A Genetic Switch: Phage Lambda Revisited

INTRODUCTION
CHAPTER ONE
THE MASTER ELEMENTS OF CONTROL
Components of the Switch
DNA
RNA Polymerase
The Repressor
Cro
The Action of Repressor and Cro
Negative Control
Positive Control
Cooperativity of Repressor Binding
Induction—Flipping the Switch
Cooperativity—Switch Stability and Sensitivity
The Effect of Autoregulation
Other Cases
CHAPTER TWO
PROTEIN-DNA INTERACTIONS AND GENE CONTROL
The Operator
Repressor
Cro
Amino Acid-Base Pair Interactions
The Promoter
Gene Control
CHAPTER THREE
CONTROL CIRCUITS—SETTING THE SWITCH
A Brief Overview of λ Growth
The Genetic Map
Circularization
Gene Expression
Integration
Control of Transcription
Very Early
Early
Late Lytic
Late Lysogenic
The Decision
Control of Integration and Excision
Case 1—Establishing Lysogeny
Case 2—Lytic Growth
Case 3—Induction
Other Phages
The SOS Response
λ Pathways and Cell Development
Regulatory Genes
Switches
Patterns of Gene Expression
CHAPTER FOUR
HOW DO WE KNOW—THE KEY EXPERIMENTS
The Repressor Idea
Clear and Virulent Mutants
Observations
Explanation
Immunity and Heteroimmunity
Observations
Explanation
Asymmetry in Bacterial Mating
Observations
Explanation
The Repressor Problem in the Early 1960s
Repressor Isolation and DNA Binding
Making More Repressor
The Claims of Chapters One and Two
The repressor is composed of two globular domains held together by a linker of some 40
amino acids
The repressor dimerizes, largely through interaction between its carboxyl domains
A repressor dimer binds, through its amino domains, to a 17 base pair operator site
A single operator site binds one dimer of repressor
Dimers form before DNA binding
The amino domains contact DNA
There are three 17 base pair repressor binding sites in the right operator. At each site
repressor and Cro bind along the same face of the helix
Chemical probes
Operator mutations
Binding to supercoiled and linear DNA
Repressor binds to three sites in OR with alternate pairwise cooperativity. The cooperativity
is mediated by interactions between carboxyl domains of adjacent dimers
In a lysogen repressor is typically bound to OR1 and OR2. The bound repressors turn off
rightward transcription of cro and stimulate leftward transcription of cI. At higher
concentrations, repressor binds to OR3 to turn off transcription of cI
Cro binds first to OR3, then to OR1 and OR2, thereby first turning off PRM, then PR
Some background about Cro
Cro in vivo
Cro in vitro
RecA cleaves repressor to trigger induction
When Cro is bound at OR3 the switch is thrown
Repressor and Cro bind to the operator as shown in Figures 2.6, 2.8, 2.10, and 2.11
Crystallography
The “helix swap” experiment
Specific amino acid-base pair contacts
The role of the arm of λ repressor
Repressor activates transcription of cI by binding to OR2 and contacting polymerase with its
amino domain
Positive control mutants
Positive control in vitro
Conclusion
CHAPTER FIVE
2004: NEW DEVELOPMENTS
1. Long-range Cooperativity and Repression of PRM
An Octamer of Repressor Binds OR and OL
Autonegative Regulation of Repressor Synthesis
How Do We Know
Long-range Interactions and Repression of PR
Long-range Interactions and Repression of PRM
Activation and Repression of PRM
Repressor Structure
2. Positive Control (Activation of Transcription)
Polymerase and Promoter
The Mechanism of Activation
How Do We Know
Activating Region Variants
A Suppressor of a pc Mutant
Crystallography
Activator Bypass
Changing Activating Regions and Target Context
3. The Structure of the Repressor Monomer and the Mechanism of Repressor Cleavage
How Do We Know
4. Evolving the Switch
Changing the Affinities of Sites in OR for Repressor
Eliminating Positive Control
Eliminating Cooperativity between DNA-binding Dimers
5. CII and the Decision