The Fitness Landscapes of -Acting Binding Sites in Different Promoter and Environmental Contexts

English version České info

The biophysical nature of the interaction between a transcription factor and its target sequences in vitro is sufficiently well understood to allow for the effects of DNA sequence alterations on affinity to be predicted. But even in relatively simple in vivo systems, the complexities of promoter organization and activity have made it difficult to predict how altering specific interactions between a transcription factor and DNA will affect promoter output. To better understand this, we measured the relative fitness of nearly all Escherichia coli binding sites in different promoter and environmental contexts by competing four randomized promoter libraries controlling the expression of the tetracycline resistance gene (tet) against each other in increasing concentrations of drug. We sequenced populations after competition to determine the relative enrichment of each −35 sequence. We observed a consistent relationship between the frequency of recovery of each −35 binding site and its predicted affinity for that varied depending on the sequence context of the promoter and drug concentration. Overall the relative fitness of each promoter could be predicted by a simple thermodynamic model of transcriptional regulation, in which the rate of transcriptional initiation (and hence fitness) is dependent upon the overall stability of the initiation complex, which in turn is dependent upon the energetic contributions of all sites within the complex. As implied by this model, a decrease in the free energy of association at one site could be compensated for by an increase in the binding energy at another to produce a similar output. Furthermore, these data show that a large and continuous range of transcriptional outputs can be accessed by merely changing the , suggesting that evolved or engineered mutations at this site could allow for subtle and precise control over gene expression.

Vyšlo v časopise: The Fitness Landscapes of -Acting Binding Sites in Different Promoter and Environmental Contexts. PLoS Genet 6(7): e32767. doi:10.1371/journal.pgen.1001042
Kategorie: Research Article
prolekare.web.journal.doi_sk: https://doi.org/10.1371/journal.pgen.1001042

Souhrn

Zdroje

1. von HippelPH

BergOG

1986 On the specificity of DNA-protein interactions. Proc Natl Acad Sci USA 83 1608 1612

2. SchneiderTD

1997 Information content of individual genetic sequences. J Theor Biol 189 427 441

3. MaerklSJ

QuakeSR

2007 A systems approach to measuring the binding energy landscapes of transcription factors. Science 315 233 237

4. ShultzabergerRK

RobertsLR

LyakhovIG

SidorovIA

StephenAG

2007 Correlation between binding rate constants and individual information of E. coli Fis binding sites. Nucleic Acids Res 35 5275 5283

5. DombroskiAJ

JohnsonBD

LonettoM

GrossCA

1996 The sigma subunit of Escherichia coli RNA polymerase senses promoter spacing. Proc Natl Acad Sci USA 93 8858 8862

6. MartinRG

GilletteWK

RheeS

RosnerJL

1999 Structural requirements for marbox function in transcriptional activation of mar/sox/rob regulon promoters in Escherichia coli: sequence, orientation and spatial relationship to the core promoter. Mol Microbiol 34 431 441

7. HawleyDK

McClureWR

1983 Compilation and analysis of Escherichia coli promoter DNA sequences. Nucleic Acids Res 11 2237 2255

8. McClureWR

1985 Mechanism and control of transcription initiation in prokaryotes. Annu Rev Biochem 54 171 204

9. BrowningDF

BusbySJ

2004 The regulation of bacterial transcription initiation. Nat Rev Microbiol 2 57 65

10. BintuL

BuchlerN

GarciaH

GerlandU

HwaT

2005 Transcriptional regulation by the numbers: models. Current opinion in genetics & development 15 116 124

11. ShultzabergerRK

ChenZ

LewisKA

SchneiderTD

2007 Anatomy of Escherichia coli σ70 promoters. Nucleic Acids Res 35 771 788

12. GertzJ

SiggiaE

CohenB

2008 Analysis of combinatorial cis-regulation in synthetic and genomic promoters

13. MustonenV

KinneyJ

CallanC

LassigM

2008 Energy-dependent fitness: A quantitative model for the evolution of yeast transcription factor binding sites. Proceedings of the National Academy of Sciences 105 12376

