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Regulation of Gene Expression ( Prokaryotes and Eukaryotes)
Introduction
All the activities of an organism are controlled by genes. Most of the genes of an organism
express themselves by producing proteins. The genes which produce proteins are called
structural genes or cistrons. Every cell of an organism posses all the genes. But all of them
are not functional all the time. If all the genes function all the time, enzymatic chaos will
prevail and there will not be much cell differentiation. The products of many genes are
needed only occasionally by the cell. Therefore, those proteins are synthesized only when the
substrate on which they act is present or when they are needed by the cell. In highly
differentiated cells of eukaryotes only a few genes are functional and all other genes are
permanently shut off. Even a lowly E. coli bacterium expresses only some of its genes at any
given time out of the total of about three thousand genes. Various mechanisms exist in the
cell, which control and regulate the expression of genes. The regulatory system turns the
genes “on” when needed and turns “off’ when not needed. This proves that gene activity can
be regulated. There are various stages at which the expression of a gene can be regulated but
most common is the initiation of transcription. It is here that bulk of the gene regulation takes
place. Other levels of gene regulation are transcriptional elongation, mRNA processing
during translation and post translation stage.
Gene regulation in prokaryotes
In bacteria the expression of genes is controlled by extracellular signals often present in the
medium in which bacteria are grown. These signals are carried to the genes by regulatory
proteins. Regulatory proteins are of two types. They are positive regulators called activators
and negative regulators called repressors. These activators and repressors are DNA binding
proteins.
Negative Regulators or Repressors
The repressor or inhibitor protein binds to the target site (operator) on DNA. These block the
RNA polymerase enzyme from binding to the promoter, thus preventing the transcription.
The repressor binds to the site where it overlaps the polymerase enzyme. Thus, activity of the
genes is turned off. It is called negative control mechanism. An anti-repressor or antiinhibitor called inducer is needed to inactivate the repressor and thereby activating the genes.
Thus, the genes are switched on. This is demonstrated by lactose operon.
Operon
In bacteria cistrons or structural genes, producing enzymes of a metabolic pathway are
organised in a cluster whose functions are related. Polycistronic genes of prokaryotes along
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with their regulatory genes constitute a system called operon. Operon is a unit of expression
and regulation.
Lactose Operon or Lac Operon
This is a negative control mechanism. In 1961 Francois Jacob and Jacques Monod proposed
operon model for the regulation of gene expression in E. coli. The synthesis of enzyme (3galactosidase has been studied in detail. This enzyme causes the breakdown of lactose into
glucose and galactose. In the absence of lactose, β-galactosidase is present in negligible
amounts. As soon as lactose is added from outside, the production of β- galactosidase
increases thousand times. As soon as the lactose in consumed, the production of the enzyme
again drops. The enzymes whose production can be increased by the presence of the substrate
on which it acts are called inducible enzymes.
Addition of lactose to the culture medium of E. coli induces the formation of three enzymes
(5-galactosidase, permease and transacetylase, which degrade lactose into glucose and
galactose. The genes, which code for these enzymes lie in a cluster and are called cistrons or
structural genes. They are transcribed simultaneously into a single mRNA chain, which has
codons for all the three enzymes. The mRNA transcribed from many genes is called
polycistronic. The functioning of structural genes to produce mRNA is controlled by
regulatory genes.
There are three structural genes Z, Y and A, which code for enzymes p-galactosidase, lac
permease and transacetylase respectively. Regulatory genes consist of Regulator I, Promoter
P and a control gene called operator gene O. Regulator I gene produces a protein called
repressor or inhibitor. The repressor is active and binds to the operator gene O and switches it
“off” and the transcription is stopped. This happens because RNA polymerase enzyme which
binds to the promoter is unable to do so because binding site of RNA polymerase and the
binding site of repressor on operator overlap each other. Hence in negative control
mechanism, the active genes are turned “off” by the repressor protein.
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When the inducer (lactose) in supplied from outside, the inducers binds to the repressor. The
lactose on entering the bacteria changes into allolactase. Allolactose changes the shape of the
repressor (conformational changes) which renders it inactive and unable to bind to the
operator. The operator becomes free and is “turned on” and thus transcription starts. In this
way, the presence of the inducer permits the transcription of Lac operon, which is no longer
blocked by the repressor protein. The synthesis of enzymes in response to the presence of
specific substrate (lactose) is called induction. It is also called de-repression.
The inducible system operates in a catabolic pathway. In the absence of lactose, E. coli cells
have an average of only three molecules of P-galactosidase enzyme per cell. Within 2-3
minutes of induction of lactose, 3000 molecules of P-galactosidase are produced in each cell.
Tryptophan Operon
It is also a negative control system but forms a biosynthetic pathway. It is known as
repressible system. It works on the principle that when the amino acid tryptophan is present,
there is no need to activate the tryptophan operon.
Repressor protein is activated by the co-repressor (tryptophan-the end product) and it binds
the operator to switch it “off’. Tryptophan is synthesized in five steps, each step requiring a
particular enzyme. The genes for encoding these enzymes lie adjacent to one another, called
trp E, trp D, trp C, trp B and trp A.
