Different kinds of cells

C0NTROL OF GENE
EXPRESSION
• Control of gene expression is
essential to all organisms: in bacteria
it allows the cell to take advantage of
changing environmental conditions,
in multicellular organisms it is critical
for directing development and
maintaining homeostasis.
• One way to control gene expression is to regulate initiation of
transcription. In order for a gene to be transcribed, RNA-
polymerase must have access to the DNA-helix and must be
capable of binding to the gene’s promoter, a specific sequence
of nucleotides at one end of the gene that tells the polymerase
where to begin transcribing. How is the initiation of
transcription regulated?
• Protein- binding nucleotide sequences on the DNA
regulate the initiation of transcription by modulating the
ability of RNA-polymerase to bind to the promoter. This
protein- binding sites are usually 10-15 nucleotides in
length. Hundreds of these sequences have been
characterized and each provides a specific binding site
for a specific protein able to recognize the sequence.
Binding of protein to the regulatory sequence either
blocks transcription by getting the way RNA-polymerase,
оr stimulates transcription by facilitating the binding of
RNA-polymerase to the promoter.
 Regulation of gene
expression in Prokaryotes
• The Operon model for the regulation of gene
expression.
• In 1961 F. Jacob and J. Monod at the Pasteur
Institute in Paris proposed that metabolic
pathways are regulated as a unit. They were
awarded Nobel prize in 1965. Based on genetic
studies of the production of the enzymes
involved in lactose metabolism Jacob and
Monod proposed the operon model to explain
gene induction in prokaryotes. Although there
are many operons in bacterial cells, the
lactose (lac.operon) discovered by Jacob and
Monod is the classic example of all operons
 Lactose metabolism in
Escherichia coli.
• In the absence of glucose, E.coli can use lactose, if
present as a source of carbon and energy.
• β-Galactosidase is the key enzyme in the metabolism of
lactose
• by E.coli. It catalyzes the following reaction:
• Lactose + H2 O > Galactose + Glucose
• In the absence of lactose, there are fewer than 10
molecules of β-galactosidase per cell
• In the presence of lactose, and no other energy source,
the number of β-galactosidase molecules can increase to
5000 molecules per cell within several minutes.
• When the sugar lactose is added to the
cultures of E.coli it induces three enzymes
necessary to
• break down the lactose into glucose and
galactose. These enzymes are synthesized
together in a coordinated manner and the
unit is called lac operon.
• Since the addition of lactose itself
stimulates the production the enzymes it is
also known as inducible system.
 Lactose (lac) operon.
• An operon is a group of coordinately
regulated genes, the
• products of which typically catalyze a
multienzyme metabolic
• pathway and their contralling elements.
The purpose of the lac
• operon is to make the enzymes
necessary to metabolize lactose
• (i.e., β-galatozidase, galatoside permease,
thiogalactosidase transacetylase)
 Two classes of
genes are needed to make a functional
operon: structural genes and regulatory
genes.
• a. The products of operons are produced by structural
genes .
• Structural genes code for products that may be
enzyme or
• transfer RNA (t-RNA), ribosomal RNA (rRNA), or
ribosomal protein. Structural gene products are
essential for the life of the cell.
• b. Regulatory genes code for products that regulate
the level
• of expression of structural genes. Although regulatory
genes are
• often not considered part of operons because they can
be located at sites remote from structural genes they
regulate, they are key elements of operons.
 Structure of the lac-
operon.
• The lac operon is a region of DNA in the
genome that contains
• the following.
• Three linked structural genes.
• 1. The lac Z gene that codes for B-
galactosidase
• 2. The lac Y gene that codes for galactoside
permease
• 3. The lac A gene that codes for
thiogalactosidase transacetylase.
Structure of the lac-
operon.
• A single promoter directs proper
initiation of transcription.
• The lac Z, lac Y, and lac A genes are
expressed as a polycistronic message
from this common promoter.
• An operator region lies adjacent to the
promoter and spans the transcriptional
initiation site. A regulatory protein called
the
• lac repressor binds to this site and
blocks initiation of
• transcription.
 Regulation of lac operon
expression
Negative regulation by the lac repressor.
• In the absence of lactose, the cell has no
need for the production of B-galactosidase
and galactoside permease.
• A regulatory molecule, the lac repressor,
prevents expression of the lac operon in the
absence of lactose. This is the example of
negative regulation.
 Structure of the lac-
repressor.
• The lac- repressor is a tetrameric protein,
with each subunit having a binding site for
an inducer. The lac - repressor is a
diffusible product of the regulatory lac I
gene.
• The lac I – gene is adjacent to the lac
operon.
• The lac I gene maintains the very low level
of lac - repressor needed for the regulation
of the lac-operon.
• In the absence of inducer, the lac
repressor binds tightly to the
operator, which blocks initiation of
transcription of the structural
genes.
• The presence of inducer (allolactose) relieves
• the negative regulation by the lac repressor. Upon
binding of the inducer to the lac repressor, the
repressor undergoes a conformational change to
a shape that no longer binds the operator
tightly. With the repressor no longer blocking
the initiation site RNA polymerase initiates
transcription.
• Since prokaryotes do not have a nucleus,
there is no physical structure such as a
nuclear membrane to separate translation,
therefore trancription and translation are
coupled in prokaryotes. Ribosomes bind to
the polycistronic lac messenger RNA (lac-
mRNA) and initiate translation even before
transcription of lac operon is complete.
Structure of the lac-
operon
• As long as lactose is present, then inducer present, and
• transcription of the lac operon continues. The result is
the
• continued production of the enzymes needed for the
metabolism of
• lactose.
• After the inducer is removed, expression of the lac
operon
• stops quickly. This occurs due to the fact that as with
most
• prokaryotic m-RNAs, the lac m-RNA is unstable and
decays within
• minutes.
 The Need for Gene
Regulation in Eukaryotes
• While eukaryotes can respond to their environment , the
main reason higher eukaryotes need to regulate their
genes is cell specialization. Whereas prokaryotes are
simple, unicellular organisms, multicellular eukaryotes
consist of hundreds of different cell types, each
differentiated to serve a different specialized function.
Each cell type differentiates by activating a different
subset of genes. Because of the multitude of cell types,
the regulation of gene expression required to bring about
such differentiation is necessarily complex. One way this
complexity is demonstrated is in multiple levels of
regulation of gene expression.
 Levels of Regulation
• Eukaryotic cells are more complex than
prokaryotic cells.
• Nucleus in eukaryotic cells separates
transcription from translation in a way not
seen in prokaryotes.
• Eukaryotic transcripts must be processed
before they can be translated. Here is a
diagram outlining the steps involved in the
production of a protein in eukaryotic cells:
Diagram outlining the steps involved in the
production of a protein in eukaryotic cells:
• Regulation can occur at any point in
this pathway; specifically, it occurs at
the levels of transcription, RNA
processing, mRNA lifetime
(longevity), and translation. Each of
these types of regulation will be
considered in turn.
 Regulation of RNA
Processing
• After transcription, the RNA must be processed
before it can be translated. RNA processing
involves addition of a 5' cap, addition of a 3' poly
(A) tail, and removal of introns. This processing
represents another level of regulation of gene
expression, particularly in regard to splicing out of
introns. Regulation can be of two types: a)
whether an RNA gets processed; and b) which
exons are retained in the mRNA.
• The first type of regulation can determine whether or
not an mRNA gets translated. If an RNA is not
processed, it will not be transported out of the nucleus,
and will not be translated.
• The second type of regulation can affect the function of
the protein produced. Some genes have exons that can
be exchanged in a process known as exon shuffling.
For example, a gene with four exons might be spliced
differently in two different cell types. In cell 1, exons 1,
2, and 4 would be used in the mRNA:
• . In cell 1, exons 1, 2, and 4 would be used
in the mRNA:

