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Prokaryotic Gene Regulation
Prokaryotic gene regulation mechanisms allow bacteria to quickly adapt to their
environments.
Bacterial strains.
Scanning electron micrograph of a variety of species of bacteria from the human intestine.
Magnification is 8,000x.
David M. Phillips/Science Source.
Topics Covered in this Module
Gene Regulation in Bacteria
Major Objectives of this Module
Describe the organization of bacterial DNA into operons.
Explain the function of RNA polymerase.
Describe the role of riboswitches in regulating gene expression.
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Principles of Biology
51 Prokaryotic Gene Regulation
Gene Regulation in Bacteria
Prokaryotes, such as bacteria, lack a nuclear membrane and are generally
unicellular organisms. The lack of membrane-bound organelles means that
processes involved in genetic expression or regulation occur without physical
separation (Figure 1). In order for genes to be expressed at the right time
and location, gene expression must be regulated carefully.
Figure 1: Coupled transcription and translation in E. coli.
The long fiber running from left to right is a segment of the E. coli
chromosome. Transcription is occurring at multiple points along the DNA
where RNA polymerase attaches. mRNA is transcribed as the polymerase
moves along the DNA from left to right. Translation begins even while
transcription is still progressing; the ribosomes attach to the nascent
mRNA strands and assemble amino acids into polypeptide chains as they
move toward the DNA strand.
Professor Oscar Miller/Science Source.
RNA polymerases in prokaryotes and eukaryotes differ. Prokaryotes use a
single type of RNA polymerase, but eukaryotes have at least three different
types of RNA polymerase. All of the genetic information contained within
prokaryotes and eukaryotes is considered their genome. In some cases, the
cells interact with each other or their environment to regulate gene
expression.
Let's focus on a classical example of prokaryotic gene expression. Escherichia
coli is a bacterial species that is common in the human large intestine,
consuming nutrients provided by the host. Individual E. coli require a
continuous supply of certain amino acids, such as tryptophan, to survive.
However, tryptophan is not always available in the intestinal environment. As
a result, E. coli is capable of synthesizing tryptophan itself by activating a
metabolic pathway when it is unavailable in the environment. When
tryptophan again becomes available in the environment, the cell can
conserve energy by halting synthesis and using environmental tryptophan
contents
instead (Figure 2). This is a way that bacteria adjust their metabolism to
environmental changes.
Figure 2: Feedback inhibition in the E. coli tryptophan biosynthesis
pathway.
In E. coli, when tryptophan levels are low, tryptophan is synthesized from a
precursor by three enzyme complexes. When tryptophan is abundant in
the bacterium, the biosynthetic pathway undergoes feedback inhibition.
Tryptophan binds to the first enzyme complex, inhibiting the pathway from
producing additional tryptophan.
© 2014 Nature Education All rights reserved.
Figure Detail
Bacterial genes are organized into operons.
An operon is a set of genes that are transcribed together as a unit and
under the control of a single set of regulators. A basic regulatory region
consists of an operator and a promoter. The promoter is the region of the
DNA that has a specific sequence that the RNA polymerase binds to for
initiation of transcription. The ability of RNA polymerase to access the
promoter is regulated by the operator and transcription factors. An operator
is a specific sequence within the DNA that binds transcription factors to turn
transcription on or off. The operator is found within the promoter or between
the promoter and the coding region of the gene. The operator, promoter and
coding regions of the genes make up the operon.
The operon can be turned off by a repressor, a transcription factor that
binds to the operator and inhibits RNA polymerase from binding to the
promoter. Blocking RNA polymerase from accessing the promoter prevents
transcription from taking place. Repressor proteins have a specific binding
site on an operator, and therefore, have no effect on any other operator sites
within the given genome.
The regulatory genes that encode repressor proteins are continuously
expressed at a low rate. The operons regulated by the repressors are not
permanently shut down because the binding of a repressor to its operator is
reversible. A majority of regulatory proteins are allosteric proteins. Allosteric
proteins are proteins that change their shape when bound to a specific
molecule called an allosteric effector. This change in shape may activate
the allosteric protein to bind the operator sequence and shut down
expression of the operon or in other cases to let go of the operator and turn
on expression of the operon.
An example of this gene regulation can be seen with the lac operon, which
codes for three proteins (Figure 3). Import and metabolism of lactose is
controlled by these lac operon-encoded proteins. The presence and absence
of lactose regulates the expression of the lac operon. When lactose is absent
from the environment, the lactose repressor protein, LacI, will bind the
operator and repress the lac operon by preventing RNA polymerase from
accessing the promoter. The allosteric effector of LacI is allolactose, an
isomer of lactose. When lactose is present, the cell modifies some of it into
allolactose. In turn, allolactose binds to LacI and changes its conformation so
that it can no longer bind to the operator. Without the LacI repressor in the
way, the transcription of the three lac genes can proceed. This is an example
of gene expression regulation that allows the organism to adapt to
environmental changes quickly and efficiently.
