genomewide characterization of host-pathogen interactions by ...
genomewide characterization of host-pathogen interactions by ...
genomewide characterization of host-pathogen interactions by ...
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log 2 (ratio)<br />
log 2 (ratio)<br />
log 2 (ratio)<br />
log 2 (ratio)<br />
log 2 (ratio)<br />
log 2 (ratio)<br />
log 2 (ratio)<br />
log 2 (ratio)<br />
log 2 (ratio)<br />
log 2 (ratio)<br />
log 2 (ratio)<br />
log 2 (ratio)<br />
Maren Depke<br />
Results<br />
Pathogen Gene Expression Pr<strong>of</strong>iling<br />
For the new transcripts included in the 2.5 h internalization signature, 6 cluster groups were<br />
defined (Fig. R.5.37 A). Cluster group 1 contained 26 transcripts. Expression was increased in the<br />
2.5 h samples and still slightly stronger in the 6.5 h samples. The serum/CO 2 controls at both time<br />
points also exhibited increase <strong>of</strong> expression. Cluster group 2 harbored 32 transcripts which<br />
featured increase in expression at the 2.5 h and to a lesser extent at the 6.5 h internalization time<br />
points and in the later 6.5 h serum/CO 2 control whereas the 2.5 h serum/CO 2 control exhibited in<br />
average no change. Transcripts with decreased expression in the 2.5 h internalized and less<br />
strongly in the 6.5 h sample belonged to cluster group 3 which included 42 transcripts. The 2.5 h<br />
serum/CO 2 control exhibited no apparent trend <strong>of</strong> regulation whereas the 6.5 h control exhibited<br />
a decrease in expression. In cluster group 4, 26 transcripts were classified, which exhibited a<br />
decrease <strong>of</strong> expression in both internalization samples, a slight increase in the 2.5 h serum/CO 2<br />
controls and a decline <strong>of</strong> expression to the reference level in the 6.5 h serum/CO 2 controls.<br />
Internalization samples in cluster group 5 showed an increase <strong>of</strong> expression 2.5 h after start <strong>of</strong><br />
infection, which again declined in samples after 6.5 h. Contrarily, expression in the serum/CO 2<br />
controls was induced at the 2.5 h time point and further increased at the 6.5 h time point. This<br />
cluster contained 49 sequences. Finally, cluster group 6 contained 25 sequences which were<br />
increased in both internalized samples, and possessed a variable expression pattern in the 2.5 h<br />
serum/CO 2 control with increased, repressed and unchanged values, while the 6.5 h serum/CO 2<br />
control samples exhibited a trend <strong>of</strong> decreased expression.<br />
A<br />
B<br />
cluster group 1<br />
cluster group 2<br />
cluster group 1<br />
cluster group 2<br />
6<br />
4<br />
2<br />
0<br />
-2<br />
-4<br />
-6<br />
1 h<br />
n. ad.<br />
2.5 h<br />
int.<br />
6.5 h<br />
int.<br />
2.5 h<br />
co.<br />
6.5 h<br />
co.<br />
6<br />
4<br />
2<br />
0<br />
-2<br />
-4<br />
-6<br />
1 h<br />
n. ad.<br />
2.5 h<br />
int.<br />
6.5 h<br />
int.<br />
2.5 h<br />
co.<br />
6.5 h<br />
co.<br />
6<br />
4<br />
2<br />
0<br />
-2<br />
-4<br />
-6<br />
1 h<br />
n. ad.<br />
2.5 h<br />
int.<br />
6.5 h<br />
int.<br />
2.5 h<br />
co.<br />
6.5 h<br />
co.<br />
6<br />
4<br />
2<br />
0<br />
-2<br />
-4<br />
-6<br />
1 h<br />
n. ad.<br />
2.5 h<br />