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Tryptophan operon codes for five enzymes that are required for the synthesis of amino acid
tryptophan. In repressible system, the regulatory gene produces a repressor protein, which is
normally inactive and unable to bind to operator on DNA. The repressor upon joining the corepressor (which is the end product tryptophan in this case) undergoes conformational
changes that activate it and enable it to bind to the operator. This prevents the binding of
RNA polymerase enzyme to the promoter. This is opposite to the situation of lac operon in
which the repressor is active on its own and loses the affinity for the operator when bound to
the inducer.
Here the availability of tryptophan which is the end product regulates the expression of this
operon and represses the synthesis of tryptophan. In this way the synthesis of enzymes of a
metabolic pathway is stopped by the end product of the metabolic chain. This mechanism
enables the bacteria to synthesize enzymes only when they are required. This is known as
feed back repression. In feed back inhibition the end product of a metabolic pathway acts as
an allosteric inhibitor of the first enzyme of the metabolic chain. Induction and repression
save valuable energy by preventing the synthesis of unnecessary enzymes.
Positive Control of Transcription
The system of regulation in lactose and tryptophan operon is essentially a negative control in
the sense that the operon is normally “on” but is kept “off’ by the regulator protein. In other
words the structural genes are not allowed to express unless required.
Catabolic Repression
Lac operon also shows positive control by catabolic repression. This is an additional control
system, which binds the repressor-operator. In E. coli, in the presence of both glucose and
lactose, the glucose in first fully utilized and then lactose is taken up for production of
energy. Glucose is richest and more efficient source of energy. Glucose has an inhibitory
effect on the expression of lac operon. The mechanism of positive control enables E. coli to
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adapt more efficiently to the changing environment of its natural habitat, which is the human
intestine.
In the presence of glucose, synthesis of β-galactosidase enzyme becomes suppressed. The
inhibitory effect of glucose is due to the marked drop in the level of a nucleotide called cyclic
AMP (c-AMP), which inhibits the transcription of mRNA. Lactose operon transcription
requires not only cyclic AMP but also another protein called catabolic activator protein
(CAP). The cAMP and CAP form a complex called cAMP-CRP complex, which is necessary
for the functioning of lactose operon. A catabolic breakdown product of glucose, called glucose catabolite, prevents the activation of lac operon by lactose. This effect is called
catabolic repression. When glucose concentration increases, the cAMP concentration decreases and vice versa. High concentration of cAMP is necessary for the activation of lac
operon. Normally in the presence of glucose, the lactose operon remains inactive. Glucose
catabolite prevents the formation cAMP-CRP complex.
In this way cAMP-CRP system is positive control because expression of lac operon requires
the presence of an activating signal which is this case in cAMP-CRP complex. There are
some promoters on DNA at which RNA polymerase cannot initiate transcription without the
presence of some additional protein factors such as cAMP-CRP complex. These factors are
positive regulators because their presence is necessary to switch on the cistrons. These are
called activators or stimulators.
Gene regulation in Eukaryotes
Gene regulation refers to the control of the rate or manner in which a gene is expressed. In
other words, gene regulation is the process by which the cell determines [through interactions
among DNA, RNA, proteins, and other substances] when and where genes will be activated
and how much gene product will be produced. Thus, the gene expression is controlled by a
complex of numerous regulatory genes and regulatory proteins. The gene regulation has been
studied in both prokaryotes and eukaryotes.
Eukaryotic gene expression can be regulated at many stages
In the articles that follow, we’ll examine different forms of eukaryotic gene regulation. That
is, we'll see how the expression of genes in eukaryotes (like us!) can be controlled at various
stages, from the availability of DNA to the production of mRNAs to the translation and
processing of proteins. Eukaryotic gene expression involves many steps, and almost all of
them can be regulated. Different genes are regulated at different points, and it’s not
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uncommon for a gene (particularly an important or powerful one) to be regulated at multiple
steps.
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Chromatin accessibility. The structure of chromatin (DNA and its organizing proteins) can
be regulated. More open or “relaxed” chromatin makes a gene more available for
transcription.
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Transcription. Transcription is a key regulatory point for many genes. Sets of transcription
factor proteins bind to specific DNA sequences in or near a gene and promote or repress its
transcription into an RNA.
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RNA processing. Splicing, capping, and addition of a poly-A tail to an RNA molecule can be
regulated, and so can exit from the nucleus. Different mRNAs may be made from the same
pre-mRNA by alternative splicing.

RNA stability. The lifetime of an mRNA molecule in the cytosol affects how many proteins
can be made from it. Small regulatory RNAs called miRNAs can bind to target mRNAs and
cause them to be chopped up.
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Translation. Translation of an mRNA may be increased or inhibited by regulators. For
instance, miRNAs sometimes block translation of their target mRNAs (rather than causing
them to be chopped up).
