Coordinated regulation of gene expression by E ... - Jacobs University

More documents

Recommendations

Info

DISCUSSION topological deformation of DNA, underscoring the importance of understanding the relationships between DNA topology and DNA architectural proteins. 4.1 Interdependency of supercoiling and NAPs Supercoiling in E.coli is predominantly controlled by gene products of gyrA, gyrB and topA. While gyrA & gyrB promoters are repressed by FIS [Schneider et al., 1999], in vitro studies on binding of FIS at topA promoter has been shown to activate its expression depending on the concentration of FIS [Weinstein-Fischer & Altuvia, 2007]. Since supercoiling in vivo is maintained by the opposing actions of these enzymes [DiNardo et al., and Pruss et al., 1982], their relative activities could be regulated to produce various levels of supercoiling which also has been shown to be modulated by FIS [Schneider et al., 1997]. Thus FIS affects both the activity and the expression of topoisomerases and helps in maintaining a constant superhelical energy during exponential growth in E.coli. Part of this homeostatic control is also due to the regulation of topoisomerase expression in response to supercoiling [Menzel et al., 1983; Tse-Dinh et al., 1985]. H-NS mutant has been reported to show an increase in unconstrained supercoiling [Mojica & Higgins, 1997]. Homeostatic control of supercoiling is exerted by topoisomerases responding to a change in the number of unrestrained supercoils, which is a consequence of change in binding of NAPs or change in external environment. The only known effector for DNA gyrase to sense the nutritional richness of the environment is the in vivo ATP level. The ATP level has been implicated in the recovery of supercoils after starvation, but the relaxation of DNA was seen to be not directly correlated with the cellular ATP levels [Balke and Gralla, 1987]. Transcription and RNAP binding also change the dynamics of supercoiling [Gamper & Hearst, 1982]. In spite of different perturbations, the bacteria during steady-state growth strive to maintain a constant level of superhelical energy. Hence a fundamental question that arises is how the cell determines the level of supercoiling that is appropriate to each physiological situation? 59
DISCUSSION In a study by Balke & Gralla, the changes in supercoiling has been shown to be dependent on the growth phase rather than on the growth rate [Balke & Gralla, 1987]. In the current work, though there are differences in the growth rates of wild-type, fis (lower growth rate compared to wild-type) & hns (higher growth rate compared to wildtype) mutants the supercoiling does not differ much (Figure 12). Since it is known that in slower-growing cells, transcription from stable RNA promoters is decreased relative to that in fast-growing cells, the decrease in growth rate in the fis mutant or the increase in the growth rate of hns mutant can be a direct effect of these proteins on the stable RNA transcription. The molecular basis of this type of growth control and the role of FIS will be discussed later. H-NS, the other NAP studied here forms a filament like structure bridging two DNA duplexes and acting as a zipper [Dame, 2005]. The increase in negative supercoiling shown here is consistent with the previous reports [Mojica et al., 1997]. The reason for this increase is not clearly understood. The role derepression of transcription in hns mutant leading to an increased global supercoiling (according to twin-domain model of supercoiling distribution) has been ruled out [Mojica et al., 1997]. Our recent study identified a 10bp AT-rich consensus recognized by H-NS and serving as a nucleation site to restrain negative supercoiling [Lang et al., 2007]. FIS protein (1 FIS molecule per 450bp of E.coli genome) is more abundant than H-NS (1 H-NS dimer per 1400bp) during exponential growth. These two proteins are known to compete for binding to DNA and their binding also depends on the supercoiling. FIS is known to preferentially bind DNA with intermediate superhelical density and it also has been shown to protect DNA of intermediate superhelical density from relaxation by topoisomerases I [Schneider et al., 1997]. Most of the effect of FIS is in fine tuning of transcription according to the prevailing situation, as it can both activate and repress the expression of a certain gene based on the growth phase. An example for this phenomenon can been seen in a work with osmE promoter by Bordes et al., [Bordes et al., 2002]. The effect of H-NS on transcription is more straightforward, it mainly acts as global repressor. In hns mutant 70% of the regulated genes are associated with Rep group while in fis mutant the Rep genes increased transiently only during exponential phase (Table 1 & Figure 14). Double-coded genes (Figure 14) from 60
