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Chapter 13
Regulation of Cellular
Processes
1
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Two Approaches to Regulation
• regulation of gene expression
– transcription initiation
– transcription elongation
– translation
• alter activity of enzymes and proteins
– posttranslational
• three domains of life differ in genome
structure and regulatory mechanisms
used
2
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Steps Leading from the
Information Coded in DNA to a
Functional Protein
Bacteria
Archaea
3
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Figure 13.1
4
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Regulation of Transcription
Initiation
• replacement of degraded enzymes
– constitutive genes
• are housekeeping genes that are expressed
continuously by the cell
– inducible genes
• are genes that code for inducible enzymes
needed only in certain environments
– such as b-Galactosidase
5
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Inducible Genes
b-Galactosidase Enzyme
• inducible enzyme functions in a catabolic
pathway
• inducible enzymes are present only when
their substrate (inducer - effector
molecule) is available
• β-galactosidase reaction catalyzed is
lactose hydrolysis into galactose and
glucose
6
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Figure 13.2
7
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Repressible Genes
• enzymes that function in
biosynthetic pathways are products
of repressible genes
• generally these enzymes are always
present unless the end product in the
biosynthetic pathway is available
8
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Control of Transcription
Initiation by Regulatory
Proteins
• induction and repression occur
because of the activity of regulatory
proteins and DNA binding domains
• these proteins either inhibit
transcription (negative control) or
promote transcription (positive
control)
9
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Negative Transcriptional
Control
• binding of regulatory protein (repressor)
at DNA regulatory site (operator) inhibits
initiation of transcription
– mRNA expression is reduced
• repressor proteins
– exist in active and inactive forms
– inducers (substrates) and corepressors
(enzymatic products) alter activity of
repressor by binding
10
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Figure 13.3(a) and (b)
11
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Positive Control
• binding of a regulatory protein (activator
protein) at a regulatory region on DNA
(activator binding sites) promotes
transcription initiation
– mRNA synthesis is increased
• activation
– inactive protein is activated by inducer
(activator protein)
– active protein is inactivated by inhibitor
12
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Figure 13.3(c) and (d)
13
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“Decision” Process in Gene
Expression
• enzymes of a catabolite pathway are
only needed (increased mRNA
synthesis) when the preferred
substrate is available
• enzymes not synthesized when
substrate absent
• efficient use of energy and materials
14
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Figure 13.4
15
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Operon Structure in Archaea
and Bacteria
• promoter and operator or activatorbinding sites, along with functionally
related structural genes, are grouped
together in the DNA
• polycistronic mRNA is produced
• regulatory proteins control gene
expression
16
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Negative Control of Lactose
(Lac) Operon
• inducible genes
– three structural genes coding for
lactose uptake and metabolism
– lac repressor (lacI) binds operator
• inhibits transcription
• enzymes normally not produced
unless lactose present
17
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lac Operon
Figure 13.5
18
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lac Repressor
• tetramers of repressor form and
bind to three operator sites
– O1, O2, O3
• bend DNA and prevent RNA
polymerase from accessing promoter
• presence of allolactose binds
repressor – no longer binds operator
19
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Figure 13.6
20
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Regulation of the lac Operon
by the lac Repressor
Figure 13.7
21
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Positive Control of the lac
Operon
• regulated by catabolite activator
protein (CAP)
– regulates in response to presence or
absence of glucose
– allows for preferential use of glucose
22
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The Tryptophan (trp) Operon
• consists of 5 structural genes which
code for enzymes needed to
synthesize tryptophan
• negative transcriptional control of
repressible genes by trp repressor
• the operon functions only in the
absence of tryptophan
23
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Regulation of the trp Operon by
Tryptophan and the trp Repressor
Figure 13.8
24
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The Arabinose (ara) Operon
• transcriptional control by a protein
(AraC) that acts both positively and
negatively
– activity depends on environmental
conditions
– inactive when arabinose present
– active when arabinose absent
25
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Positive and Negative Control
of ara Operon by AraC
• Figure 13.9
26
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Two-Component Regulatory
Systems and Phosphorelay
Systems
• many genes and operons are turned
on or off in response to
environmental conditions
– the regulatory proteins involved are
part of a two-component signal system
which links external events to
regulation of gene expression
27
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Two Component Regulatory
System
• found in all three domains of life
