Coen Lab - Research

From Flower Development, E Coen , Cell & Developmental Biology Department - JIC UK

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Overview of Research Programme

Our research programme involves collaborations with several labs, including Computer scientists and ecological population geneticists. We study developmental and evolutionary problems at a range of spatial and temporal scales, from subcellular events at minute intervals, to events at geographic scale over millions of years. Our long term goal is to link these different scales of analysis to give an integrated understanding of evolution and development. Each person in the lab has a main project while also working in collaboration with others in the group.

A major focus of our group is on leaf development in Arabidopsis, which has the advantage of good genetics and convenience for imaging. Starting at the subcellular scale, we are analysing the role of microtubules and nuclear movements in the control of leaf cell division and growth. This involves extensive confocal imaging, image processing and computational modelling1,2,3. This work is closely related to studies at the cellular scale on division, growth and differentiation in developing leaves, involving a combination of time-lapse imaging and modelling. At the tissue scale we are studying leaf development through clonal analysis, time-lapse imaging6 and Optical Projection Tomography9. Leaf growth is also being modelled at this scale by treating the leaf as a growing sheet of material8. In addition to these studies on leaf development, we are analysing flower and leaf development using similar approaches in Antirrhinum8,9,10 and in monocots such as barley and maize.

Another major focus of our work is on the evolution of flower colour, using the Antirrhinum species group as a model system. One aspect is to study evolutionary events over a few generations, at a naturally occuring hybrid zone in the Pyrenees. This uses a combination of DNA pedigree, phenotypic and pollinator studies10,11,12,13. We also aim to identify the key genes controlling flower colour that are under selection. On a longer evolutionary timescale we are studying variation in the function and structure of genes controlling flower colour between different Antirrhinum species using a combination of molecular and genetic approaches10,12.

Studies on the evolution of leaf shape are also being extended to carnivorous plants which have elaborate leaves that entice and digest animal prey, allowing them to thrive in nutrient poor environments. Carnivorous cup-shaped leaves with lids are effective animal traps very similar in structure to each other. They are found in four independent lineages; Asian pitcher plants (Nepenthes), Albany pitcher plants (Cephalotus), American pitcher plants (Sarracenia, Darlingtonia and Heliamphora) and bladderworts (Utricularia). This example of convergent evolution might hint at similar developmental mechanisms underlying the growth of these cup-shaped leaves. We are establishing Utricularia gibba as a model carnivorous plant to discover whether developmental rules discovered in Arabidopsis are adapted to grow cup-shaped leaf traps and whether these different plants are using similar genetic mechanisms to grow their traps4,5,7,14 . We are using a combination of genetic analysis, imaging and mathematical modelling to understand how these leaves grow themselves.


1 - Modelling Microtubule Dynamics

Microtubule edge segmentation, orientation colouring and segment joining

Jacob Newman - email in collaboration with Jordi Chan

Microtubules in Arabidopsis leaf cells have been observed to undergo persistent reorientation. Much work has been done to quantify the dynamics and behaviour of microtubules, however little is known about the processes by which inter-microtubule interactions lead to the patterning observed in biological data. This work uses computer modelling software developed by Andrew Bangham and Richard Kennaway (MTtbox) to create a model which can reproduce the ordering and reordering of microtubule arrays, using realistic assumptions regarding microtubule dynamics.

In tandem with the modelling, an image processing project (in collaboration with Paul Southam) seeks to automatically quantify the dynamics and interactions of microtubules in time-lapsed, electron microscopy image data. This quantification serves to support the manual observations by Jordi Chan, to allow the processing of more data than could realistically be analysed by eye, and to extract parameters which will feed directly into our microtubule modelling. I hope to apply these methods to other systems such as Utricularia.

2 - Modelling Dynamic Growth of Cup-Shaped Leaves and Petals

Model of cup-shaped structure.

Jie Cheng - email in collaboration with Chris Whitewoods, Beatriz Gonçalves, Man Yu and John Fozard.

Leaves and petals are generally flat sheet-like structures. However, some groups such as buttercup family (eg. Nigella) and carnivorous plants (eg. Utricularia) have evolved cup-shaped petals and leaves independently. The genetic mechanisms behind this kind of transition is unclear. To seek a general principal, I am using volumetric finite element method to model the dynamic transformation of 2D sheets and 3D cups theoretically. Then, I will adopt the experimental results from 3D imaging and molecular genetics to test our models. I am using the similar strategy to model the tissue outgrowth such as ligules in monocot leaves.