14. FryC

FarnhamP

1999 Context-dependent transcriptional regulation

15. HochschildA

DoveS

1998 Protein–Protein Contacts Minireview that Activate and Repress Prokaryotic Transcription. Cell 92 597 600

16. RoyS

GargesS

AdhyaS

1998 Activation and repression of transcription by differential contact: two sides of a coin

17. KimJ

ShapiroD

1996 In simple synthetic promoters YY1-induced DNA bending is important in transcription activation and repression. Nucleic acids research 24 4341

18. SheridanS

BenhamC

HatfieldG

1998 Activation of gene expression by a novel DNA structural transmission mechanism that requires supercoiling-induced DNA duplex destabilization in an upstream activating sequence. Journal of Biological Chemistry 273 21298 21308

19. MartinR

GilletteW

MartinN

RosnerJ

2002 Complex formation between activator and RNA polymerase as the basis for transcriptional activation by MarA and SoxS in Escherichia coli. Molecular Microbiology 43 355 370

20. ShultzabergerRK

ChiangDY

MosesAM

EisenMB

2007 Determining physical constraints in transcriptional initiation complexes using DNA sequence analysis. PLoS ONE 2 e1199 doi:10.1371/journal.pone.0001199

21. MandeckiW

ReznikoffWS

1982 A lac promotor with a changed distance between −10 and −35 regions. Nucleic Acids Res 10 903 912

22. MartinR

JairK

WolfR

RosnerJ

1996 Autoactivation of the marRAB multiple antibiotic resistance operon by the MarA transcriptional activator in Escherichia coli. Journal of bacteriology 178 2216 2223

23. SchneiderTD

StormoGD

GoldL

EhrenfeuchtA

1986 Information content of binding sites on nucleotide sequences. J Mol Biol 188 415 431

24. SchneiderTD

1991 Theory of molecular machines. II. Energy dissipation from molecular machines. J Theor Biol 148 125 137

25. DekelE

AlonU

2005 Optimality and evolutionary tuning of the expression level of a protein. Nature 436 588 592

26. NguyenT

PhanQ

DuongL

BertrandK

LenskiR

1989 Effects of carriage and expression of the TnlO tetracycline-resistance operon on the fitness of Escherichia coli K12. Molecular Biology and Evolution 6 213 225

27. LenskiR

SouzaV

DuongL

PhanQ

NguyenT

1994 Epistatic effects of promoter and repressor functions of the Tn10 tetracycline-resistance operon on the fitness of Escherichia coli. Molecular Ecology 3 127 135

28. EllingerT

BehnkeD

BujardH

GrallaJ

1994 Stalling of Escherichia coli RNA polymerase in the+ 6 to+ 12 region in vivo is associated with tight binding to consensus promoter elements. Journal of molecular biology 239 455 465

29. FentonMS

LeeSJ

GrallaJD

2000 Escherichia coli promoter opening and −10 recognition: mutational analysis of σ70. EMBO J 19 1130 1137

30. SclaviB

ZaychikovE

RogozinaA

WaltherF

BuckleM

2005 Real-time characterization of intermediates in the pathway to open complex formation by Escherichia coli RNA polymerase at the T7A1 promoter. Proc Natl Acad Sci USA 102 4706 4711

31. SambrookJ

FritschEF

ManiatisT

1989 Molecular Cloning, A Laboratory Manual Cold Spring Harbor, New York Cold Spring Harbor Laboratory, second edition

32. ZhengL

BaumannU

ReymondJ

2004 An efficient one-step site-directed and site-saturation mutagenesis protocol. Nucleic acids research 32 e115

33. SchneiderTD

StephensRM

1990 Sequence logos: A new way to display consensus sequences. Nucleic Acids Res 18 6097 6100

34. ShultzabergerRK

SchneiderTD

1999 Using sequence logos and information analysis of Lrp DNA binding sites to investigate discrepancies between natural selection and SELEX. Nucleic Acids Res 27 882 887