabolic network of the plant cell. Stable isotope labelling provides a powerful approach for probing the operation of these pathways, both at the level of establishing the occurrence of specific metabolic steps and at the level of quantifying the metabolic fluxes that they support. Typically these tasks are achieved using 13C- or 15N-labelled precursors with either nuclear magnetic resonance (NMR) spectroscopy or mass spectrometry (MS) as the detection technique.

We have developed two distinct experimental approaches for analysing plant mitochondrial metabolism using stable isotopes. In the first approach we use NMR to observe metabolic events non-invasively in dilute suspensions of isolated mitochondria in defined respiratory states. This versatile method gives the experimentalist many degrees of freedom, since the mitochondria can be easily studied under a wide range of conditions, and we use this method to study the contribution of particular steps or pathways to mitochondrial metabolism, and also to investigate substrate channelling. In the second approach we use NMR and gas chromatography-MS to analyse the redistribution of label in steady-state experiments on Arabidopsis cell suspensions. This allows us to produce flux maps that reveal the links between the mitochondria and the rest of central metabolism.

There is considerable scope for further technical development of these powerful approaches, and the focus of the project will be to explore the analytical options for increased sensitivity and throughput. The resulting methods will then be used to characterise poorly understood mitochondrial processes, such as the dynamic association of hexokinase with the outer mitochondrial membrane or the conditions under which the tricarboxylic acid cycle becomes non-cyclic.

Recent Publications

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

N.J. Kruger, M.A. Troncoso-Ponce and R.G. Ratcliffe (2008) 1H-NMR metabolite fingerprinting and metabolomic analysis of perchloric acid extracts from plant tissues. Nature Protocols 3, 1001-1012.

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.

L.J. Sweetlove, K.F.M. Beard, A. Nunes-Nesi, A.R. Fernie and R.G. Ratcliffe (2010) Not just a circle: flux modes in the plant TCA cycle. Trends in Plant Science 15, 462-470.

L.J. Sweetlove and R.G. Ratcliffe (2011) Flux-balance modelling of plant metabolism. Frontiers in Plant Science 2:38. doi: 10.3389/fpls.2011.00038

Evolution of metabolic regulation in plants

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 (FKP; fructose-6-phosphate 2-kinase, fructose-2,6-bisphosphatase). All higher plants have one or more FKP 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 FKP isozymes in Fru-2,6-P2 metabolism, (ii) the significance of the amino terminal domain in FKP function, and (iii) the role played by this regulatory metabolite in plants through its novel interaction with PFP.

J.E. Markham and N.J. Kruger (2002) Kinetic properties of 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase from spinach leaves. European Journal of Biochemistry 269, 1267-1277.

P.A.M. Michels and D.J. Rigden (2006) Evolutionary analysis of fructose 2,6-bisphosphate metabolism. IUBMB Life 58, 133 – 141.

Recent Publications

N.J. Kruger, M.A. Troncoso-Ponce and R.G. Ratcliffe (2008) 1H-NMR metabolite fingerprinting and metabolomic analysis of perchloric acid extracts from plant tissues. Nature Protocols 3, 1001-1012.

T.C.R. Williams, L. Miguet, S.K. Masakapalli, N.J. Kruger, L.J. Sweetlove and R.G. Ratcliffe (2008) Metabolic network fluxes in heterotrophic Arabidopsis cells: stability of the flux distribution under different oxygenation conditions. Plant Physiology 148, 704-718.

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, 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.P. Howard et al. (2011) Antisense suppression of the small chloroplast protein CP12 in tobacco alters carbon partitioning and severely restricts growth Plant Physiology (in press); doi:10.1104/pp.111.183806.

Metabolic flux analysis of Rhizobium leguminosarum

Project Description

Nitrogen fixation by legumes plays an important role in sustainable agriculture and the global nitrogen cycle. The process depends on the establishment of 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 assimilation, 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, and in particular it requires measurements of the metabolic fluxes supported by the bacterial and plant cell metabolic networks. To address this problem, we intend to develop a novel strategy for analyzing cell-specific metabolism based on stable isotope labelling of cell-specific marker proteins.

