Download Invstigating Modelling with ePlant

Investigating Biological Modelling With ePlant
Teaching Notes
ePlant is an online tool that allows scientists to visualise genetic data related to Arabidopsis
thaliana, the model plant, in 3D. It allows you to select a gene and follow its influence over the
plant from the genome all the way up to the whole organism.
This resource contains two activities: Activity One demonstrates the link between expression
of different genes in a plant organ and its function, Activity Two shows how protein function is
influenced by its structure.
Selecting a gene of interest
To use ePlant, you must first choose a gene of interest, referred to by its AGI ID (the
identification number for the gene given by the Arabidopsis Genome Initiative). We have preselected a number of genes for you to use in this resource. The following genes are
particularly relevant to the A-level biology syllabus. The AGI ID is shown in italics:



At2g39730 A protein used to synthesise rubisco, the enzyme which catalyses the
Calvin cycle in the light independent reactions of photosynthesis.
At1g19300 A protein involved in xylem wall synthesis.
At3g47730 A transcription factor which controls plant development and flowers.
Using ePlant – a brief overview
1. Open your web browser and enter http://bar.utoronto.ca/eplant/ into the address bar.
Figure 3: The home page of ePlant.
2. Enter an AGI ID into the text box at the top left of the screen.
Science & Plants for Schools: www.saps.org.uk
Investigating Biological Modelling with ePlant – teachers’ notes
This document may be reproduced for educational use in any institution taking part in the SAPS programme. It may
not be reproduced for any other purpose. Revised 2011.
-1-
3. To choose a function, press one of the buttons from the bar across the top of the
window: Homologs and Polymorphisms, Plant Expression, Tissue Expression,
Subcellular Location, Interactors or Protein Model. In these activities, we will use only
two functions, Plant Expression and Protein Model.
4. To switch between different functions, select one of the other function buttons on the
bar across the top of the window. Your gene of interest will remain the same.
To discover where the raw data originated, select the function you are interested in. In the
bottom right corner of the window, there is a link to a paper which supplied the raw data.
Activity 1: Demonstrating the Relationship between Expression of Different
Genes and the Function of the Tissue
ePlant provides a clear visual demonstration of the way that the protein complement of a cell
is dependent on its location and its function; cells in different tissues express only the genes
which are necessary for the specific function of that tissue.
In this activity, we suggest that you introduce each gene and its role, and then ask the
students where in the plant they expect it to be expressed.
Question 1 – Where will an enzyme involved in chlorophyll biosynthesis be expressed?
At5g18660 is a gene for a Divinyl Reductase (DVR) enzyme, which is involved in chlorophyll
biosynthesis.
As students will understand the role of chlorophyll in the light dependent stage of
photosynthesis, they should be able to predict that genes for chlorophyll biosynthesis would
be highly expressed in the leaves.
1) Enter the AGI ID At5g18660 into the box at the top left side of the window.
2) Select ‘Plant Expression’ from the top menu. The programme will now draw a 3D
image of the Arabidopsis plant and highlight the areas where the gene is expressed.
It may take up to 30 seconds to fully load and highlight the image. (A progress
indicator is shown in the bottom right-hand corner of the screen.)
3) Areas of the plant where expression levels of the gene are high are shown in red, and
areas where expression levels are low are shown in yellow. Note that only key areas
of the plant are highlighted, and the rest of the image remains in grey.
4) Tick the ‘rotate’ box to rotate the plant a few times, and then untick the box to hold the
plant still.
The image shows that the gene is most highly expressed in the mature leaves, leaf 8 and leaf
10. There is no expression in the other organs, as you may expect. The expression is slightly
lower in less mature leaves, indicating lower levels of chlorophyll biosynthesis enzymes here.
At present there is no known reason why this might be.
Question 2 – Where will an enzyme for xylem biosynthesis be expressed?
At1g19300 is a gene for an enzyme for xylem biosynthesis
Students will understand the role of xylem in the plant, and so should be able to predict where
the At1g19300 gene will be expressed (i.e. high levels in the stem and the root, and very low
levels in the flower and the leaves).
