Transcript 2/17/12 Gene regulation

8.1 Major Modes of Regulation
• Gene expression: transcription of gene into mRNA
followed by translation of mRNA into protein
(Figure 8.1)
• Most proteins are enzymes that carry out
biochemical reactions
• Constitutive proteins are needed at the same level
all the time
• Microbial genomes encode many proteins that are
not needed all the time
• Regulation helps conserve energy and resources
© 2012 Pearson Education, Inc.
Figure 8.1
Upstream
region
Downstream
region
DNA
Start of
transcription
Shine-Dalgarno
sequence (ribosomebinding site)
Transcription
terminator
Transcription
mRNA
Start codon:
Translation
starts here
Stop codon:
Translation
ends here
Translation
Protein
© 2012 Pearson Education, Inc.
8.2 DNA-Binding Proteins
• mRNA transcripts generally have a short half-life
– Prevents the production of unneeded proteins
• Regulation of transcription typically requires
proteins that can bind to DNA
• Small molecules influence the binding of
regulatory proteins to DNA
– Proteins actually regulate transcription
© 2012 Pearson Education, Inc.
8.2 DNA-Binding Proteins
• Most DNA-binding proteins interact with DNA in a
sequence-specific manner
• Specificity provided by interactions between amino
acid side chains and chemical groups on the bases
and sugar–phosphate backbone of DNA
• Major groove of DNA is the main site of protein
binding
• Inverted repeats frequently are binding site for
regulatory proteins
© 2012 Pearson Education, Inc.
8.2 DNA-Binding Proteins
• Homodimeric proteins: proteins composed of
two identical polypeptides
• Protein dimers interact with inverted repeats
on DNA
– Each of the polypeptides binds to one inverted
repeat (Figure 8.2)
© 2012 Pearson Education, Inc.
Figure 8.2
Domain containing protein–protein
contacts, holding protein dimer together
DNA-binding domain fits in
major grooves and along
sugar–phosphate backbone
Inverted repeats
Inverted repeats
© 2012 Pearson Education, Inc.
8.2 DNA-Binding Proteins
• Several classes of protein domains are critical for
proper binding of proteins to DNA
– Helix-turn-helix (Figure 8.3)
• First helix is the recognition helix
• Second helix is the stabilizing helix
• Many different DNA-binding proteins from Bacteria
contain helix-turn-helix
– lac and trp repressors of E. coli
© 2012 Pearson Education, Inc.
Figure 8.3
Stabilizing
helix
Turn
Recognition
helix
DNA
Subunits
of binding
protein
© 2012 Pearson Education, Inc.
8.2 DNA-Binding Proteins
•
Multiple outcomes after DNA binding are
possible
1. DNA-binding protein may catalyze a specific
reaction on the DNA molecule (i.e.,
transcription by RNA polymerase)
2. The binding event can block transcription
(negative regulation)
3. The binding event can activate transcription
(positive regulation)
© 2012 Pearson Education, Inc.
8.3 Negative Control of Transcription:
Repression and Induction
•
Several mechanisms for controlling gene
expression in bacteria
–
–
–
These systems are greatly influenced by
environment in which the organism is growing
Presence or absence of specific small
molecules
Interactions between small molecules and DNAbinding proteins result in control of transcription
or translation
© 2012 Pearson Education, Inc.
8.3 Negative Control of Transcription:
Repression and Induction
• Negative control: a regulatory mechanism
that stops transcription
– Repression: preventing the synthesis of an
enzyme in response to a signal (Figure 8.5)
• Enzymes affected by repression make up a
small fraction of total proteins
• Typically affects anabolic enzymes
(e.g., arginine biosynthesis)
© 2012 Pearson Education, Inc.
8.3 Negative Control of Transcription:
Repression and Induction
• Negative Control (cont’d)
– Induction: production of an enzyme in response to
a signal (Figure 8.6)
• Typically affects catabolic enzymes (e.g., lac
operon)
• Enzymes are synthesized only when they are
needed
– no wasted energy
© 2012 Pearson Education, Inc.
Figure 8.5
Relative increase
Repression
Cell number
Total
protein
Arginine added
Arginine
biosynthesis
enzymes
Time
© 2012 Pearson Education, Inc.
