Transcript Microarray Analysis of Drosophila Development During

Microarray Analysis of
Drosophila Development During
Metamorphosis
K. P. White, S. A. Rifkin P. Hurban
and D. S. Hogness
Goal
• Use of DNA microarrays to study changes in
gene expression patterns during Drosophila
metamorphosis, induced by ecdysone peaks.
• Studies were performed during six time points
relative to the puparium formation (PF)
stages that span two ecdysone pulses.
Drosophila metamorphosis : material
The arrays
• Array elements : 6'240 cDNA clones from the
Berkeley Drosophila Genome Project, covering
about 40% of the total estimated number of
genes in Drosophila, plus known ecdysoneregulated control genes
• Interesting subset : 534 elements displaying
threefold or more differential expression during
metamorphosis
• Number of experiments : 19, covering 6 time
points relative to PF
Drosophila metamorphosis : material
The samples
• Reference sample Cy3-labelled : Animals in
puparium formation (PF) stage, 0 hour
• Time-course experiments, Cy5-labelled :18
and 4 hours before PF , and 3, 6, 9, 12 hours
after PF
Drosophila metamorphosis : methods
Two methods were used to interpret the data :
1- The Stanford clustering method.
2- A self-organizing map (SOM) algorithm.
This method allows the user to choose the
number of desired clusters.
The result is displayed in graphs which
represent the clusters. Time points are
represented in these graphs from left to right.
Drosophila metamorphosis : results
Stanford clustering
Differentially expressed genes fall
into two main categories. The first
category contains genes that are
expressed at 18 hours BFP
(before the late larval ecdysone
pulse) but then fall to low or
undetectable levels during this
pulse. This category is colored red
in the first column. They are
potentially repressed by ecdysone
The second category consists of
genes expressed at low or
undetectable levels before the late
larval ecdysone pulse but then are
induced during this pulse. They
are potentially induced by
ecdysone.
Drosophila metamorphosis : results
SOM clustering
The label in the upper left
corner of each cluster
indicates the cluster number.
The number in the top center
of each box indicates how
many elements (out= of 534)
are in each cluster. Yellow
boxes show the clusters
containing the control genes
(Known ecdysone-controlled
genes)
Blue lines are the mean
expression profiles; Red lines
indicate SD.
Drosophila metamorphosis : results
More detailed results
Genes encoding
structural and regulatory
elements of muscle
formation and other
muscle specific proteins
are down-regulated at 4
Hr BPF.
This prepares the larval
muscle breakdown
which begins
aproximately 2 hours
APF
Drosophila metamorphosis : results
More detailed results
The CNS undergoes
dramatic
differentiation during
early metamorphosis.
The genes involved
in this differenciation
are induced at the
stage 4 hours BPF,
probably by the first
ecdysone peak
Drosophila metamorphosis : results
More detailed results
Genes involved in
programmed cell-death, as
reaper, caspase-1 are
induced by the fisrt ecdysone
peak, whereas genes
encoding cell death inhibitors
are induced by the second
peak.
DHR3 is a repressor of
ecdysone-inductible genes,
but can induce genes active
during midprepupal
development (BFTZ-F1,
LD24139)
Drosophila metamorphosis : results
More detailed results
Genes encoding enzymes in the
glycolytic pathway are downregulated, suggesting that some
cells temper their metabolic activity
during metamorphosis.
The figure shows the Glycolytic
pathway. Enzymes with their gene
present on the array are listed in red.
The fold repression during the first
time-course experiment is shown by
numbers in red.