• two proteins govern pathway
– sensor kinase
• extracellular receptor for metabolite
• intracellular communication pathway
– response-regulator protein
• activated by sensor kinase
• DNA binding protein
– activator – enhances transcription needed
– repressor – inhibits transcription unless needed
28
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Regulation of Porin Proteins by a Two
Component Signal Transduction
System
Figure 13.10
29
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Phosphorelay System of Porin
Proteins
• Env Z (sensor kinase)
– autophosphorylates in high osmolarity
• OmpR (response regulator)
– phosphorylated and regulates transcription
• Regulate expression of porin proteins
(OmpC and OmpF) depending on
osmolarity
30
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Porin Protein Functions
• OmpC is smaller porin protein
– dominant when E. coli is in high
osmolarity intestinal tract
– lower levels of diffusion
• Omp F is larger porin protein
– dominant when E. coli is in dilute
environment
– allows more diffusion of solutes
31
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Regulation of Transcription
Elongation
• transcription can also be regulated by
controlling transcription termination
• this type of regulation, called attenuation,
was first demonstrated with trp operon
• more recently riboswitches have been
demonstrated to also play a regulatory
role
32
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Trp Operon Attenuation
• in addition to transcription initiation
control, transcription continuation is
also controlled in this operon
• attenuation is termination of
transcription within the leader
region (leader peptide)
• occurs through stem-loop structures
in the mRNA depending on trp level
33
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Attenuation of the trp Operon
Figure 13.11
34
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Riboswitches (Sensory RNAs)
• a specialized form of transcription
attenuation
• folding of mRNA leader sequence (the
riboswitch) determines if transcription
will continue or be terminated
• folding pattern altered in response to
mRNA binding of an effector molecule
• riboswitches in gram-positive bacteria
function in transcriptional termination
35
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Riboswitch of the Riboflavin
(rib) Operon of Bacillus subtilis
Figure 13.12
36
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Regulation of Gene Expression
by Riboswitches
Table 13.1
37
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Regulation of Translation
• riboswitches in gram-negative
bacteria regulate translation of
mRNA
– effector binding elements at 5’ end
alters mRNA leader folding pattern
• translation initiation can also be
controlled by some small RNA
molecules
38
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Regulation of Translation by a
Riboswitch
Figure 13.13
39
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Regulation of Translation by
Small RNA Molecules
• small (sRNAs) or noncoding
(ncRNAs) RNAs
– do not function as mRNA, tRNA, or
rRNA
– some (antisense RNAs) are
complementary to mRNA and function
by base pairing
– may inhibit or enhance translation
40
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Translation Regulation by
Antisense RNA
Figure 13.14
41
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Regulation of Gene Expression
by Small Regulatory RNAs
Table 12.2
42
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Posttranslational Regulation
• regulatory enzymes control
chemotaxis proteins by
– phosphorelay system and
– covalent modification
• irreversible (proteolysis)
• reversible
– methylation/demethylation
– phosphorylation/dephosphorylation
43
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Chemotaxis in E. coli
• Methyl-accepting chemotaxis proteins
(MCPs) are chemoreceptors in membrane
– binds environmental chemicals
– initiates a series of interactions with cytoplasmic
proteins that affects flagellar rotation
• activates sensor kinase CheA (with help of
CheW) which autophosphorylates
• CheA phophorylates the response regulator
CheY
• CheY governs rotation of flagella
44
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Figure 13.15
45
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Attractant Concentration Is
High
• MCP is methylated (CheR)
• CheA/CheW is active (phosphorylated)
• CheY is active
– diffuses to flagellar motor and switches to
clockwise (CW) rotation
– tumbling occurs
• concentration of attractant is measured
every few seconds (receptors reset)
46
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Attractant Concentration
Decreases
• MCP is not methylated (CheB)
• CheA is inactive (not phosphorylated)
– CheY is inactive
– flagellar rotation is counter-clockwise (CCW)
and running occurs
• regulatory system allows E. coli to
respond to and adapt to very small
amounts of attractant
47
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Methyl-Accepting Chemotaxis
Proteins in E. coli Membrane
Figure 13.16
48
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Global Regulatory Systems
• regulatory systems that affect many
genes and pathways simultaneously
• important for bacteria since they
must respond rapidly to a wide
variety of changing environmental
conditions
49
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Specialized Nomenclature
• regulon
– genes or operons controlled by a
common regulatory protein
• modulon
– operon network under control of a
common global regulatory protein but
individual operons are controlled
separately by their own regulators
50
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Mechanisms Used for Global
Regulation
• global regulatory systems often use
many types of regulation such as:
– regulatory proteins
– alternative sigma factors
– two component signal transduction
systems
– phosphorelay systems
51