Figure 8.6
Induction
Relative increase
Total protein
Cell number
-Galactosidase
Lactose added
Time
© 2012 Pearson Education, Inc.
8.3 Negative Control of Transcription:
Repression and Induction
• Inducer: substance that induces enzyme synthesis
• Corepressor: substance that represses enzyme
synthesis
• Effectors: collective term for inducers and
repressors
• Effectors affect transcription indirectly by binding
to specific DNA-binding proteins
– Repressor molecules bind to an allosteric
repressor protein
– Allosteric repressor becomes active and binds to
region of DNA near promoter called the operator
© 2012 Pearson Education, Inc.
Figure 8.7
arg Promoter arg Operator
argC
RNA
polymerase
argB
argH
Transcription proceeds
Repressor
arg Promoter arg Operator
RNA
polymerase
argC
argH
Corepressor
Transcription blocked
(arginine)
Repressor
© 2012 Pearson Education, Inc.
argB
Figure 8.8
lac Promoter lac Operator
RNA
polymerase
lac Promoter lac Operator
RNA
polymerase
lacZ
lacY
Transcription blocked
Repressor
lacZ
lacY
lacA
Transcription proceeds
Repressor
Inducer
© 2012 Pearson Education, Inc.
lacA
8.4 Positive Control of Transcription
• Positive control: regulator protein activates the
binding of RNA polymerase to DNA (Figure 8.9)
• Maltose catabolism in E. coli
– Maltose activator protein cannot bind to DNA
unless it first binds maltose
• Activator proteins bind specifically to certain DNA
sequence
– Called activator-binding site, not operator
© 2012 Pearson Education, Inc.
Figure 8.9
Activatorbinding site
mal Promoter
malE
malF
malG
No transcription
RNA
polymerase
Maltose activator protein
Activatorbinding site
mal Promoter
RNA
polymerase
malE
malF
Transcription proceeds
Maltose activator protein
Inducer
© 2012 Pearson Education, Inc.
malG
8.4 Positive Control of Transcription
• Promoters of positively controlled operons only
weakly bind RNA polymerase
• Activator protein helps RNA polymerase
recognize promoter
– May cause a change in DNA structure
– May interact directly with RNA polymerase
• Activator-binding site may be close to the
promoter or several hundred base pairs away
(Figure 8.11)
© 2012 Pearson Education, Inc.
Figure 8.11
Activatorbinding site
Promoter
RNA
polymerase
Activator
protein
Activator protein
Promoter
RNA
polymerase
Transcription
proceeds
Transcription
proceeds
Activatorbinding site
© 2012 Pearson Education, Inc.
8.5 Global Control and the lac Operon
• Cyclic AMP and CRP
– In catabolite repression, transcription is controlled
by an activator protein and is a form of positive
control (Figure 8.14)
– Cyclic AMP receptor protein (CRP) is the
activator protein
– Cyclic AMP is a key molecule in many metabolic
control systems
• It is derived from a nucleic acid precursor
• It is a regulatory nucleotide
© 2012 Pearson Education, Inc.
Figure 8.14
CRP protein
cAMP
RNA
polymerase
lac Structural genes
DNA
Transcription
mRNA
lacI
Active
repressor
binds to
operator
and
blocks
transcription
Transcription
mRNA lacZ
lacY
lacA
Translation
Translation
Inducer
Active
repressor
Lactose catabolism
Inactive
repressor
© 2012 Pearson Education, Inc.
8.5 Global Control and the lac Operon
• Dozens of catabolic operons affected by
catabolite repression
– Enzymes for degrading lactose, maltose, and
other common carbon sources
• Flagellar genes are also controlled by
catabolite repression
– No need to swim in search of nutrients
© 2012 Pearson Education, Inc.
8.7 Two-Component Regulatory Systems
• Prokaryotes regulate cellular metabolism in
response to environmental fluctuations
– External signal is transmitted directly to the target
– External signal detected by sensor and
transmitted to regulatory machinery (Signal
transduction)
• Most signal transduction systems are twocomponent regulatory systems
© 2012 Pearson Education, Inc.