Page 1 and 2:
Coordinated regulation of gene expr
Page 3 and 4:
Parts of this work has been publish
Page 5 and 6:
ABSTRACT promoter region determines
Page 7 and 8:
ACKNOWLEDGEMENTS first impression o
Page 9 and 10:
TABLE OF CONTENTS 2.2.5 Preparation
Page 11 and 12:
LIST OF TABLES List of tables Table
Page 13 and 14:
LIST OF FIGURES List of Figures Fig
Page 15 and 16:
ABBREVIATIONS Abbreviations ∆61
Page 17 and 18:
ABBREVIATIONS TBE Tris TS tyrT35U t
Page 19 and 20:
INTRODUCTION Salmonella: “In more
Page 21 and 22:
INTRODUCTION Figure 1: Schematic il
Page 23 and 24:
INTRODUCTION UP element -35 -10 Spa
Page 25 and 26: INTRODUCTION can also influence elo
Page 27 and 28: INTRODUCTION have more base-pairs t
Page 29 and 30: INTRODUCTION Figure 6: Graphical re
Page 31 and 32: INTRODUCTION shown that H-NS reache
Page 33 and 34: INTRODUCTION formed as a repercussi
Page 35 and 36: MATERIALS AND METHODS 2 Materials a
Page 37 and 38: MATERIALS AND METHODS NaCl 10 gm YT
Page 39 and 40: MATERIALS AND METHODS LZ41 3 pyrF28
Page 41 and 42: MATERIALS AND METHODS 2.2 Methods 2
Page 43 and 44: MATERIALS AND METHODS given using a
Page 45 and 46: MATERIALS AND METHODS phenol: chlor
Page 47 and 48: MATERIALS AND METHODS Filtered (by
Page 49 and 50: MATERIALS AND METHODS free water (A
Page 51 and 52: RESULTS 3 Results Growth phase-depe
Page 53 and 54: RESULTS Neither the fis nor the hns
Page 55 and 56: RESULTS these identified genes (by
Page 57 and 58: RESULTS furthermore the colour of t
Page 59 and 60: RESULTS During the entire growth cy
Page 61 and 62: RESULTS transcripts of a growth pha
Page 63 and 64: RESULTS phosphorylation, are associ
Page 65 and 66: RESULTS Table 4: RT-PCR analysis of
Page 67 and 68: RESULTS Figure 16: Schematic repres
Page 69 and 70: RESULTS parC) which allows the supe
Page 71 and 72: RESULTS Figure 19: In vitro transcr
Page 73 and 74: RESULTS contrast the tyrTD promoter
Page 75: DISCUSSION 4 Discussion The idea of
Page 79 and 80: DISCUSSION cells maintain viability
Page 81 and 82: DISCUSSION writhe in the UAS by FIS
Page 83 and 84: DISCUSSION 4.3.2 Mechanism of trans
Page 85 and 86: CONCLUSION 5 Conclusion Regulation
Page 87 and 88: REFERENCES Bordes, P., Conter, A.,
Page 89 and 90: REFERENCES Gruber, T.M., and Gross,
Page 91 and 92: REFERENCES molecular basis for sele
Page 93 and 94: REFERENCES Pan, C.Q., Finkel, S.E.,
Page 95 and 96: REFERENCES Schroder, O., and Wagner
Page 97 and 98: REFERENCES Willenbrock, H., and Uss
Page 99 and 100: APPENDIX I gene blattner ratio p-va
Page 127 and 128:
APPENDIX I Supplementary table - VI
Page 129 and 130:
APPENDIX I fabB b2323 0.6336 0.0043
Page 131 and 132:
APPENDIX I ibpA b3687 2.4960 0.0181
Page 133 and 134:
APPENDIX I pepP b2908 1.2182 0.0368
Page 135 and 136:
APPENDIX I speG b1584 0.4151 0.0001
Page 137 and 138:
APPENDIX I ycjX b1321 4.0841 0.0141
Page 139 and 140:
APPENDIX I yhfY b3382 1.4063 0.0453
Page 141 and 142:
APPENDIX I gene blattner ratio p-va
Page 143 and 144:
Page 145 and 146:
Page 147 and 148:
Page 149 and 150:
Page 151 and 152:
Page 153 and 154:
Page 155 and 156:
Page 157 and 158:
APPENDIX I gene blattner ratio A/B
Page 159 and 160:
Page 161 and 162:
Page 163 and 164:
Page 165 and 166:
Page 167 and 168:
Page 169 and 170:
Page 171 and 172:
Page 173 and 174:
Page 175 and 176:
Page 177 and 178:
Page 179 and 180:
Page 181 and 182:
Page 183 and 184:
Page 185 and 186:
Page 187 and 188:
Page 189 and 190:
Page 191 and 192:
Page 193 and 194:
Page 195 and 196:
Page 197 and 198:
Page 199 and 200:
Page 201 and 202:
Page 203 and 204:
Page 205 and 206:
Statement of sources Declaration I
Page 207:
190
show all

Coordinated regulation of gene expression by E ... - Jacobs University

Create successful ePaper yourself

Delete template?

Save as template?