In collaborating with Man Yu and John Fozard, I am using Arabidopsis cotyledon to explore whether external mechanical forces can influence the tissue and cell polarities.

My visiting is supported by the UCAS Joint PhD Training Program and the National Natural Science Foundation of China Grants(91631308).

3 - Microtubule dynamics during growth and division

Pre-prophase band formation in a leaf epidermis cell.

Jordi Chan - email in collaboration with Paul Southam and Jake Newman

Microtubules are highly dynamic subcellular filaments that adopt different patterns of alignment during growth, division and differentiation. During growth, microtubules are found in close association with the plasma-membrane where they are needed to guide the movement of cellulose synthase to generate appropriate wall architectures. Like animal cells, microtubules are required for chromosome segregation during cell division via the spindle apparatus. However, unlike animal cells, plant microtubules bunch up at the onset of mitosis to form a cortical ring, known as the pre-prophase band (PPB), which is involved in determining the future plane of cell division. Since plant cells within tissues are immobilised by their own walls and can only respond to cellular and environmental cues by altering their directions of growth and division planes, microtubules are indispensible elements of plant morphogenesis.

How microtubules change alignment and construct different arrays during growth and cell division is unknown but is likely to reside within fundamental properties of the microtubules themselves, such as the dynamic behaviour of their filamentous ends, regulation of their sites and angles of nucleation (birth) and outcomes of encounters between themselves and the geometry of the cell. It is interesting to think that these simple subcellular behaviours or rules concerning microtubule dynamics may not only shape the cell but also be amplified during morphogenesis from cell-to-cell to shape the entire plant.

The aim of my work is to reconstitute microtubule dynamics and cortical array organisation in-silico to understand and test mechanisms behind microtubule reorientation and division plane alignment (PPB formation) in-planta. This will involve time-lapse microscopy of growing and dividing leaf cells expressing appropriate fluorescent reporter genes, the development of software to quantify microtubule dynamics at both subcellular and tissue levels, and computer modelling. Models will be tested using cytoskeletal-specific drugs and mutants harbouring defective cytoskeletal genes. I will also apply these methods to other systems such as Utricularia.

4 - An integrated understanding of cup-shape development in Utricularia gibba

Spiral meristem of Utricularia gibba

Beatriz Gonçalves - email in collaboration with Chris Whitewoods

The transition from a two-dimensional to a three-dimensional body plan is thought to be key in the evolution of complex shapes and the colonization of different habitats. Substantial work has been done to elucidate the mechanisms behind the morphogenesis of flat organs such as leaves, while the mechanisms underlying the transition from 2D to 3D body plans and organ shapes remain poorly understood. One approach for elucidating the developmental mechanisms behind the transition from flat leaf primordia to complex cup structures is to model tissue growth as a 2D sheet that deforms out-of-plane into 3D. The tissue growth patterns behind these deformations may involve differences in growth rates, orientations or a combination of both, the genetic basis of which is an enduring challenge of developmental biology.

My project focuses on cup-shape acquisition during Utricularia gibba bladder development and aims to elucidate the patterns of tissue growth and the genes controlling its morphogenesis.

The Utricularia gibba, a carnivorous bladderwort, has been adopted as a model system for the study of complex leaf shapes in our lab and chosen especially for its small size and small diploid genome as well as its small aquatic traps that are transparent and amenable to a scope of imaging techniques including live-imaging. Currently I am using a combination of imaging, genetics, expression studies and modelling to test tissue growth hypotheses and the roles of candidate genes borrowed from the flat leaf development such as CUP and TCP related genes.

My work is funded by a Long-term Fellowship from the Federation of European Biochemical Societies (FEBS).

5 - Developmental genetics of Utricularia gibba

Man Yu - email in collaboration with Beatriz Gonçalves, Karen Lee and Chris Whitewoods

Tiny developing Utricularia bladder.