Figure 3: The lac operon of E. coli.
In this operon, expression is controlled
by a repressor, an operator and RNA
polymerase. The lac operon of E. coli is a
segment of DNA that includes a
promoter (P), an operator (O), and the
three structural genes (lacZ, lacY, and
lacA) that code for proteins involved in
lactose import and lactose metabolism.
In the absence of lactose (a), the
lactose repressor protein, LacI (I),
which is encoded by the lacI gene, binds
to the operator and inhibits transcription
of the lac operon. The repressor protein
prevents RNA polymerase from
transcribing lacZ, lacY, and lacA. In the
presence of lactose (b), some lactose is
modified to allolactose, which binds to
the allosteric site of the LacI repressor
protein. This binding changes the
conformation of LacI, preventing it from
binding to the operator. The release of
LacI from the operator allows RNA
polymerase to transcribe lacZ, lacY, and
lacA.
© 2013 Nature Education All rights
reserved.
Gene regulation can be positive or negative. In negative gene regulation, the
primary function of a regulatory protein is to inhibit the expression of a gene.
Operons controlled by negative gene regulation can be categorized into
repressible operons or inducible operons depending on the effect of the
allosteric effector on repressor-operator binding. In repressible operons,
the allosteric effector causes the repressor to inhibit the expression of the
operon. An example of this is tryptophan and the trp operon (Figure 4).
Without tryptophan as an allosteric effector, the repressor cannot bind to the
operator, and transcription of the trp operon is allowed to proceed. When
tryptophan is present, however, it binds to the repressor, causing a
conformational change that allows it to bind to the operator and block
transcription of the trp operon. In effect, the operon is repressible by addition
of tryptophan to the environment.
In contrast, inducible operons inhibit the expression of the operon when the
allosteric effector is absent. However, the expression of the operon is turned
on, or induced, when the allosteric effector is present and binds to the
regulatory protein. An example of this is the LacI repressor and allolactose of
the lac operon. Without the allosteric effector allolactose, LacI remains bound
to the operator, preventing lac operon transcription. However, the presence of
lactose (and therefore allolactose) changes the conformation of LacI so that
it releases from the operator and permits lac operon transcription.
A third type of gene regulation is positive gene regulation, in which the
interaction between a protein and its allosteric effector results in the
activation of gene expression. The lac operon also includes an example of
positive gene regulation. In E. coli, the metabolite cyclic AMP (cAMP)
accumulates to high levels when glucose levels in the cell are low.
Conversely, cAMP levels fall when cellular glucose levels increase. cAMP is
the allosteric effector for an allosteric regulatory protein called catabolite
activator protein (CAP). In the absence of cAMP, CAP is inactive, but when
CAP is bound to cAMP, it changes into an active conformation, which is able
to bind a specific DNA sequence called the activator sequence. This
activator sequence is located upstream of the lac promoter. When the
activator sequence is not bound by CAP-cAMP, the affinity of RNA
polymerase for the adjacent promoter is low, but when CAP-cAMP is bound
to the activator sequence, the affinity of RNA polymerase for the promoter
increases, increasing the expression of the lac operon genes. Therefore, in
the absence of glucose, cAMP levels are high, and lac operon expression is
also high. In the presence of glucose, however, cAMP levels are low, and
therefore lac operon expression is low as well.
The overall benefit of negative and positive gene regulation of the lac operon
is that the expression levels of the lactose metabolism genes are directly
influenced by the nutritional environment of the cell. That is, when lactose is
absent, nucleotides, amino acids and energy are not wasted in expressing
the lactose metabolism genes. The inducible nature of the lac operon helps
ensure that the operon is not expressed unless lactose is present to induce
its expression. Furthermore, even if lactose is present, it would be wasteful
to express the lactose metabolism genes at maximal levels if other energy
sources — such as glucose — had not yet been exhausted. This wasteful
potential outcome is avoided thanks to positive gene regulation by the
CAP-cAMP system, which helps ensure that lac operon expression is
maximized only when glucose is absent and lactose is present.
Figure 4: Regulation of the trp operon.