The project builds on our work in the field of steady-state metabolic flux analysis (MFA). Recently we have extended the scope of this method by using GFP-binding nanobodies, derived from a heterologous expression system, to purify GFP from the roots of transgenic Arabidopsis thaliana plants that express GFP in specific cell types. This will allow us to use GFP as a marker protein to interrogate the metabolic state of specific cell types following incubation with 13C-labelled substrates.

The challenge is to 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 for primary metabolism for the bacteroid within its host cell for the first time.

Recent Publications

N.J. Kruger, M.A. Troncoso-Ponce and R.G. Ratcliffe (2008) 1H-NMR metabolite fingerprinting and metabolomic analysis of perchloric acid extracts from plant tissues. Nature Protocols 3, 1001-1012.

T.C.R. Williams, L. Miguet, S.K. Masakapalli, N.J. Kruger, L.J. Sweetlove and R.G. Ratcliffe (2008) Metabolic network fluxes in heterotrophic Arabidopsis cells: stability of the flux distribution under different oxygenation conditions. Plant Physiology 148, 704-718.

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, 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.

An interdisciplinary approach to understand development and evolution of leaf shape

Project Description

Two key challenges in biology are to understand how biological forms are generated, and to elucidate the basis for their diversity. Addressing these challenges is difficult because bioscience methodologies are insufficient for conceptualizing how complex biological shapes are generated. This is because final form is produced by a cascade of developmental processes that include complex feedback loops of genetic regulation, signaling, cell division and tissue growth, which take place at different levels of organization. The aim of this project is to use a combination of computational modelling and developmental genetics to address these problems in the case of divergent leaf forms of seed plants. We will focus on the dramatic difference in leaf shape between A.thaliana a model organism with simple leaves and its relative C.hirsuta which has compound leaves divided to leaflets. Other than being taxonomically related these plants show exceptional genetic tractability thus providing excellent opportunities for comparative studies elucidating the basis of morphological diversity. We will compare leaf ontogenesis and growth in mutants, natural variants and transgenic lines of these two species to produce predictive models that capture essential aspects of leaf development and diversity. Modeling methods will be advanced as needed for these analyses. As such this is an interdisciplinary project situated at the interface between evolutionary developmental biology and computational modeling. Computational aspects of the project will be co-supervised by Prof Przemyslaw Prusinkiewicz (Univ of Calgary) and Dr Yiannis Ventikos (Univ of Oxford Dept. of Engineering) and the project will run within the context of an HFSP funded grant to MT and PP. This project will suit a biologist with expertise in computational programming.

Recent Publications

Bilsborough, G., Runions, A., Barkoulas, M., Jenkins, H., Hasson, A., Galinha, C., Laufs, P., Hay, A., Prusinkiewicz, P. and Tsiantis, M. Model for the regulation of Arabidopsis thaliana leaf margin development PNAS. 108, 3424-3429

Piazza, P., Bailey, C.D., Cartolano, M., Krieger, J., Cao, J., Ossowski, S., Schneeberger, K., Hall, N., Fei He, Juliette de Meaux, MacLeod, N., Filatov D., Hay, A. and Tsiantis, M. (2010). Arabidopsis thaliana leaf form evolved via loss of KNOX expression in leaves in association with a selective sweep. Current Biology 20, 2223-2228.

Grigg, S., Galinha, C., Kornet N, Canales, C., Scheres, B. and Tsiantis, M. (2009). Repression of apical HD-ZIP III homeobox genes is required for Arabidopsis embryonic root development. Current Biology,

Barkoulas, M. Hay, A. Kouyioumoutzi, E and Tsiantis, M. (2008). “A developmental framework for dissected leaf formation in the Arabidopsis relative Cardamine hirsuta.” Nature Genetics 40, 1136 – 1141.

Hay, A. and Tsiantis, M. (2006). The genetic basis for differences in leaf form between Arabidopsis and its wild relative Cardamine hirsuta. Nature Genetics 38, 942-7.

Diversification of gene regulatory networks sculpting leaf shape

Project Description

A key problem in biology is to understand how diversity in organismal form is generated. We study this problem by investigating the genetic mechanisms underlying variation in form of the predominant photosynthetic organ of plants, the leaf. Leaf form can be classified as simple, where the leaf blade is entire, or dissected where the blade is divided into distinct units called leaflets. Mechanisms that specify dissected versus entire leaf shape and regulate the number, position and timing of leaflet production are poorly understood and are the focus of this project.