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
Protein activity. Proteins can undergo a variety of modifications, such as being chopped up
or tagged with chemical groups. These modifications can be regulated and may affect the
activity or behavior of the protein.
Although all stages of gene expression can be regulated, the main control point for many
genes is transcription. Later stages of regulation often refine the gene expression patterns that
are "roughed out" during transcription.
Regulation of gene expression involves many different mechanisms
In prokaryotes, regulatory mechanisms are generally simpler than those found in eukaryotes.
Prokaryotic regulation is often dependent on the type and quantity of nutrients that surround
the cell as well as a few other environmental factors, such as temperature and pH. A
combination of activators, repressors and occasionally enhancers control transcription.
Prokaryotic gene expression also happens in the same space as translation, reducing the
opportunities for compartmentalization of regulation. Multicellular organisms have more
complex genomes and the presence of a nucleus and separate cytoplasm provide a more
compartmentalized structure. There are a number of different stages at which gene expression
may be regulated in eukaryotes (figure 1)
Figure 1: Regulation of gene expression in eukaryotes may take place at several different
stages.
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In the nucleus, the process of chromatin remodeling regulates the availability of a gene for
transcription. Once transcribed, the primary transcript of mRNA, or pre-mRNA, undergoes
RNA processing, which involves splicing and the addition of a 5′ cap and a 3′ poly(A) tail to
produce a mature mRNA in the nucleus. The mature mRNA is then exported from the
nucleus to the cytoplasm, where its life span varies. Once outside the nucleus, localization
factors may target mature mRNAs to specific regions of the cytoplasm where they are
translated into polypeptides. The resulting polypeptides can undergo post-translational
modifications, which can regulate protein folding, glycosylation, intracellular transport,
protein activation, and protein degradation.
Several gene expression mechanisms involve chromatin structure
When eukaryotic cells aren't dividing, chromosomes exist in an uncondensed state called
chromatin. Chromatin consists of DNA wrapped around a histone protein core. The wrapped
DNA isn't as available for transcription as the DNA of prokaryotes, and as we'll discuss,
mechanisms exist to relieve this repression. Also in eukaryotes, the RNA polymerase doesn't
bind directly to the DNA, but instead binds via a set of proteins: the transcription initiation
complex. Two different types of chromatin can be seen during interphase: euchromatin and
heterochromatin. Euchromatin, which is a lightly packed form, contains areas of DNA that
are undergoing active gene transcription. Not all of the chromatin is undergoing gene
transcription, however. Heterochromatin, in contrast, is mostly inactive DNA that is being
actively inhibited or repressed in a region-specific manner. The chromatin state can change in
response to cellular signals and gene activity. This is facilitated by enzymes that modify
histones by adding methyl and acetyl groups to their N-terminal tails. Acetylation reduces the
net positive charge of the histones, loosening their affinity for DNA, and increasing
transcription factor binding. Methylation, in contrast, leads to increased binding of histones to
DNA, and decreases the availability of DNA for transcription. Figure 2 shows an example of
how acetylation and methylation of histones may affect transcriptional activity in a normal
cell compared to a cancer cell with inappropriate gene expression.
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Figure 2: Modifications of methyl and acetyl groups in histones affect transcriptional activity.
The grey cylinders represent histone octamers. Acetylation (blue circles) and methylation
(green circles) of histone subunits are shown. In normal cells, the promoters of tumorsuppressor genes show acetylation of histone subunits, associated with active transcription. In
contrast, in cancer cells, the promoters of tumor-suppressor genes are not acetylated, and the
genes are not actively transcribed. In normal cells, the heterochromatic regions at the ends of
the chromosomes do not show acetylation, and the genes are not actively transcribed. In
cancer cells, the heterochromatic regions at chromosome ends are acetylated and
transcriptionally active.
Transcription is an important stage for gene regulation
In eukaryotes, the RNA immediately transcribed from a DNA template, the pre-mRNA,
undergoes a number of processing events before it becomes a mature mRNA (Figure 3). The
RNA polymerase needs transcription factors to initiate transcription. Some general
transcription factors are required for all genes. Some bind to specific sequences of DNA such
as the TATA box. Others bind to proteins such as RNA polymerase II. These general
transcription factors don't usually produce high rates of transcription, and for that reason,
gene-specific transcription factors called activators or repressors are also required. These
factors bind to proximal or distal control elements, which are specific DNA sequences that
are usually four to eight base pairs long. The rate of gene expression may be greatly affected
by binding of specific transcription factors to control elements. Proximal control elements are
close to the promoter. Distal control elements may be grouped as enhancers, and may be
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thousands of nucleotides removed from the gene. Although one gene may have more than
one enhancer, a given enhancer is usually associated with only one gene.
Figure 3: The structure of a eukaryotic gene and its transcript.
Each gene has a promoter, the DNA sequence where RNA polymerase, along with
transcription factors, binds and begins transcription. RNA processing removes introns and
splices the exons together using structures called spliceosomes, and a 5′ cap and poly(A) 3′
tail are added to the mRNA transcript. The mRNA is then translated into a polypeptide.