The trp operon regulates the expression of five genes in response to
tryptophan levels inside the cell. The trpR gene, regulated separately from
the trp operon, encodes the Trp repressor protein. In the absence of
tryptophan (a), the Trp repressor is unable to bind to the operator of the trp
operon. As a result, RNA polymerase binds to the promoter and
transcribes five genes — trpE, trpD, trpC, trpB, and trpA — which encode
enzymes required for tryptophan synthesis. Tryptophan levels in the cell
increase as a result. Once tryptophan levels exceed a critical
concentration (b), tryptophan molecules bind to the allosteric sites of the
Trp repressor, changing its conformation. In its new conformation, the
repressor binds to the operator, blocking RNA polymerase from binding to
the promoter and preventing the expression of the five trp genes.
© 2014 Nature Education All rights reserved.
Bacterial transcription is regulated by a single type of RNA polymerase.
All transcription in bacteria is carried out by a single RNA polymerase, which
has five core subunits and a sigma subunit. RNA polymerase binds to
specific sequences on the DNA called promoters. What does the sigma
subunit do? In 1969, Andrew Travers and Richard Burgess discovered the
role of RNA polymerase sigma. They found that in in vitro assays of bacterial
transcription with template DNA and purified RNA polymerase, transcription
did not start at specific sites. In vivo, the RNA polymerase did start at specific
sites, even with a minimal amount of polymerase. This was because sigma
factor was not purified with the core RNA polymerase that was used in the in
vitro assays. Without sigma factor, the core RNA polymerase made errors in
start point choice, which led to RNA synthesis at inappropriate sites. Travers
and Burgess concluded that sigma factors were necessary for specificity in
the initiation of transcription (Figure 5).
Figure 5: The sigma cycle.
Sigma factors are necessary for
specificity in the initiation of
transcription. The core RNA
polymerase joins with the sigma factor
and attaches to the DNA. After RNA
synthesis is initiated, RNA polymerase
continues transcription, but the sigma
factor is released and can bind to other
RNA polymerases.
© 2011 Nature Education All rights
reserved.
Figure Detail
Bacterial sporulation of Bacillus
subtilis provides a good example of sigma factors that specifically bind specific
promoters. These bacteria exist in both the vegetative state and endospore state. During endospore formation, specific
genes are expressed, but they are not expressed during the vegetative state. The first genes for sporulation are turned on
after the expression of a gene that encodes an alternative sigma factor. Different sigma factors are subsequently
expressed to activate new genes that are needed later in the sporulation. Each of these sigma factors recognizes
promoters of the genes in its group. By using the same core RNA polymerase with different sigma factor subunits, the
organism is able to regulate expression of different sets of genes.
Riboswitches frequently regulate gene expression in bacteria.
Some mRNA transcripts contain riboswitches, or regions of RNA that can
switch conformations depending on the presence or absence of ligands such
as metabolites or metal ions. In bacterial mRNA, riboswitches are commonly
found in the 5′ untranslated region (5′-UTR) preceding the start codon
(Figure 6). Similar to the interaction between an allosteric effector and its
allosteric protein, the binding of a riboswitch to its appropriate ligand
changes the conformation of the mRNA molecule in a variety of ways that
can influence the expression levels of the genes contained in the mRNA.
Figure 6: Bacterial mRNA with a riboswitch.
Riboswitches are typically found in the 5′ untranslated region of an mRNA
transcript. In this type of riboswitch, the conformation of the mRNA
determines whether the remainder of the mRNA is transcribed past the
riboswitch region.
© 2013 Nature Education All rights reserved.
Figure Detail
Because RNA is primarily single stranded, it can fold into secondary and
tertiary structures, much like proteins can. This ability of RNA to adopt threedimensional structures allows mRNA to possess binding sites for other
molecules and switch between conformations depending on whether these
binding sites are occupied. The aptamer of a riboswitch is the region that
binds to a specific ligand, such as enzyme cofactors, nucleotide precursors,
amino acids or metal ions. When the aptamer binds the appropriate ligand, it
induces a conformational change in a nearby region of mRNA known as the
expression platform.
Conformational changes in the expression platform can have a variety of
effects on gene expression. One example involves a mechanism of
transcription termination that depends on the formation of a stem-loop
structure. In some genes, the DNA template contains a special sequence
such that the mRNA transcribed from it folds back on itself and forms a
hairpin loop. The hairpin is created by base pairing of nearby complementary
G and C bases within the mRNA. The region immediately following the
hairpin is rich in U bases in the mRNA — and therefore A bases in the
corresponding DNA template. The formation of the stronger GC base pairs in
the hairpin, combined with the weaker UA base pairs between the mRNA
and the DNA template, separates the mRNA from the DNA and terminates
transcription.