We will study leaflet production in the small mustard plant Cardamine hirsuta a species that we recently developed into a powerful genetic system suitable for studying for studying evolution of form. One advantage of C.hirsuta is that it is closely related to the model organism Arabidopsis thaliana that bears entire leaves. Thus, the availability of two related genetically tractable species that differ in leaf shape allows us to perform in depth functional studies to understand the precise regulatory changes that result in dissected versus simple leaf form. Experimental work will involve characterisation of C.hirsuta mutants that show perturbed leaflet formation, comparative functional studies of leaf patterning genes in A.thaliana and C.hirsuta. The project will provide training in developmental genetics and Molecular Biology. These experiments will help understand how leaf development programmes have been reconfigured to result in the dissected leaf form. This will be an important step in understanding how distinct leaf forms arise in Nature. A BBSRC funded project is running on this topic which will create opportunities for synergy within the lab.

Recent Publications

Bilsborough, G., Runions, A., Barkoulas, M., Jenkins, H., Hasson, A., Galinha, C., Laufs, P., Hay, A., Prusinkiewicz, P. and Tsiantis, M. Model for the regulation of Arabidopsis thaliana leaf margin development PNAS. 108, 3424-3429

Piazza, P., Bailey, C.D., Cartolano, M., Krieger, J., Cao, J., Ossowski, S., Schneeberger, K., Hall, N., Fei He, Juliette de Meaux, MacLeod, N., Filatov D., Hay, A. and Tsiantis, M. (2010). Arabidopsis thaliana leaf form evolved via loss of KNOX expression in leaves in association with a selective sweep. Current Biology 20, 2223-2228.

Grigg, S., Galinha, C., Kornet N, Canales, C., Scheres, B. and Tsiantis, M. (2009). Repression of apical HD-ZIP III homeobox genes is required for Arabidopsis embryonic root development. Current Biology,

Barkoulas, M. Hay, A. Kouyioumoutzi, E and Tsiantis, M. (2008). “A developmental framework for dissected leaf formation in the Arabidopsis relative Cardamine hirsuta.” Nature Genetics 40, 1136 – 1141.

Hay, A. and Tsiantis, M. (2006). The genetic basis for differences in leaf form between Arabidopsis and its wild relative Cardamine hirsuta. Nature Genetics 38, 942-7.

Evolution of gene regulation as driving force for morphological change in A. thaliana relatives

Project Description

A key problem in biology is to understand how diversity in organismal form is generated. This project will seek to use function-based assays informed by genomics approaches to identify genes that contributed to diversity of leaf forms in relatives of the model plant A. thaliana. The project will provide training in genetics molecular biology and would suit students with an interest in deciphering the mechanistic basis of how morphology evolves. A BBSRC funded project is running on this topic which will create opportunities for synergy within the lab.

Recent Publications

Bilsborough, G., Runions, A., Barkoulas, M., Jenkins, H., Hasson, A., Galinha, C., Laufs, P., Hay, A., Prusinkiewicz, P. and Tsiantis, M. Model for the regulation of Arabidopsis thaliana leaf margin development PNAS. 108, 3424-3429

Piazza, P., Bailey, C.D., Cartolano, M., Krieger, J., Cao, J., Ossowski, S., Schneeberger, K., Hall, N., Fei He, Juliette de Meaux, MacLeod, N., Filatov D., Hay, A. and Tsiantis, M. (2010). Arabidopsis thaliana leaf form evolved via loss of KNOX expression in leaves in association with a selective sweep. Current Biology 20, 2223-2228.

Grigg, S., Galinha, C., Kornet N, Canales, C., Scheres, B. and Tsiantis, M. (2009). Repression of apical HD-ZIP III homeobox genes is required for Arabidopsis embryonic root development. Current Biology,

Barkoulas, M. Hay, A. Kouyioumoutzi, E and Tsiantis, M. (2008). “A developmental framework for dissected leaf formation in the Arabidopsis relative Cardamine hirsuta.” Nature Genetics 40, 1136 – 1141.

