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.

At the subcellular scale, we are analysing the role of microtubules and nuclear movements in the control of cell division and growth. This involves extensive confocal imaging, image processing and computational modelling1,3,. 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 imagingand Optical Projection Tomography2.

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 studies5,6,7,8,. 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 approaches.

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 traps4,2,9 . We are using a combination of genetic analysis, imaging and mathematical modelling to understand how these leaves shape themselves.


Funding



1 - Microtubule dynamics during growth and division

Pre-prophase band formation in a leaf epidermis cell.

Jordi Chan - email in collaboration with Richard Kennaway

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.


2 - 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 Rob Bellow and Chris Whitewoods.

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 ╬▓-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


3 - Modelling Growth and Development

Model for Snapdragon cyc dich mutant

Richard Kennaway - email

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


4 - Genetic Control of Growth Dynamics in a Complex Leaf

Utricularia gibba mutant trap


Robert Bellow - email in collaboration with Karen Lee and Chris Whitewoods

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.


5 - Evolution of organ size and shape between Antirrhinum species

Antirrhinum wild-type flower

Lucy Copsey - email in collaboration with Des Bradley, Daniel Richardson 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.


6 - Evolutionary Dynamics Underlying Species Diversification

Bee polinating A. pseudomajus

Matthew Couchman - email in collaboration with Lucy Copsey, Des Bradley and Daniel Richardson.

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.


7 - Natural variation of flower colour

Antirrhinum flower colour variation.

Desmond Bradley - email

Collaborators: Annabel Whibley, 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.

8 - 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.


9 - Genetic analysis of Utricularia gibba trap development

A mature U. gibba trap.

Chris Whitewoods - in collaboration with Karen Lee, Rob Bellow and Beatriz Gon├žalves

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.

modified on 14 November 2020 at 17:37 ••• 1,040,977 views