The stem-loop structure can form the basis of an expression platform that
determines whether an mRNA is transcribed to completion (Figure 7). If the
ligand for the riboswitch is absent, the stem-loop structure does not form,
and transcription proceeds normally. However, if the ligand is present and
binds to the aptamer, the conformation of the expression platform changes
such that the stem-loop structure forms and prematurely terminates
transcription. In effect, the presence of the ligand prevents mRNA past the
riboswitch from being transcribed. In other riboswitches, the converse may
be true — the absence of the ligand results in transcription termination, and
the presence of the ligand is required for normal levels of transcription.
Figure 7: The aptamer and expression platform of a riboswitch.
The riboswitch changes between different conformations depending on
the presence or absence of a metabolite ligand (M). In this example, if the
ligand is not bound (left), a hairpin loop forms, but it is too far from the run
of U bases to behave as a transcription termination signal. The remainder
of the mRNA is transcribed in this case. If a ligand binds to the aptamer
(right), the entire riboswitch changes conformation such that the
expression platform is folded into a hairpin loop adjacent to the run of U
bases. This is the stem-loop structure that prematurely aborts transcription
and prevents the expression of the genes in the remainder of the mRNA.
© 2013 Nature Education All rights reserved.
Because bacteria lack a nuclear membrane to separate transcription and
translation into different areas, translation of polypeptides can occur as soon
as mRNA is transcribed. Some types of riboswitches take advantage of the
fact that transcription and translation can occur in the same space in bacteria
to regulate gene expression at the translational level. For example, some
riboswitches can either expose or sequester the ribosome-binding site of the
mRNA in response to ligand molecules binding to their aptamers. In this way,
an mRNA can be transcribed to completion, but its riboswitch's conformation
will decide whether it will actually be translated into protein. Other
riboswitches contain codons that influence the position of a ribosome along
the 5′-UTR as it translates a short "leader" polypeptide. The ability of the
ribosome to pass these codons depends on the availability of specific amino
acids. In turn, this ribosome's position determines whether a stem-loop
structure forms and whether the full mRNA is transcribed and later
translated. In all cases, however, riboswitches are an important way to
regulate gene expression in response to chemical cues from the
environment.
Biomedical research.
The ability of riboswitches to regulate gene expression makes them possible
drug targets in biomedical research. Studies of aptamers have shown that
RNA can distinguish between structures that are closely related and that
they can do this as well as an antibody can. As a result, riboswitches in
pathogenic bacteria and fungi are becoming a new target for drug design. In
2006, Rebecca Montange and Robert Batey at the University of Colorado at
Boulder found the structure of a riboswitch in bacterial mRNA bound to
S-adenosylmethionine. Investigating the complex folded structure of the
riboswitch, they identified the structural changes that result from the ligand
binding and that prevent transcription. New drugs that target the binding sites
that are regulated by the riboswitch could repress transcription, which could
kill the bacteria. As riboswitch research continues, with scientists elucidating
the three-dimensional structures of more and more riboswitches, researchers
will have a greater chance of creating new riboswitch-targeting
antimicrobials.
IN THIS MODULE
Gene Regulation in Bacteria
Summary
Test Your Knowledge
WHY DOES THIS TOPIC MATTER?
Synthetic Biology: Making Life from
Bits and Pieces
Scientists are combining biology and
engineering to change the world.
PRIMARY LITERATURE
Controlling inflammation to stop
sepsis
Amelioration of sepsis by inhibiting
sialidase-mediated disruption of the
CD24-SiglecG interaction.
View | Download
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Principles of Biology
51 Prokaryotic Gene Regulation
Summary
Describe the organization of bacterial DNA into operons.
Some bacterial genes are organized into operons, in which the genes are
under the control of a single promoter and a regulatory region called an
operator. Bacteria exhibit three major types of gene regulation. In repressible
operons, an allosteric effector triggers the repression of an otherwise
expressed operon. In inducible operons, the effector induces the expression
of an operon that is repressed in the absence of the effector. In positive gene
regulation, the effector increases the expression of the operon above some
baseline level.
OBJECTIVE
Explain the function of RNA polymerase.
RNA polymerase initiates transcription at specific sequences called
promoters. Promoter choice by RNA polymerase is regulated by sigma
factors. In many operons, the binding of a repressor protein to the operator
obstructs the RNA polymerase from binding the promoter, which prevents
transcription of the operon genes.
OBJECTIVE
Describe the role of riboswitches in regulating gene
expression.
Riboswitches are mRNA molecules that bind ligands and change
conformation as a result. Some of these alternative structures serve as
signals to continue or terminate transcription or translation. As a result,
riboswitches serve as a way for mRNA molecules to sense the environment
and regulate gene expression.