Hay, A. and Tsiantis, M. (2006). The genetic basis for differences in leaf form between Arabidopsis and its wild relative Cardamine hirsuta. Nature Genetics 38, 942-7.

Genetic basis of petal loss in Cardamine hirsuta

Project Description

This project aims to investigate the genetic control of divergent petal number between the model organism Arabidopsis thaliana and its close relative Cardamine hirsuta. A. thaliana has a typical mustard flower with four petals while C. hirsuta differs by having fewer petals. Recessive mutants have been isolated in C. hirsuta that are sufficient to produce flowers with four petals. The objectives of this project are to identify the genes responsible for the four petal mutant phenotypes in C. hirsuta and to determine whether their function is conserved between C. hirsuta and A. thaliana or specific to C. hirsuta.

Recent Publications

McKim, S and Hay, A. (2010) Patterning and evolution of floral structures – marking time. Current Opinion in Genetics & Development, 20(4): 448-453

P Piazza, C Bailey, M Cartolano, J Krieger, J Cao, S Ossowski, K Schneeberger, F He, J de Meaux, N Hall, N MacLeod, D Filatov, A Hay and M Tsiantis (2010) Arabidopsis thaliana Leaf Form Evolved via Loss of KNOX Expression in Leaves in Association with a Selective Sweep. Current Biology, 20(24): 2223-8

Molecular Cell biology: The evolution of endocytic and secretory pathways in plants

Project Description

We are interested in understanding the molecular organisation and evolution of membrane trafficking mechanisms in plant cells. Comparative genomics and cell biology suggest that secretory and endocytic membrane trafficking pathways diversified independently in the evolution of multicellular plants and animals (2,4). Work in my group has provided some of the first experimental support for this hypothesis (1-4).. The unique biosynthetic and trafficking functions of these pathways are responsible for some of the most biologically and socially important features of plant cells including the establishment of complex cell polarity, cell wall biogenesis, and cytokinesis.

We study a group of important regulatory GTPases of the Rab family. Rab proteins catalyse the assembly of specific macromolecular complexes that give individual membrane domains a distinct identity and orchestrate specific vesicle transport steps. One class of Rab GTPase involved in post-Golgi secretion, endocytic sorting, and cytokinesis (1-3) has diversified enormously during land plant evolution and the hypothesis is that this diversification accompanied the evolution of plant-specific membrane sorting events. To test this hypothesis a major aim of the group is to identify the molecular targets and regulators of these Rab GTPases.

Projects may involve the identification and characterisation of novel interactors or a study of the evolution of these proteins by characterising their paralogues in basal-branching land plants such as the liverwort Marchantia polymorpha. Projects will provide extensive training in molecular biology, protein biochemistry, genetics, and live-cell microscopy.

We have also completed genetic screens for Arabidopsis mutants that are defective in secretory membrane traffic (4,5). The screens are based on the use of ratiometric GFP reporters that were developed in my lab. These mutants may define novel plant-specific aspects of membrane traffic. Over 50 such mutants, many of which are seedling lethal, are assigned to chromosomes and are available for identification and characterisation.

Recent Publications

Camacho, L, Smertenko, A., Hussey, P.J, Moore, I. (2009) Arabidopsis Rab-E GTPases exhibit a novel interaction with a plasma-membrane phosphatidylinositol-4-phosphate 5-kinase. J. Cell Sci. 122; 4383-4392.

Woollard, A.A, Moore, I. (2008) The functions of Rab GTPases in plant membrane traffic Current Opinion in Plant Biology.

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.

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.

Rico, A., Bennett, M.H., Forcat, S., Huang, W.E., and Preston, G.M. (2010). Agroinfiltration reduces ABA levels and suppresses Pseudomonas syringae-elicited salicylic acid production in Nicotiana tabacum. PLoS ONE 5, e8977.

Proteomic analysis of plant membrane trafficking pathways in health and disease

Project Description

The unique biosynthetic and trafficking functions of these pathways are responsible for some of the most biologically and socially important features of plant cells including the establishment of complex cell polarity, cell wall biogenesis, cytokinesis, and pathogen defense. Comparative genomics and cell biology suggest that secretory and endocytic membrane trafficking pathways diversified independently in the evolution of multicellular plants and animals (2,4). Our work has provided some of the first experimental support for this hypothesis (1-4).

