genomewide characterization of host-pathogen interactions by ...

Maren Depke
Results
Host Cell Gene Expression Pattern in an in vitro Infection Model
and time point had the highest similarity to each other, and thus, the four treatment/time point
groups defined the three lowest level clusters (Fig. R.4.2, orange dashed line). When further
analyzing the next level of clustering the high similarity between the 2.5 h groups, i. e. infected
samples and control samples, was obvious (Fig. R.4.2, red dashed line). In the following cluster
level, the 6.5 h control samples showed higher similarity to both 2.5 h samples groups than to the
6.5 h infected samples. Summing up, the biological replicates exhibited a high reproducibility. The
highest similarity on the level of treatment groups was observed between 2.5 h infected and
control samples, and not between the control samples of both time points as it might have been
expected. The medium control samples of the 6.5 h timepoint were more similar to the 2.5 h
samples (infection and control) than to the 6.5 h infected samples.
medium control
infected; GFP positive
time point 2.5 h
time point 6.5 h
biological replicate 1
biological replicate 2
biological replicate 3
biological replicate 4
Fig. R.4.2: Hierarchical clustering of 16 array data sets from S9 infection experiment.
The following clustering algorithms were applied on log-transformed data: Agglomerative clustering with average linkage using
euclidian distance weighted by error as similarity measure. Sequences which were not expressed on all 16 arrays (defined by p > 0.01
on intensity profile level in Rosetta Resolver software) and control sequences were excluded from the cluster analysis.
The orange dashed line indicates the three lowest level clusters which led to a segmentation into the the four treatment/time point
groups. The next level of clustering grouped the 6.5 h control samples next to 2.5 h control and infected samples (red dashed line).
Comparison of treatment groups and assessment of differentially regulated genes
For identification of differential gene expression, infected samples were compared to their
time-matched controls. Additionally, same treatments were compared between the two time
points for control purposes. In detail, four comparisons of sample groups were carried out:
2.5 h infected GFP-positive samples vs. 2.5 h medium control
6.5 h infected GFP-positive samples vs. 6.5 h medium control
6.5 h infected GFP-positive samples vs. 2.5 h infected GFP-positive samples
6.5 h medium control vs. 2.5 h medium control
All comparisons are depicted by scatter plots (Fig. R.4.3). A small difference appeared
between the infected and control samples 2.5 h after infection (Fig. R.4.3, panel ), while a
strong reaction of infected cells compared to the time-matched control became visible after 6.5 h
(Fig. R.4.3, panel ). Consequently, the infected samples strongly differed between both time
points (Fig. R.4.3, panel ). But also in the control samples a certain change of signal intensities
depending on incubation time was recorded. This change mainly consisted of lower signal values
after 6.5 h than after 2.5 h (Fig. R.4.3, panel).
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