Science & Plants for Schools: www.saps.org.uk
Investigating Biological Modelling with ePlant – teachers’ notes
This document may be reproduced for educational use in any institution taking part in the SAPS programme. It may
not be reproduced for any other purpose. Revised 2011.
-2-
In order to view the location of expression of this gene in Arabidopsis, enter the AGI ID
At1g19300 into the box at the top left side of the window. Then follow steps 2-4 outlined
above.
The 3D image shows a high level of expression in the stem and the root (shown in red),
where xylem tissue is located. By contrast, the flower and the leaves show far lower levels of
expression (shown in yellow). This demonstrates that the protein complement of a cell is
dependent on its location and its function. The first node in the plant is orange, showing a
slightly lower level of xylem than in the rest of the stem. This is because there is little vascular
tissue around the leaf node because vascular tissue here is diverted towards the leaf. This is
known as the ‘leaf gap’.
Question 3 – Where will a gene involved in stomatal opening be expressed?
At1g08810 is a gene involved in stomatal opening.
In order to view the location of expression of this gene in Arabidopsis, enter the AGI ID
At1g08810 into the box at the top left side of the window. Then follow steps 2-4 outlined
above.
Students will recognise the role of the stomata, and are likely to predict that the highest level
of expression would be in the leaves.
However, this gene encodes a transcription factor (a protein which controls the expression of
other genes). This transcription factor also regulates growth responses as well as stomatal
opening. As a result, the gene displays even higher levels of expression in the seed siliques,
the organs which will create the seed pod. The leaves appear light orange, while the seed
siliques are red, indicating a higher level of expression here.
This shows that many proteins can perform more than one function, which means that a plant
only needs one gene to perform multiple functions. This means that the plant can act more
efficiently and use fewer resources.
Question 4 – Where will a gene relating to the activation of Rubisco be expressed?
At2g39730 (Rubisco activase) is an enzyme which activates Rubisco, the enzyme which
catalyses the Calvin Cycle in photosynthesis.
Students are likely to predict that the gene will appear in very similar places to those of
At5g18660 in question 1.
In order to view the location of expression of this gene in Arabidopsis, enter the AGI ID
At2g39730 into the box at the top left side of the window. Then follow steps 2-4 outlined
above.
High levels of expression are seen throughout the plant (with the exception of the roots).
Highest levels of expression are seen in the leaves (which are red), the main site of
photosynthesis.
Students may compare these high levels of Rubisco to the comparatively low levels of
At5g18660, the chlorophyll biosynthesis enzyme shown above, which seems to have a far
lower level. They may speculate why this is - it is because there are many photosynthetic
compounds, such as the carotenes and xanthophylls, as well as chlorophylls a and b.
Science & Plants for Schools: www.saps.org.uk
Investigating Biological Modelling with ePlant – teachers’ notes
This document may be reproduced for educational use in any institution taking part in the SAPS programme. It may
not be reproduced for any other purpose. Revised 2011.
-3-
Activity 2: Demonstrating the Relationship between Protein Structure and
Function
ePlant links through to JMol, a tool for visualising molecules in 3D. You will need to have Java
installed on your computer in order to use JMol. This tool provides a useful way of discussing
the relationship between the structure of a protein and its function by showing a 3 dimensional
model of the protein structure.
You may wish to show the students the protein structure first, and invite them to suggest what
its function might be. You can also switch to the ‘Plant Expression’ view (as in Activity 1), to
show that the gene is found throughout the plant.
Question 1 – How does the structure of an aquaporin relate to its function?
Selected gene: At2g25810 (an aquaporin).
Aquaporins are a pore proteins which transport water molecules across cell membranes by
facilitated diffusion.
1. Enter the AGI ID At2g25810 into the box at the top left side of the window.
2. Select the ‘Protein Model’ from the top menu.
3. There is a list of links divided into two sections. Select the first link from the top
section.
4. The protein should appear as eight alpha helices arranged in a ring. (This may take
up to 30 seconds to load when you first run it.)
