genomewide characterization of host-pathogen interactions by ...

More documents

Recommendations

Info

Maren Depke Discussion and Conclusions genes repressed (Garzoni et al. 2007) or induced (this study), saeRS repressed (Garzoni et al. 2007) or induced (this study). Clearly, the comparability is limited due to experimental differences, which underlines the importance of performing global molecular studies (transcriptome and proteome profiling) in different models to finally achieve a complete picture of the characteristics of host-pathogen interactions. Such different models include e. g. other epithelial cell lines, like the S9 cells used in this study, but can also be extended to immune cells. Such a study was performed by Voyich and colleagues in 2005. The authors analyzed the gene expression profile of different S. aureus strains upon phagocytosis by human neutrophils (Voyich et al. 2005). Again, several experimental parameters differed between the studies of Voyich and colleagues (2005), Garzoni and colleagues (2007), and this study, partly resulting from the very different host cell type employed by Voyich and coworkers. The authors described immune evasion mechanisms of S. aureus, which are initiated by transcriptional changes. First, the authors observed induction of stress response genes, e. g. superoxide dismutases sodA and sodM and several other genes involved in counteracting the influence of reactive oxygen intermediates. An as strong response was not observed in this study, which can be explained by the innate function of neutrophils to kill pathogens, which is not given for epithelial cells. Thus, staphylococci internalized in S9 cells probably encounter less harmful molecules than staphylococci after uptake by neutrophils. The induction of TCA cycle genes after phagocytosis by neutrophils (Voyich et al. 2005) corresponds with results from this study using S9 cells. Voyich and coworkers observed repression of genes involved in cell envelope synthesis, cell division, and replication, which was not as obvious in S9 cell internalized staphylococci. Nevertheless, cell wall remodeling processes were concluded from gene expression data (this study). Furthermore, staphylococci in neutrophils induced several toxins and adhesins (Voyich et al. 2005), of which some were also observed after S9 cell internalization, e. g. hlgB, hlgC, lukD, lukE; fnbB, coa, clfA (this study). The toxin gene induction appeared to be stronger in the neutrophil than in the S9 study, although hla was induced after internalization in S9 cells and not after phagocytosis by neutrophils. Anyway, the different staphylococcal strains, which were used in the studies, contribute considerably to the characteristics of the gene expression signature (Voyich et al. 2005). In the neutrophil phagocytosis dependent gene expression pattern, differential expression of regulatory genes was reported (Voyich et al. 2005). These included increased expression of vraRS, saeRS, and sarA. While induction or a trend of induction of saeRS and vraRS was also visible in this study, sarA was not differentially expressed when staphylococci were internalized in S9 cells. Furthermore, repression of agr or sigB was not observed in this study, but in the study of Voyich and coworkers. In total, after comparison of the results of Voyich and colleagues (2005), Garzoni and colleagues (2007), and this study, it becomes clear that similarities in certain aspects do not necessarily lead to completely consistent results. This probably depends to a large extent on the combination of host cell and bacterial strain, but also on further experimental conditions which varied strongly between the studies. Nevertheless, parts of the results were also found in other studies. Since an admittedly slightly imprecise attempt to specify real internalization specific gene expression that did not occur in control samples revealed virulence associated genes in the study described in this thesis, the recorded gene expression pattern is probably physiologically relevant. Therefore, the gene expression profiling turned out to be an important basic study which will entail follow-up experiments like studies of bacterial mutants, comparison of different 204