OBJECTIVE
Key Terms
allosteric effector
A small molecule that binds to the allosteric site of a protein, thereby changing the
protein's conformation and activity.
allosteric protein
A protein whose activity is regulated by the binding of a small regulatory molecule
(an allosteric effector) to the allosteric site of the protein, which results in a
conformational change.
aptamer
The region of a riboswitch that serves as a receptor for a ligand; binding of the
ligand to the aptamer triggers conformational changes in the expression platform
that influence gene expression.
expression platform
The region of a riboswitch that changes conformation in response to a ligand
molecule binding to the aptamer.
inducible operon
A negatively regulated operon whose activity is stimulated (induced) by the
presence of an allosteric effector; expression of the operon is off when the effector
is absent and on when the effector is present.
operator
A specific DNA sequence within an operon that serves as the binding site for a
repressor protein.
operon
A cluster of genes that are expressed together and regulated from a single
promoter; includes the genes themselves, the promoter and the regulatory
sequences, such as the operator.
contents
promoter
A specific DNA sequence where RNA polymerase binds to the DNA to initiate
transcription.
repressible operon
A negatively regulated operon whose activity is inhibited (repressed) by the
presence of an allosteric effector; expression of the operon is on when the effector
is absent and off when the effector is present.
repressor
A regulatory protein that binds to an operator and inhibits RNA polymerase from
binding to the promoter.
riboswitch
A type of regulatory region in RNA that contains a region (the aptamer) that can
bind a ligand and a region that changes conformation as a result of ligand binding
(the expression platform); involved in regulation of gene expression at the
transcriptional and/or translational levels.
stem-loop structure
A sequence of RNA containing internal GC base pairing such that a hairpin loop
forms during its transcription, which serves as a mechanism to terminate
continued transcription.
IN THIS MODULE
Gene Regulation in Bacteria
Summary
Test Your Knowledge
WHY DOES THIS TOPIC MATTER?
Synthetic Biology: Making Life from
Bits and Pieces
Scientists are combining biology and
engineering to change the world.
PRIMARY LITERATURE
Controlling inflammation to stop
sepsis
Amelioration of sepsis by inhibiting
sialidase-mediated disruption of the
CD24-SiglecG interaction.
View | Download
page 262 of 989
1 pages left in this module
Principles of Biology
51 Prokaryotic Gene Regulation
Test Your Knowledge
1. Complete the following sentence: Prokaryotes differ from eukaryotes by...
lacking a nucleus.
lacking proteins.
lacking RNA.
lacking DNA.
lacking ribosomes.
2. How would a mutation in the operator of the lac operon that prevented the LacI
repressor from binding affect the expression of the genes in the lac operon?
The lac operon genes would be expressed continuously.
The lac operon genes would not be expressed at all.
The mutation would repress lactose.
The mutation would inhibit lactose.
The lac operon genes would be expressed in a regulated manner.
3. Complete the following sentence: The lac repressor is an allosteric molecule
because...
it catalyzes the synthesis of lactose.
it lowers the transcription rate.
it inhibits the synthesis of lactose.
it catalyzes mutations on the operon.
it interacts with another molecule to change the repressor's active site.
4. What is the function of the sigma factor in transcription?
to prevent the binding of the polymerase to the promoter
to enable specific binding of DNA polymerase to RNA
to enable specific binding of promoters to operons
to enable specific binding of RNA polymerase to promoters
to enable specific binding of RNA to DNA
5. Which environmental condition results in maximal expression levels of the lac
operon genes?
low glucose, high lactose
high glucose, high lactose
low glucose, low lactose
high glucose, low lactose
All of these conditions will result in the same level of lac operon gene expression.
contents
6. Which mutation is expected to have a similar effect to a mutation in which the LacI
repressor is no longer able to bind to the operator?
a mutation in the operator that enables it to bind but not release the LacI repressor
a mutation in the operator that prevents recognition by the LacI repressor
a mutation in the LacI repressor in which it can still bind the operator but lacks an
allosteric site
a mutation in the promoter that prevents RNA polymerase from binding
None of the answers are correct.
Submit
IN THIS MODULE
Gene Regulation in Bacteria
Summary
Test Your Knowledge
WHY DOES THIS TOPIC MATTER?
Synthetic Biology: Making Life from
Bits and Pieces
Scientists are combining biology and
engineering to change the world.
PRIMARY LITERATURE
Controlling inflammation to stop
sepsis
Amelioration of sepsis by inhibiting
sialidase-mediated disruption of the
CD24-SiglecG interaction.
View | Download
page 263 of 989