Exercises: Gene Ontology - Theoretical Biology & Bioinformatics

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Oct 1, 2013 (4 years and 1 month ago)

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Exercises:
Gene Ontology


Assignment 1:

Functional overrepresentation of PLETHORA downstream targets


Growth in plants is located in the meristems. In the root meristem, there
are stem cells surrounding a small group of quiescent cells, the quiescent
center (QC). In the group of Ben Scheres research focuses on gene
regulatory networks responsible for the identity and maintenance of the
stem cell niche in the root meristem. The PLETHORA (PLT) transcription
factor family has turned out to be key componen
ts in these processes. PLT
expression is in the QC and depends on auxin accumulation. Double mutants
(
plt1 plt2
) mis
-
specify the QC and cannot maintain stem cells. Moreover,
when ectopically expressed, these genes specify new QC and root stem cells
in any
position. However, the molecular mechanism to achieve this is
unclear. Hence there have been done micro
-
array experiments overexpressing
(35S::PLT
1 and 35S::PLT2
) to elucidate targets of plethora transcription
factors.


Left panel: PLT genes expressed in
stem cells
region of root tip.

Right panel:
plt1plt2
double mutants do not
maintain the root meristem.





Left panel: overexpression of PLT2 induces roots.

Right panel: ectopic induction of PLT2 in the
shoot induce root meristem formation.





Download from
http://bioinformatics.bio.uu.nl/snel/support/

the datasets
named PLT
1
_
targets
.txt and PLT
2
_
targets
.txt. In the web browser, go to
VirtualPlant (
http://virtualplant.bio.nyu.edu/cgi
-
bin/vpweb/
) and upload
each list of probes in separate datasets.


-

Rename each dataset and compare with the initial list. Is there any
difference? Why?

-

How many targets are shared by PLT
1
and P
LT
2
? Which ones are unique
for each?

-

If we suspect that PLT
1
and PLT
2

are redundant, which dataset will
be the more meaningful to use for functional analysis?

-

How would you summarize the functions of the transcriptional program
carried out by PLT
1

and PLT
2
?


Assignment 2:
Shade avoidance


pathogen response: which one will prevail?



Since plants cannot run away from predators, they have to be creative when
it comes to fighting them away. Similarly, if plants find themselves being
overshadowed by leaves
from neighboring plants, they have to adjust growth
strategies. At the same time, other interactions with the environment such
as attacks by pathogens also force the plant to allocate its limited
energy and nutrient budget.

In the ecophysiology group, Ronald Pierik and Mieke de Wit have uncovered
hints that there is “cross
-
talk” between the “shade avoidance pathway” and
the pathogen response that is mediated by the plant hormone jasmonic acid.
Therefore they performed microar
ray analysis of control growth, growth
under far infrared light (the signal sensed for shade avoidance), growth
under the addition of jasmonic acid, and a combimation of the latter two
conditions. This should help to elucidate the biology of the cross
-
talk

of
the processes.


Download / copy from
http://bioinformatics.bio.uu.nl/snel/support/

the 3
datasets named 1) Me_JA_DEGs.txt (only jasmonate treatment); 2)
R_Frlight_DEGs.txt (only light treat
ment); 3)
light
_
JA
_combined_DEGs.txt
(both treatments) corresponding to the differentially expressed genes in
each of the experiments.


-

How much overlap there is between the different treatments? Use
Venny website (
http://bioinfogp.cnb.csic.es/tools/venny/
) to obtain
a graphical representation.


Install cytoscape (see
http://www.cytoscape.org/
). This can take a while.
Open cytoscape and go to plugins
-
> manag
e plugins and install BINGO v2.44
(this can also take a while). Now you can use BINGO with the different
lists you obtained in Venny to answer the next questions:


-

For JA treatment, in the table output for the process “response to
jasmonic acid stimulus” h
ow many genes in our gene cluster are in
the process “response to jasmonic acid stimulus”?

-

How many genes are in our gene cluster?

-

How many genes in the genome/background are in “response to jasmonic
acid stimulus?

-

How many genes are in the background?

-

Compare these numbers to a few other highly significantly enriched
GO terms and to a few other not so significantly enriched GO terms.

-

Compare the table output to the network / tangled hierarchy output
of BINGO. What do you experience as strengths and wea
knesses of each
mode of display?

-

Also do the GO analysis for molecular function and for subcellular
compartment.

-

Inspect the results and speculate on which pathway(s) is
repressed/dominant when both stimuli are applied compared to the
conditions when only

one stimulus applied?


Assignment 3:
What about your own data?


Following a similar procedure for assignment 2, perform a functional
analysis in BINGO (i.e. not only GO process) for your set of
differentially expressed genes.