Maren Depke Discussion and Conclusions host cell lines, but also confirmation and validation experiments by application of different techniques like quantitative real-time RT-PCR. Of course, the broadening of experiments to in vivo studies would be most interesting. Here, the most prominent problems are difficulties in recovery of bacterial cells from tissue and the resulting low amount of sample or the sample contamination with host material. In this setting, application of quantitative real-time RT-PCR probably will be the method of choice, which has already proven to successfully help resolving colonization expression patterns (Burian et al. 2010a, 2010b). Until now, the tiling array data have been used as expression data set under restriction to already annotated genomic content of S. aureus. The analysis is not yet finished. Although new transcribed units have been defined for which interesting regulation patterns in the infection experiments were visible, the sequences will have to be categorized and validated. Internalization dependently regulated sequences later will be targets for further characterization experiments. It will be an interesting task to compare the resulting information on new genes with that generated with the competing approach of high-throughput transcriptome sequencing (Beaume et al 2010a, 2010b). 205
Page 1 and 2:
G E N O M E W I D E C H A R A C T E
Page 3 and 4:
Maren Depke C O N T E N T S GENOMEW
Page 5 and 6:
Maren Depke Z U S A M M E N F A S S
Page 7 and 8:
Maren Depke Zusammenfassung der Dis
Page 9 and 10:
Maren Depke S U M M A R Y O F D I S
Page 11 and 12:
Maren Depke Summary of Dissertation
Page 13 and 14:
Maren Depke I N T R O D U C T I O N
Page 15 and 16:
Maren Depke Introduction Besides op
Page 17 and 18:
Maren Depke Introduction Extracellu
Page 19 and 20:
Maren Depke Introduction with heigh
Page 21 and 22:
Maren Depke Introduction 2005). SaP
Page 23 and 24:
Maren Depke Introduction contains a
Page 25 and 26:
Maren Depke Introduction via fibron
Page 27 and 28:
Maren Depke Introduction cathelicid
Page 29 and 30:
Maren Depke Introduction shock are
Page 31 and 32:
Maren Depke Introduction STUDIES OF
Page 33 and 34:
Maren Depke Introduction are effect
Page 35 and 36:
Maren Depke Introduction cell stimu
Page 37 and 38:
Maren Depke Introduction channel CF
Page 39 and 40:
Maren Depke M A T E R I A L A N D M
Page 41:
Maren Depke Material and Methods Li
Page 44 and 45:
Maren Depke Material and Methods Ki
Page 47 and 48:
Maren Depke Material and Methods GE
Page 49:
Maren Depke Material and Methods Ge
Page 52 and 53:
Maren Depke Material and Methods Ho
Page 54 and 55:
Maren Depke Material and Methods Ho
Page 56 and 57:
Maren Depke Material and Methods Pa
Page 58 and 59:
Page 60 and 61:
Page 63 and 64:
corticosterone [pg/ml plasma] corti
Page 65 and 66:
Maren Depke Results Liver Gene Expr
Page 67 and 68:
lood glucose levels [nmol/l] resist
Page 69 and 70:
LBP [ng/ml] CPR [ng/ml] Maren Depke
Page 71 and 72:
Maren Depke Results Liver Gene Expr
Page 73 and 74:
liver weight [g] Maren Depke Result
Page 75 and 76:
infection rate cfu / 10 mg tissue c
Page 77 and 78:
Maren Depke Results Kidney Gene Exp
Page 79 and 80:
Table R.2.1: Genes displaying stati
Page 81 and 82:
Maren Depke BR1 sign DsigB vs wt BR
Page 83 and 84:
Page 85 and 86:
Page 87 and 88:
Page 89 and 90:
Page 91 and 92:
Maren Depke Results GENE EXPRESSION
Page 93 and 94:
Maren Depke Results Gene Expression
Page 95 and 96:
log2(ratio C57BL/6 / BALB/c) at IFN
Page 97 and 98:
Page 99 and 100:
Page 101 and 102:
Page 103 and 104:
Page 105 and 106:
Page 107 and 108:
Page 109:
Page 112 and 113:
Maren Depke Results Host Cell Gene
Page 114 and 115:
Page 116 and 117:
Page 118 and 119:
Page 120 and 121:
Page 122 and 123:
Page 124 and 125:
Page 126 and 127:
Page 128 and 129:
Page 130 and 131:
Page 132 and 133:
Maren Depke Results Pathogen Gene E
Page 134 and 135:
Page 136 and 137:
Page 138 and 139:
S. aureus RN1HG sample conditions a
Page 140 and 141:
Page 142 and 143:
Page 144 and 145:
Page 146 and 147:
Page 148 and 149:
mean normalized intensity mean norm
Page 150 and 151:
Page 152 and 153:
Page 154 and 155: mean normalized intensity mean norm
Page 156 and 157: Maren Depke Results Pathogen Gene E
Page 158 and 159: mean normalized intensity Maren Dep
Page 160 and 161: Maren Depke Results Pathogen Gene E
Page 168 and 169: log 2 (ratio) log 2 (ratio) log 2 (
Page 171 and 172: Maren Depke D I S C U S S I O N A N
Page 173 and 174: Maren Depke Discussion and Conclusi
Page 203: Maren Depke Discussion and Conclusi
Page 208 and 209: Maren Depke References Athanasopoul
Page 210 and 211: Maren Depke References Byrne GI, Le
Page 212 and 213: Maren Depke References Demling R. T
Page 214 and 215: Maren Depke References Foss GS, Pry
Page 216 and 217: Maren Depke References Hagopian K,
Page 218 and 219: Maren Depke References Jin T, Bokar
Page 220 and 221: Maren Depke References Kwan T, Liu
Page 222 and 223: Maren Depke References Luster AD, U
Page 224 and 225: Maren Depke References Namiki S, Na
Page 226 and 227: Maren Depke References Puccetti P.
Page 228 and 229: Maren Depke References Siewert E, B
Page 230 and 231: Maren Depke References van den Akke
Page 232 and 233: Maren Depke References Yang XL, Sch
Page 235: Maren Depke A F F I D A V I T / E R
Page 238 and 239: Maren Depke Acknowledgments Kidney
show all

genomewide characterization of host-pathogen interactions by ...

Create successful ePaper yourself

Delete template?

Save as template?