I am using the carnivorous plant Utricularia gibba as a model to understand the development of three-dimensional structures in plants. U. gibba produces cup-shaped traps to capture tiny invertebrates. The traps’ transparency, small size and quick growth make it possible to analyse their development and the rules underpinning it.

In collaboration with Karen Lee I am characterising the development of these traps using a combination of light and confocal microscopy. We will use this information to test a computational model of 3D trap development.

To understand the genes underlying these developmental transitions I am also analysing mutants in trap morphology from a forward genetic screen we have performed in the lab. In collaboration with Chris Whitewoods I will characterise mutant phenotypes and ultimately map the causative mutations.

In combination these approaches will help us to understand the genetics and development of 3D structures in plants.

6 - Dynamic Growth Maps of Leaf Development

Arabidopsis leaf one expressing GFP in the cell membranes

Samantha Fox - email

A major challenge in biology is to understand how buds comprising a few cells can give rise to complex plant and animal appendages, like leaves or limbs. We have addressed this problem through a combination of time-lapse imaging of growing leaf buds, clonal analysis and computational modelling. We have generated a model that shows how leaf shape can arise according to a few simple rules and that growth is coordinated by an in-built polarity system. Experimental tests through partial leaf ablation support this model, and allow re-evaluation of previous experimental studies. Our model allows a range of observed leaf shapes to be generated and predicts observed clone patterns in different species. Thus our experimentally validated model may underlie the development and evolution of diverse organ shapes.

Watch my Youtube movie to find out more:

7 - Exploring Inner Worlds of Plants in 3D with Optical Projection Tomography

Optical Projection Tomography Volume Rendered Plant Specimens
Inside a Cephalotus follicularis pitcher leaf trap

Karen Lee - email in collaboration with Claire Bushell, Paul Southam, Grant Calder and Jerome Avondo

Please see 3D Gallery for images of carnivorous plants and other beautiful plant specimens.

We have explored the use of Optical Projection Tomography (OPT) as a method for capturing 3D morphology and gene activity at a variety of developmental stages and scales from plant specimens, in collaboration with James Sharpe and Bioptonics . OPT can be conveniently applied to a wide variety of plant material including seedlings, leaves, flowers, roots, seeds, embryos and meristems. At the highest resolution large individual cells can be seen in the context of the surrounding plant structure. 3D domains of gene expression can be visualized using either marker genes such as β-glucuronidase, or more directly by whole-mount in situ hybridization. For naturally semi-transparent structures, such as roots or Bladderwort suction traps, live 3D imaging using OPT is possible. 3D gene expression patterns in living transgenic plants expressing fluorescent GFP markers can also be visualised. To interactively analyse and quantify OPT data, software was developed to visualise 3D volumes, accurately place points on volumes in 3D space and extract growth measurements.

Using these tools to capture leaf shape and growth, in combination with mathematical modelling, we are studying mechanisms controlling growth and shape from earliest stages of Arabidopsis leaf growth to maturity in 3D.

I am initiating a new project exploring carnivores. Carnivorous plants are amazing. They seem to turn the natural order around by being able to entice, capture and consume animal prey, when we normally think of plants as passive suppliers of nutrition for the animal world. Taking what we have learned from our Arabidopsis research we want to discover whether rules of growth underlying the development of simple leaves in Arabidopsis are are adapted to grow cup-shaped leaf traps of carnivorous plants. Using a combination of 3D imaging, genetic analysis and modelling, we aim to explore how these complex leaves develop.

Another project I am working on is a website showcasing the Inner World of Carnivorous Plants

Our carnivorous plant work is also featured in a Sightings article- 3D Carnivorous Plants, in American Science

Some of our images can also be found on the Cell Picture Show- Plant Biology

8 - Modelling Growth at the Tissue Scale

Model for Snapdragon cyc dich mutant

Richard Kennaway - email in collaboration with Erika Kuchen and Xana Rebocho

I am developing finite element methods for modelling the growth and development of curved two-dimensional tissues such as leaves and petals. The interactive software tool I am developing, called GFtbox, is available for download.