int.<br />
6.5 h<br />
int.<br />
2.5 h<br />
co.<br />
6.5 h<br />
co.<br />
cluster group 3<br />
cluster group 4<br />
cluster group 3<br />
cluster group 4<br />
6<br />
4<br />
2<br />
0<br />
-2<br />
-4<br />
-6<br />
1 h<br />
n. ad.<br />
2.5 h<br />
int.<br />
6.5 h<br />
int.<br />
2.5 h<br />
co.<br />
6.5 h<br />
co.<br />
6<br />
4<br />
2<br />
0<br />
-2<br />
-4<br />
-6<br />
1 h<br />
n. ad.<br />
2.5 h<br />
int.<br />
6.5 h<br />
int.<br />
2.5 h<br />
co.<br />
6.5 h<br />
co.<br />
6<br />
4<br />
2<br />
0<br />
-2<br />
-4<br />
-6<br />
1 h<br />
n. ad.<br />
2.5 h<br />
int.<br />
6.5 h<br />
int.<br />
2.5 h<br />
co.<br />
6.5 h<br />
co.<br />
6<br />
4<br />
2<br />
0<br />
-2<br />
-4<br />
-6<br />
1 h<br />
n. ad.<br />
2.5 h<br />
int.<br />
6.5 h<br />
int.<br />
2.5 h<br />
co.<br />
6.5 h<br />
co.<br />
cluster group 5<br />
cluster group 6<br />
cluster group 5<br />
cluster group 6<br />
6<br />
4<br />
2<br />
0<br />
-2<br />
-4<br />
-6<br />
1 h<br />
n. ad.<br />
2.5 h<br />
int.<br />
6.5 h<br />
int.<br />
2.5 h<br />
co.<br />
6.5 h<br />
co.<br />
6<br />
4<br />
2<br />
0<br />
-2<br />
-4<br />
-6<br />
1 h<br />
n. ad.<br />
2.5 h<br />
int.<br />
6.5 h<br />
int.<br />
2.5 h<br />
co.<br />
6.5 h<br />
co.<br />
6<br />
4<br />
2<br />
0<br />
-2<br />
-4<br />
-6<br />
1 h<br />
n. ad.<br />
2.5 h<br />
int.<br />
6.5 h<br />
int.<br />
2.5 h<br />
co.<br />
6.5 h<br />
co.<br />
6<br />
4<br />
2<br />
0<br />
-2<br />
-4<br />
-6<br />
1 h<br />
n. ad.<br />
2.5 h<br />
int.<br />
6.5 h<br />
int.<br />
2.5 h<br />
co.<br />
6.5 h<br />
co.<br />
Fig. R.5.37: K-means clustering <strong>of</strong> gene expression data from newly identified transcripts, which exhibited differential expression in<br />
the 2.5 h internalized sample (A) or in the 6.5 h internalized sample (B) in comparison to the 1 h serum/CO 2 control with statistical<br />
testing and multiple testing correction (p* < 0.05) and a minimal absolute fold change cut<strong>of</strong>f <strong>of</strong> 2. For k-means clustering, the<br />
following parameters were applied: Transcripts should be assigned to the user-defined number <strong>of</strong> 6 groups and sorted <strong>by</strong> deviation<br />
using the metric type cosine correlation. No additional data preparation or statistical cuts were applied.<br />
The ratio data set (experimental condition normalized to the 1 h serum/CO 2 control) employed for clustering consisted <strong>of</strong> the five<br />
experimental conditions <strong>of</strong> non-adherent (1 h), internalized (2.5 h and 6.5 h), and serum/CO 2 control (2.5 h and 6.5 h) staphylococci.<br />
Although not being continuous data, values were depicted as line graphs instead <strong>of</strong> bar charts for better facility <strong>of</strong> inspection.<br />
(1 h n. ad. – non-adherent staphylococci after 1 h <strong>of</strong> infection; 2.5 h int. − 2.5 h internalization; 6.5 h int. − 6.5 h internalization;<br />
2.5 h co. − 2.5 h serum/CO 2 control; 6.5 h co. − 6.5 h serum/CO 2 control)<br />
Correspondingly, also for the new transcripts included in the 6.5 h internalization signature,<br />
6 cluster groups were defined (Fig. R.5.37 B). Here, cluster group 1 contained 19 transcripts with<br />
168