We study a group of important regulatory GTPases of the Rab family. Rab proteins catalyse the assembly of specific macromolecular complexes that give individual membrane domains a distinct identity and orchestrate specific vesicle transport steps. Some Rab GTPases involved in post-Golgi secretion, endocytic sorting, and cytokinesis (1-3) exhibit novel molecular interactions (1) while others have diversified enormously during land plant evolution. The hypothesis is that this molecular evolution accompanied the evolution of plant-specific membrane sorting events associated with processes such as cytokinesis, cell polarity, and disease resistance. To test this hypothesis a major aim of the group is to identify the trafficking pathways and molecular cargoes that are regulated by the individual Rab GTPase subclasses.

In association with a recently funded BBSRC research project, the aim is to apply proteomic methods to characterise the secreted and plasma-membrane proteomes of plants expressing inhibitory mutants of various Rab GTPase subclasses. We will also ask how each Rab pathway affects the secretory response to various pathogen-associated elicitors in the root and shoot system.

Recent Publications

Camacho, L, Smertenko, A., Hussey, P.J, Moore, I. (2009) Arabidopsis Rab-E GTPases exhibit a novel interaction with a plasma-membrane phosphatidylinositol-4-phosphate 5-kinase. J. Cell Sci. 122; 4383-4392.

Woollard, A.A, Moore, I. (2008) The functions of Rab GTPases in plant membrane traffic Current Opinion in Plant Biology.

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.

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.

Rico, A., Bennett, M.H., Forcat, S., Huang, W.E., and Preston, G.M. (2010). Agroinfiltration reduces ABA levels and suppresses Pseudomonas syringae-elicited salicylic acid production in Nicotiana tabacum. PLoS ONE 5, e8977.

Rooting systems and food security

Project Description

Three projects are available in our lab to study different aspects of root biology. The overarching aims of our research are to discover fundamental mechanisms controlling root development and evolution and to apply this knowledge to increase food security.

1 Rooting system evolution The invasion of the land was accompanied by the evolution of rooting systems that anchored plants to their substratum and facilitated the uptake of growth-limiting nutrients from soil. We are using a variety of approaches including genetics, genome sequencing and RNA seq. to identify and characterize the mechanisms that responsible for the diversity of rooting structures in plants. This is important because the results of this research will demonstrate the mechanisms that underpinned root system evolution in land plants. This was critical to the invasion of the land 500 million years ago, one of the pivotal events in Earth history.

2 Cell growth in plants Transcription factors control the growth of root hairs. RSL4 is a basic helix loop helix transcription factor that is necessary and sufficient for root hair growth. The laboratory is currently characterizing the mechanisms of RSL4-regualted growth with emphasis on the process that stops growth. We are using genetics to identify genes that tell root hairs when to stop growing. Mutations in these genes result in the constitutive growth of root hairs. That is, the root hairs do not stop growing. A project is available to characterize the role of one or a few of these genes. This project is important because the results provide insight into fundamental mechanisms of cell growth in eukaryotes.

3 Enhancing nutrient uptake in plants The world population will increase by over 2 billion in the next 40 years and food production will need to increase to supply these people with a sufficient diet. Increasing the efficiency of nutrient uptake and fertilizer-use will be critical for the sustainable intensification of agriculture. We have developed technologies that will enhance nutrient uptake in crops. This project aims to develop a combination of technologies to increase nutrient uptake in rice and wheat.

Recent Publications

Jang G, Yi K, Pires ND, Menand B, Dolan L (2011) RSL genes are sufficient for thizoids system development in early diverging land plants Development 128, 2273-2281

Yi K, Menand B, Bell E, Dolan L (2010) A basic helix loop helix transcription factor controls cell growth and size in root hairs Nature Genetics 42, 264-267

Menand B, Yi K, Jouannic S, Hoffmann L, Ryan E, Linstead P, Schaefer DG, Dolan L (2007) An ancient mechanisms controls the development of cells with a rooting function in land plants Science 316, 1477-1480