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Table 13.3
52
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Catabolite Repression
• regulation of transcription by both
repressors and activators
• diauxic growth
– a biphasic growth pattern
• preferential use of one carbon source over
another when both are available in environment
– lag occurs
• after preferred substrate is exhausted followed
by resumption of growth using the second source
• catabolite repression plays a role in this
pattern of growth
53
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Figure 13.7
54
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Catabolite Activator Protein
(CAP)
• also called cyclic AMP receptor
protein (CRP)
• brings about the coordinate
regulation of catabolite operons
• exists in two forms
– active form when 3’,5’-cyclic adenosine
monophosphate (cAMP) is bound
– inactive form when it is free of cAMP
55
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Figure 13.18
56
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Figure 13.19
57
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CAP Regulation of lac Operon
• all catabolite operons contain a CAP
binding site
• CAP must be bound to this site
before RNA polymerase can bind the
promoter and begin transcription
58
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Regulation of cAMP - 1
• CAP activity is modulated by cAMP
• levels of cAMP controlled by adenyl
cyclase (converts ATP to cAMP and
PPi) by PEP
– adenyl cyclase active only when little or
no glucose is present
– in absence of glucose, CAP is active
and promotes transcription of operons
used for catabolism of other sugars
59
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Regulation of the lac Operon by the lac
Repressor and CAP
Figure 13.20
60
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Regulation of cAMP - 2
• cAMP levels are also increased by the
phosphoenolpyruvate and sugar
phosphotransferase system
• when glucose is available
– enzyme system transfers phosphorus to
glucose
• when glucose is not available
– enzyme system transfers phosphorus to
adenyl cyclase and cAMP is made
61
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Figure 3.21
62
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Regulation by Other
Nucleotides
• guanine tetraphosphate (ppGpp)
– stringent response of E. coli
• cyclic dimeric GMP (c-di-GMP)
– regulates virulence genes
– regulates transition form motile to
nonmotile lifestyle in biofilm formation
63
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Figure 13.22
64
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Stringent Response
• cells are starved for amino acids
• protein synthesis cannot proceed
• cell decreases production of tRNA and
rRNA to conserve energy
– protein RelA downregulates synthesis
through production of pppGpp
– pppGpp and DksA destabilizes transition
initiation open complexes
• cell increases biosynthesis of needed
amino acids
65
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Figure 13.23
66
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Figure 13.24
67
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Quorum Sensing
• cell-to-cell communication mediated
by small signaling molecules such as
N-acyl-homoserine lactone (AHL)
• couples cells density and
intercellular communication to
transcription regulation
68
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Quorum Sensing in V. fischeri
• high concentrations of the AHL produced
by increased density of cells diffuse back
into the cell and bind to the
transcriptional regulator LuxR and
activates transcription
• LuxR stimulates transcription of the
genes for AHL synthase (luxl) and the
proteins needed for light production
69
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Figure 13.25
70
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Response to Autoinducers by
V. harveyi
• responds to three autoinducers
– maximizes expression of bioluminescence
• low cell density
– low autoinducers present
– LuxR not made, no bioluminescence
• high cell density
– any combination of inducers
– LuxR made, bioluminescence occurs
71
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Figure 13.26
72
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Sporulation in Bacillus subtilis
• another example of a global regulatory
system in which phosphorelay,
posttranslational modification of
proteins, transcription initiation
regulatory proteins and alternative
sigma factors play a role
• starvation signal induces production of
alternative sigma factors
73
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Control of Sporulation in
Bacillus subtilis
Figure 13.27
74
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Regulation of Gene Expression
in Eukarya and Archaea
• occurs at transcriptional,
translational, and posttranslation
levels (figure 13.1)
75
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Eukarya Gene Expression
• eukarya use regulatory transcription
factors to regulate transcription
initiation
• activator proteins bind enhancers
– mRNA transcription increases
• repressor proteins bind silencers
– mRNA transcription decreases
76
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Eukarya Gene Expression - 2
• sRNA molecules control gene
expression
• antisense RNAs and micro RNAs
(miRNAs) regulate translation
• sRNAs also associate with
spliceosome and increase alternative
splicing for new proteins
77
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Figure 13.28
78
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Regulation of Gene Expression
in Archaea
• most Archaea regulatory proteins function
like Bacteria activators and repressors
– they bind DNA sites near the promoter,
enhancing or blocking the binding of RNA
polymerase
• some regulatory proteins function like
Eukarya regulatory transcription factors
by interacting with general transcription
factors
79