5. To manipulate the image and get a different view of the structure, click on the protein
and drag it around the screen. The protein should move as the mouse moves.
6. To view the protein from the side: right click on the protein, select ‘View’ from the
menu which appears and then select ‘Front’.
7. To view the protein through the pore: right click on the protein, select ‘View’ from the
menu which appears and then select ‘Top’.
When observed from the side, it should appear as several alpha helices arranged in a ring. By
looking through the protein from the top, you should be able to see the pore in the
arrangement, which the water molecules will pass through.
If shown the structure of the protein from various angles, students may be able to guess the
function of this protein as a transmembrane transport protein. This may also lead to a
discussion about different types of transmembrane protein and different types of transport and
osmosis.
When viewed in the ‘Plant Expression’ view in ePlant (as in Activity 1), aquaporins appear to
be expressed all over the plant. This is because all cells regulate their water supply in order to
avoid having too much or too little water in them. Students may have learnt about plasmolysis
in cells, where the cell contains too little water and the cell membrane pulls away from the cell
wall.
Question 2 – How does this compare to another transmembrane transporter?
Selected gene: At4g13420 (A High Affinity Potassium Ion Transporter)
Science & Plants for Schools: www.saps.org.uk
Investigating Biological Modelling with ePlant – teachers’ notes
This document may be reproduced for educational use in any institution taking part in the SAPS programme. It may
not be reproduced for any other purpose. Revised 2011.
-4-
This protein is another transmembrane transporter, which is mainly present in the roots and
transports potassium ions into root hair cells from the soil surrounding the root. It is induced
as a response to potassium starvation in the cell- visible symptoms of potassium starvation
include curling and the appearance of purple spots on the leaves. The tissue between leaf
veins may begin to turn yellow, showing symptoms of chlorosis (where the leaves contain too
little chlorophyll).
Potassium and sodium have a similar charge and are almost the same diameter (1.33Å for
potassium, 0.95Å for sodium). As the cells require different concentrations of potassium and
sodium, the channel must allow the transport of potassium but not sodium, so potassium
channels have a selectivity filter. This is one reason why this channel appears more complex
than the aquaporin from the above example, as aquaporins do not have the same difficulties
with selectivity.
When looking at the protein, to look through the pore first right click on the protein, select
‘View’ and then select ‘Front’. To view the protein from the side right click on the protein,
select ‘View’ and then select ‘Top’. This will align the protein as it would appear when viewed
in the plane of the membrane.
When viewed in ‘Plant Expression’ (as in Activity One), the highest levels of expression are in
the roots. This is because potassium ion uptake occurs in the roots via the root hair cells.
More information about potassium channels:
http://proteopedia.org/wiki/index.php/Potassium_Channel
Using TAIR to Identify an Arabidopsis gene
If you wish to use a gene other than those recommended in these activities, you will need to
identify its AGI ID using a tool such as The Arabidopsis Information Resource (TAIR). TAIR
allows you to search for genes either by function or by name.
i) Enter http://www.arabidopsis.org/servlets/Search?action=new_search&type=gene
in a new tab.
Figure 1: The gene search page for The Arabidopsis Initiative Resource (TAIR).
Science & Plants for Schools: www.saps.org.uk
Investigating Biological Modelling with ePlant – teachers’ notes
This document may be reproduced for educational use in any institution taking part in the SAPS programme. It may
not be reproduced for any other purpose. Revised 2011.
-5-
ii) Search for a gene by name or function (using the key word box) using the
selectable options and click ‘Submit Query’.
iii) Enter the AGI ID (Fig.2, highlighted) into the AGI ID box on the top right hand
corner of ePlant.
Figure 2: The results of a gene search. The AGI ID is highlighted in yellow.
Science & Plants for Schools: www.saps.org.uk
Investigating Biological Modelling with ePlant – teachers’ notes
This document may be reproduced for educational use in any institution taking part in the SAPS programme. It may
not be reproduced for any other purpose. Revised 2011.
-6-