Evolution of sex chromosomes in diploids & haploids

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. Y chromosome degeneration is thought to occur due to heterozygosity of the Y-chromosome and genetic hitchhiking that reduce its effective population size. The smaller the effective population size, the larger is the chance of fixation of deleterious mutations. Purifying selection is not effective on the Y chromosome due to reduced population size, which leads to accumulation of deleterious alleles and gradual loss of function of Y-linked genes. The PhD project is devoted to testing various aspects of this theory. It will use dioecious plant Silene latifolia that evolved sex chromosomes quite recently and provides a convenient model system to study early stages of sex chromosome evolution in diploid species. Another part of this project will be devoted to the study of sex chromosome evolution in haploid fungus Microbotryum. Perhaps the most obvious difference between the species with haploid and diploid sex determination is that only Y chromosome is sex-specific and non-recombining in diploid species, while both sex chromosomes are sex-specific and non recombining in the species with haploid sex determination. Another important difference is that only one copy of each chromosome is present in haploids, thus, loss of genes from nonrecombining Y in haploid species is likely to be more deleterious than in diploid species where genes are “sheltered” by the functional copies on the X chromosome. This has led to a view that genetic degeneration on sex chromosomes may not occur in haploid species (Bull 1983), which will be tested in course of this project.

Recent Publications

Chibalina MV, Filatov DA. 2011 Plant Y chromosome degeneration is retarded by haploid purifying selection. Current Biology 17: 1475-1479.

Votintseva AA, Filatov DA. 2009 Evolutionary strata in a small mating-type-specific region of the smut fungus Microbotryum violaceum. Genetics. 182:1391-1396.

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

Armstrong SJ and Filatov DA. 2008 A cytogenetic view of sex chromosome evolution in plants.Cytogenet Genome Res.120:241-246

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

Foundation Monograph of Ipomoea

Project Description

Estimates of the number of described species of flowering plant range between 220,000 and 420,000. The discrepancy between these numbers is mainly caused by unknown levels of duplicate names (synonyms). In order to compile a more accurate list of described species of flowering plant - an important aim for conservation, measures of extinction and levels of biodiversity - duplicate names need to be identified. Furthermore, it is estimated that there are 70,000 species of flowering plant still to be discovered but at least half of these have already been collected and await description in the world's herbaria. These issues of synonymy and unidentified new species are most prevalent in large tropical genera that have not been monographed from a global perspective. The volume of specimens and dispersed literature associated with these groups of plants, make the task very daunting. This project seeks to combine expertise, smart technology with a bold vision to devise and develop methods to tackle these groups in a relatively short timeframe. Specifically, we will develop a new method to accelarate the rate at which the taxonomy of large problematic groups of tropical plants can be improved in a relatively short period of time. Specifically, we propose to produce a foundation monograph of Ipomoea, a large genus of tropical plants that has never been tackled from a global perspective.

Recent Publications

Bebber, D.P. Carine, M.A.Wood, J.R.I. Wortley, A.H. Harris, D.J. Prance, G.T. Davidse, G. Paige, J. Pennington, T.D. Robson, N.K.B. and Scotland, R.W 2010. Herbaria are a major frontier for species discovery. PNAS. 107 (51): 22169-22171.

Wood JRI & Scotland RW. 2009. New and little known species of Strobilanthes (Acanthaceae) from India and South East Asia. Kew Bulletin 64: 3-47.

Molecular basis of adaptation in plant photosynthesis

Project Description

Photosynthesis is the ultimate source of almost all organic matter on Earth, and the key biochemical reactions in this process are shared by all green plants. Yet plants perform photosynthesis in an astonishing variety of different environments. For example, temperature, light intensity and water availability may differ drastically between hot, exposed desert environments and the deep shade of forest understories. This project will focus on adaptive mechanisms that allow plants to survive and carry out photosynthesis effectively under such a wide range of environmental conditions. The project will involve a number of approaches combining molecular population genetics, evolutionary biology and plant biochemistry focusing on the molecular basis of adaptation in Rubisco, the key enzyme of photosynthetic carbon assimilation. This enzyme is the cornerstone of photosynthesis, being responsible for the conversion of inorganic carbon into organic compounds. As the performance of Rubisco may greatly affect plant growth and hence crop yields, significant efforts have been made to study the structure and function of this enzyme. This project will investigate Rubisco function using a comparative approach, studying the enzyme in C3, C4 and CAM plants, and in their appropriate sister-groups, from a wide range of different habitats. This will create a unique opportunity to combine population-genetic, phylogenetic and biochemical approaches to studying the molecular evolution of this crucial enzyme in photosynthesis.