8.7 Two-Component Regulatory Systems
• Two-component regulatory systems (Figure 8.16)
– Made up of two different proteins:
• Sensor kinase: (in cytoplasmic membrane) detects
environmental signal and autophosphorylates
• Response regulator: (in cytoplasm) DNA-binding
protein that regulates transcription
– Also has feedback loop
• Terminates signal
© 2012 Pearson Education, Inc.
Figure 8.16
Environmental signal
Sensor
kinase
Cytoplasmic
membrane
Response regulator
Phosphatase
activity
RNA
polymerase
Promoter Operator
© 2012 Pearson Education, Inc.
Transcription blocked
Structural genes
DNA
8.7 Two-Component Regulatory Systems
• Almost 50 different two-component systems
in E. coli
– Examples include phosphate assimilation,
nitrogen metabolism, and osmotic pressure
response
• Some signal transduction systems have
multiple regulatory elements
• Some Archaea also have two-component
regulatory systems
© 2012 Pearson Education, Inc.
8.9 Quorum Sensing
• Prokaryotes can respond to the presence of other
cells of the same species
• Quorum sensing: mechanism by which bacteria
assess their population density
– Ensures sufficient number of cells are present
before initiating a response that requires a certain
cell density to have an effect (e.g., toxin
production in pathogenic bacterium)
© 2012 Pearson Education, Inc.
8.9 Quorum Sensing
• Each species of bacterium produces a
specific autoinducer molecule (Figure 8.18)
– Diffuses freely across the cell envelope
– Reaches high concentrations inside cell only if
many cells are near
– Binds to specific activator protein and triggers
transcription of specific genes
© 2012 Pearson Education, Inc.
Figure 8.18
Acyl homoserine lactone (AHL)
AHL
Other cells
of the same
species
Activator protein
Quorumspecific
proteins
Chromosome
© 2012 Pearson Education, Inc.
AHL
AHL synthase
8.9 Quorum Sensing
• Several different classes of autoinducers
– Acyl homoserine lactone was the first
autoinducer to be identified
• Quorum sensing first discovered as mechanism
regulating light production in bacteria including
Aliivibrio fischeri (Figure 8.19)
– Lux operon encodes bioluminescence
© 2012 Pearson Education, Inc.
Figure 8.19
© 2012 Pearson Education, Inc.
8.9 Quorum Sensing
• Examples of quorum sensing
– P. aeruginosa switches from free living to
growing as a biofilm
– Virulence factors of Staphylococcus aureus
• Quorum sensing is present in some microbial
eukaryotes
• Quorum sensing likely exists in Archaea
© 2012 Pearson Education, Inc.
8.11 Other Global Control Networks
• Several other global control systems
–
–
–
–
–
–
Aerobic and anaerobic respiration
Catabolite repression
Nitrogen utilization
Oxidative stress
SOS response
Heat shock response
© 2012 Pearson Education, Inc.
8.11 Other Global Control Networks
• Heat shock response:
– Largely controlled by alternative sigma factors
(Figure 8.21)
– Heat shock proteins: counteract damage of
denatured proteins and help cell recover from
temperature stress
• Very ancient proteins
• Heat shock response also occurs in Archaea
© 2012 Pearson Education, Inc.
Low temperature
Figure 8.21
DnaK
RpoH
Proteins unfold at
high temperature
Degradation of
RpoH by protease
High temperature
RpoH is
released
© 2012 Pearson Education, Inc.
RpoH
DnaK binds
unfolded
proteins
RpoH is free
to transcribe
heat shock
genes
IV. Regulation of Development in
Model Bacteria
• 8.12 Sporulation in Bacillus
• 8.13 Caulobacter Differentiation
© 2012 Pearson Education, Inc.
8.12 Sporulation in Bacillus
• Regulation of development in model bacteria
– Some prokaryotes display the basic principle of
differentiation
• Endospore formation in Bacillus (Figure 8.22)
– Controlled by 4 sigma factors
– Forms inside mother cell
– Triggered by adverse external conditions (i.e.,
starvation or desiccation)
© 2012 Pearson Education, Inc.