9 - Genetic Control of Growth Dynamics in a Complex Leaf

Utricularia gibba mutant trap

Robert Bellow - email in collaboration with Beatriz Gonçalves, Karen Lee, Chris Whitewoods and Man Yu

Leaves display amazing diversity and complexity in size and shape where many leaves have the main axis of growth predominantly in two-dimensions. One of the most complex leaf structures found in nature are the three-dimensional cup-shaped (epiascidiate) leaves of certain carnivorous plants. A key question is: are the genetic and mechanistic rules underlying leaf development conserved between two-dimensional and three-dimensional leaves? To elucidate how an epiascidiate leaf forms I study the carnivorous traps of Utricularia gibba.

U. gibba has one of the smallest genomes know in the plant kingdom (~100Mb), which makes it amenable to the forward genetic screen I am currently working on where some mutant lines have an altered final trap morphology. We know that certain cellular growth dynamics are required differentially for correct trap formation, how have these dynamics changes in mutated traps? Through using molecular and bioinformatic approaches, I have isolated a gene from one of these mutants, the first time that this has been accomplished in a carnivorous plant, and am further exploring what changes have occurred in the developmental process to lead to an altered shape.

BBSRC Doctoral Training Partnership on the Norwich Research Park

My work is funded by a BBSRC Doctoral Training Partnership studentship.

10 - Evolution of organ size and shape between Antirrhinum species

Antirrhinum wild-type flower

Lucy Copsey - email in collaboration with Xana Rebocho, Des Bradley and Matt Couchman.

The "old world" Antirrhinum species, found growing naturally in southern Europe and North Africa, show an extensive range of diversity in growth habit, organ size, shape and flower colour. This variation is important as it highlights differences between individuals which may be due to either environmental effects or differences at gene level, these genetic differences underpins how diversity in form is generated through evolutionary time and is the basis of evolution. By exploiting evolutionary variation in size we hope to identify genes controlling organ size in Antirrhinum species (allometry project).

I am using a classical genetics approach involving the production of a number of plant resources developed by crossing Antirrhinum species to both our cultivated JI stock 7 line and interspecies crosses to study natural variation and domestication effects. These resources are available for use by group members and for the wider Antirrhinum community. I also supervise and coordinate the genetics of Utricularia.

11 - Evolutionary Dynamics Underlying Species Diversification

Bee polinating A. pseudomajus

Matthew Couchman - email in collaboration with Xana Rebocho, Lucy Copsey, Des Bradley and Nick Barton

We are part of a collaboration investigating factors affecting gene flow across populations. The project focuses on flower colour in two distinct subspecies of Antirrhinum. Within the Spanish Pyrenees there are a number of hybrid zones that provide ideal environments for analysing how genes influence flower colour. For each individual within our chosen hybrid zone we are annually recording their GPS location and colour scores as well as taking samples for genotyping and other molecular analysis.

My role within the project is to develop a relational database to capture these and other collaborator outputs as well as a website to act as a gateway to this database. The website will include visual tools such as a configurable map of recorded GPS positions and genetic and physical maps.

12 - Natural variation of flower colour

Antirrhinum flower colour variation.

Desmond Bradley - email

Collaborators: Christophe Thebaud, Xana Rebocho, Nick Barton, David Field, Christophe Andalo and Monique Burrus

Two subspecies of Antirrhinum (magenta and yellow), which generally occur in genetic isolation in the Pyrenees, have hybridized in nature to create a hybrid zone. Due to hybridization the Antirrhinum flowers display an array of parental as well as mixed flower colour phenotypes which are the result of genetic variation at three key loci ROSEA, ELUTA and SULFUREA. The two first genes are involved in the control of the magenta anthocyanin pigmentation while SULF is a repressor of aurone (yellow) pigmentation. As the HZ gives a great playground to study evolution in action, the goal of this research is to follow the genetic flow of these flower colour loci within the hybrid population as well as the fitness of each phenotype/genotype throughout several years. I also use similar approaches to analyse gene expression regulation in Utricularia.

13 - Evolution of novel phenotypes through regulatory small RNAs

The effect on phenotype of some flower colour genes.

Daniel Richardson - in collaboration with Des Bradley, Lucy Copsey, Matt Couchman, Annabel Whibley, Tamas Dalmay, Simon Moxon, and Leighton Folkes.

Neofunctionalisation (NF) is the process by which, following a duplication event, a given gene gains a new function. This process is typically split into two classes; coding NF refers to situations where the new function arises within the coding sequence of the duplicated gene, and regulatory NF refers to instances where the new function manifests due to changes in the expression pattern of the gene.

