genomewide characterization of host-pathogen interactions by ...

mean normalized intensity
mean normalized intensity
Maren Depke
Results
Pathogen Gene Expression Profiling
Additionally, repression was observed for hysA, htrA, and nuc in 2.5 h serum/CO 2 control.
While hysA and htrA were repressed in both analyzed time points of internalization, the
repression of sspA was only detected 2.5 h after start of infection. The other two genes of the ssp
operon were not detected as differentially expressed (Fig. R.5.16 A, C).
Staphylococci secrete proteins which help the bacterium to evade the host’s immune
response. In the experimental setting of the in vitro S9 infection and internalization model, some
of them were induced in S. aureus RN1HG after internalization in S9 cells, especially at the 6.5 h
time point: chp (chemotaxis inhibitory protein CHIPS), eap (extracellular adherence protein, also
called MHC class II analog protein Map), and efb (extracellular fibrinogen-binding protein).
Staphylokinase (sak) was repressed in internalized staphylococci at the 2.5 h time point
(Fig. R.5.17 A, B). Staphylococci own two genes coding for superoxide dismutases, sodA and
sodM. These two gene exhibited divergent regulation during internalization: sodA was induced
and sodM was repressed in at least one of the two analyzed time points (Fig. R.5.17 A, C).
A
fold change in comparison to
baseline 1 h serum/CO 2 control
S. aureus
RN1HG
sample
conditions
and time
points
chp eap efb sak sodA sodM
exponential growth phase -7.9 -2.0 -2.8 -2.8 2.4 -1.2
2.5 h internalized 1.7 1.8 1.9 -3.8 2.4 -1.7
6.5 h internalized 4.4 11.0 2.7 -1.9 3.2 -3.0
2.5 h serum/CO 2 control -1.2 2.0 -1.3 2.2 1.9 -1.1
6.5 h serum/CO 2 control -3.1 4.7 -1.3 -2.7 2.7 -2.5
2.5 h anaerobic incubation -4.6 3.1 1.2 1.1 1.3 -1.6
B
100.0
C
10.0
10.0
1.0
chp
eap
efb
sak
1.0
sodA
sodM
0.1
exp.
1 h
co.
2.5 h
int.
6.5 h
int.
2.5 h
co.
6.5 h
co.
2.5 h
anae.
S. aureus RN1HG sample conditions and time points
0.1
exp.
1 h
co.
2.5 h
int.
6.5 h
int.
2.5 h
co.
6.5 h
co.
2.5 h
anae.
S. aureus RN1HG sample conditions and time points
Fig. R.5.17: Immune evasion gene expression.
A. Overview on fold change values relative to the baseline sample “1 h serum/CO 2 control” and on significance in statistical group
comparisons. Differentially expressed genes are marked by gray filling for the corresponding sample condition/time point.
Genes which were significant in statistical testing (p* < 0.05) but did not pass the absolute fold change cutoff 2 are indicated in bold
without filling. The sample “6.5 h serum/CO 2 control” could not be included in statistical testing because of small group size (n = 2).
B, C. Overview on mean normalized gene expression intensity for different immune evasion (B) and superoxide dismutase (C) genes.
After the global normalization by inter-chip scaling and detrending, each individual gene has been normalized to the expression level
of the baseline sample “1 h serum/CO 2 control”. Although not being continuous data, values were depicted as line graphs instead of
bar charts for better facility of inspection.
(exp. − exponential growth phase; 1 h co. – 1 h serum/CO 2 control; 2.5 h int. − 2.5 h internalization; 6.5 h int. − 6.5 h internalization;
2.5 h co. − 2.5 h serum/CO 2 control; 6.5 h co. − 6.5 h serum/CO 2 control; 2.5 h anae. − 2.5 h anaerobic incubation)
Changes in the cell wall allow staphylococci defense against and evasion of the immune
response. The dlt operon for example is responsible for the incorporation of D-alanine into the
teichoic acids, which leads to a reduction of negative charge of the cell wall. Thus, these bacterial
cells are less vulnerable by antimicrobial cationic peptides due to a reduced attraction.
Surprinsingly, this operon was temporarily repressed at the 2.5 h time point of internalization. At
the same time and also 6.5 h after start of infection, the genes of cell wall modulating enzymes
lytM (peptidoglycan hydrolase, endopeptidase) and ssaA (secretory antigen precursor, amidase)
were induced in internalized staphylococci (Fig. R.5.18 A, B).
152