Recent Publications

West-Eberhard, M.J., Smith, J.A.C. and Winter, K. (2011) Photosynthesis, reorganized. Science 332, 311–312.

Kapralov, M.V., Kubien, D.S., Andersson, I. and Filatov, D.A. (2011) Changes in Rubisco kinetics during the evolution of C4 photosynthesis in Flaveria (Asteraceae) are associated with positive selection on genes encoding the enzyme. Mol. Biol. Evol. 28, 1491–1503.

Kapralov, M.V. and Filatov, D.A. (2007) Widespread positive selection in the photosynthetic Rubisco enzyme. BMC Evol. Biol. 7:73.

Kapralov, M.V. and Filatov, D.A. (2006) Molecular adaptation during adaptive radiation in the Hawaiian endemic genus Schiedea. PLoS ONE 1:e8.

Crayn, D.M., Winter, K. and Smith, J.A.C. (2004) Multiple origins of crassulacean acid metabolism and the epiphytic habitat in the Neotropical family Bromeliaceae. Proc. Natl. Acad. Sci. USA 101, 3703–3708.

The role of coevolution history in explaining contemporary plant-herbivore interactions

Project Description

Current anthropogenic environmental change is causing a rapid loss of biodiversity. Although the effects of the main causes of this loss (e.g. habitat fragmentation, climate change and invasive species) on single species have been widely studied, the impacts on species interactions are poorly understood. In particular, we do not understand how these phenomena affect the ecological and evolutionary processes and mechanisms that impact species interactions.

The diversity of life has not only resulted from the diversification of species, but also from the diversification of interactions among them. Coevolution, the reciprocal evolutionary change of interacting species, is a major force shaping patterns of adaptation and influencing diversification of interacting species and ultimately generating biological diversity. Therefore, it is important to also focus on interactions and their diversity, in particular highly specialised and tightly coevolving interactions, when attempting to conserve biological diversity. Moreover, many of the services that have been identified as indicators of ecosystem integrity are, in fact, the consequences of species interactions, not services provided by a single species acting on its own, for example, pollination, nutrient cycling, decomposition and biocontrol. We still lack an understanding of how contemporary ecological interactions are shaped by their (co)evolutionary history, although this would be crucial for better understanding and conserving species and their interactions.

This project aims at investigating how (co)evolutionary history has shaped interactions between plants from the family Vincetoxicum and their specialist and generalist herbivores. Associations between plant divergence and divergence in secondary chemistry and herbivore assembly will be studied to asses whether (co)evolutionary history explains herbivore community diversity and structure. The study will also include the invasive Vincetoxicum species providing an opportunity to investigate whether we can use the understanding of (co)evolutionary history and dynamics to better target biocontrol of invasive plant species.

Recent Publications

Muola, A., Mutikainen, P., Laukkanen, L., Lilley, M. & Leimu, R. 2010. Genetic variation in herbivore resistance and tolerance: the role of plant life-history and type of damage. Journal of Evolutionary Biology, 23:2185-2196.

Muola, A., Mutikainen, P., Lilley, M., Laukkanen, L., Salminen, J.-P. & Leimu, R. 2010. Associations of plant fitness, leaf chemistry and damage reflect selection mosaic in plant-herbivore interactions. Ecology 91:2650-2659.

Leimu, R., Vergeer, P., Angeloni, F. & Ouborg, N.J. 2010. Habitat fragmentation, climate change, and inbreeding in plants. Annals of the New York Academy of Science. Eds. Ostfeld, R. S. & Schlesinger W. H. The Year in Ecology and Conservation Biology, pp 84-98.

Leimu, R, Kloss, L. and Fischer, M. 2008. Effects of experimental inbreeding on herbivore resistance and plant fitness: the role of history of inbreeding, herbivory and abiotic factors. Ecology Letters, 11:1101-1110.

Leimu, R. & Mutikainen, P. 2005. Population history, mating system and fitness variation in a perennial herb with a fragmented distribution. Conservation Biology, 19:1-8.