Study of a natural hybrid zone of two subspecies of the garden snapdragon Antirrhinum majus has revealed that, within a subset of the population, positive selection is acting on a locus containing an inverted duplication of FLA, a gene involved in the synthesis of the yellow pigment aurone. This locus, SULF, appears to have arisen relatively recently in evolutionary time from multiple duplications of FLA, and acts to control patterns of yellow pigmentation on flowers through the generation of FLA-inhibiting regulatory small RNAs (sRNAs). SULF represents an interesting case study for a novel type of NF; although the duplication events have resulted in a change in the coding sequence of the FLA paralogue, the end result is a change in the expression pattern of the non-neofunctionalised FLA gene. It is presently unclear to what extent this mechanism, dubbed sRNA-mediated neofunctionalisation (SNF), is important in a wider evolutionary context.

My PhD project will utilise genomics and phylogenetics to investigate the involvement of SNF in the evolution of novel phenotypes in wild relatives of A. majus. SNF candidate loci will be identified through analysis of genome and transcriptome data from a range of morphologically diverse Antirrhinum species. The evolutionary history of validated candidates will be examined using phylogenetics. Taken together, these analyses should inform about the extent to which SNF has been involved in the generation of variation in wild snapdragons, how SNF regulatory elements coevolve with their targets, and how general the mechanism is for different traits.

BBSRC Doctoral Training Partnership on the Norwich Research Park

My work is funded by a BBSRC Doctoral Training Partnership studentship.

14 - Genetic analysis of Utricularia gibba trap development

A mature U. gibba trap.

Chris Whitewoods - in collaboration with Karen Lee, Beatriz Gonçalves and Man Yu.

Carnivorous plants have evolved cup-shaped insect traps four times independently. Each time they evolved from leaves. This raises the question of how the ancestral leaf developmental program has been modified to produce these complex structures and whether the same thing happened each time. Have genes involved in simple leaf development simply been modified or have novel genes been acquired to create the new shape?

I am using the aquatic carnivorous plant Utricularia gibba to answer these questions using both forward and reverse genetics approaches:

Firstly, in collaboration with Karen Lee I am performing a screen for trap mutants in an EMS-treated mutant population of U. gibba. We are mapping causative mutations using whole-genome-sequencing and analysing their role in trap development. This will not only help us understand the genetic mechanisms of trap development, but also enable us to compare the genetic pathways of trap development in U. gibba with that of leaf development in plants with simpler leaves to help us speculate how traps evolved.

Secondly, I am analysing the role of U. gibba homologues of genes known to be involved in simple leaf development in other plants. This will allow us to see how the ancestral leaf developmental program has been modified to create a cup shape.

The results from these approaches are being used in conjunction with computational modelling to refine our hypotheses about the development and evolution of these complex leaves.

15 - Understanding spacing of outgrowths in Arabidopsis

A young Arabidopsis leaf showing CUC2 localisation and serrations

Catherine Mansfield - in collaboration with Samantha Fox and Jordi Chan.

A huge variety of leaf shapes exist in nature. Different species of plant often have different patterns of serrations (outgrowths) on their leaves. In order to consistently generate leaves of the same shape, plants must have a mechanism of controlling outgrowth formation. A similar phenomenon is observed in phyllotaxis where plants are able to position their leaves or branches (outgrowths) at regular intervals at the whole plant level. The remarkable regularity of spacing in plants suggests a robust genetic control mechanism during development which may involve the plant hormone auxin. My work aims to understand the mechanisms of spacing through microscopy and live-imaging, molecular genetics, and modelling.

Genes known to be involved in serration formation, such as the transcription factor CUC2, will be ectopically expressed in Arabidopsis with spatial and temporal variation to understand how plants space their outgrowths at regular intervals. It will be important to establish how ectopic serrations interact with existing serrations, and how spatial and temporal variation may play a role in this. It would also be interesting to see if the mechanisms controlling spacing in leaves were similar in spacing of outgrowths in phyllotaxis which might involve ectopic expression of genes in the meristem.

Knowledge from this work will improve our understanding of the control of leaf and plant shape.

modified on 12 September 2019 at 16:41 ••• 1,038,116 views