genomewide characterization of host-pathogen interactions by ...

Maren Depke
Results
Pathogen Gene Expression Profiling
Proteome analysis of internalized staphylococci, which was performed by Sandra Scharf,
revealed the increased abundance of PrsA. In parallel, the response regulator VraR of the twocomponent
system VraRS was up-regulated. Also transcriptome analysis revealed induction of
prsA in internalized staphylococci at the time point 2.5 h after start of infection. The regulatory
genes vraR and vraS were not differentially expressed. Nevertheless, with a fold change of 1.8
and 1.6 for vraR and vraS, respectively, a tendency for an induction of gene expression was
visible, which fitted well to the proteome data (Fig. R.5.21 A, B).
The VraRS regulon contains 46 genes including vraR and vraS (Kuroda et al. 2003). Of the
published 46 S. aureus N315 LocusTags, 45 could be mapped to S. aureus NCTC8325 LocusTags
(NCBI’s EntrezGenome Gene Plot; http://www.ncbi.nlm.nih.gov/sutils/geneplot.cgi). When asking
for the fraction of the VraRS regulon which was regulated in internalized S. aureus RN1HG in S9
cells, 22 % (2.5 h after start of infection) and 11 % (6.5 h after start of infection) were differentially
expressed upon internalization. In detail, 7 genes were induced 2.5 h after start of infection, of
which 4 were still induced at the later time point of 6.5 h after start of infection. Repressed gene
expression was also observed: Repression was detected for 3 genes at the early 2.5 h time point,
and one of these genes still was repressed at the later 6.5 h time point (Fig. R.5.22 A, B).
A
VraRS regulon according to
Kuroda et al. 2003
B
VraRS regulon according to
Kuroda et al. 2003
35
40
7 3
0
395 360
0
4 1
0
315 307
0
induced sequences in
“2.5 h internalization” vs.
“1 h serum/CO 2 control”
repressed sequences in
“2.5 h internalization” vs.
“1 h serum/CO 2 control”
induced sequences in
“6.5 h internalization” vs.
“1 h serum/CO 2 control”
repressed sequences in
“6.5 h internalization” vs.
“1 h serum/CO 2 control”
Fig. R.5.22: Differential expression of genes belonging to the VraRS regulon in internalized S. aureus RN1HG in S9 cells.
Genes known to be regulated by VraRS (according to Kuroda et al. 2003) were compared to S. aureus RN1HG S9 internalization gene
expression signatures separately for induced and repressed genes.
A. 2.5 h after start of infection.
B. 6.5 h after start of infection.
Differentially regulated metabolic genes
First, BIOCYC “omics-Viewer” pathway mapping (BIOCYC, SRI International, CA, USA,
http://biocyc.org/expression.html) was applied for a comprehensive overview on changes in
gene expression of metabolic enzymes in internalized and control staphylococci. This tool allows
the display of gene expression data on highly abstracted metabolic pathway schemes and
therefore an intuitive comprehension of processes in the selected experimental setup.
In comparison to the baseline of 2.5 h serum/CO 2 control samples, differentially expressed
sequences in a) 2.5 h internalization, b) 6.5 h internalization, c) 2.5 h serum/CO 2 control, and
d) 2.5 h anaerobic incubation were included in the analysis.
Several changes in the expression of metabolic genes became visible, but certain functionally
associated groups of genes accumulated changes in expression (Fig. R.5.23).
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