• In cell 2 on the other hand, exons 1, 3, and

4 would be used:
In cell 2 on the other hand, exons 1, 3,
and 4 would be used:
• In each of these cases, the polypeptide produced
could have a different function. In mammals, for
example, the calcitonin gene produces a
hormone in one cell type, and a neurotransmitter
in another cell type, due to exon shuffling. In
Drosophila, alternate splicing of the sex-lethal
RNA can produce an mRNA encoding a functional
polypeptide, or one with a premature stop codon
that encodes a short, nonfunctional polypeptide.
 Regulation of RNA
Longevity
• Imagine two mRNA molecules: one lasts for five minutes in the
cytoplasm before being degraded, while the other one manages
to linger for an hour before being degraded. If both are
translated continually while they exist, it is obvious that more of
the second polypeptide will be produced than the first. This is
the principle behind regulation of RNA longevity. mRNAs from
different genes have their approximate lifespan encoded in
them; this serves to help regulate how much of each
polypeptide is produced. The information for lifespan is found in
the 3' UTR. The sequence AUUUA, when found in the 3' UTR, is
a signal for early degradation (and therefore short lifetime). The
more times the sequence is present, the shorter the lifespan of
the mRNA. Because it is encoded in the nucleotide sequence,
this is a set property of each different mRNA; the longevity of
an mRNA can't be varied.
Regulation of Translation
• Whether or not an mRNA molecule is translated can be
regulated as well. The various mechanisms of
translational regulation are incompletely understood,
but there are many documented examples
(particularly in embryonic development) of mRNA
molecules that are present routinely, but are only
translated under certain circumstances. For example,
many animals sequester large amounts of mRNA in
their eggs, and those mRNA molecules are not
translated unless the egg is fertilized.
Regulation of Transcription
• Many different proteins participate in transcription.
Some of these are required for the transcription of all
structural genes and are termed general transcription
factors. Others, labeled specific transcription factors,
have more specialized roles, activating only certain
genes at certain stages of development A key
transcriptional element is RNA-polymerase. It plays a
vital role in initiating transcription by binding to the
promoter region, but it can not locate the promoter
region on its own. Further more, it is incapable of
producing significant quantities of m-RNA by itself.
• Effective transcription requires the
interaction of a large complex of 50
or so different proteins. These
include different transcription
factors, which bind to RNA-polymrase
and to specific DNA sequences in the
promoter region ( sequences such as
TATA and others needed for
transcription initiation).
• The general transcription factors allow RNA-
polymerase to bind to the promoter region so
that it can function effectively in transcription.
The transcriptional activity of specific genes
can be greatly increased by interaction with
sequences called enhancers, which may be
located thousands of bases upstream or
downstream of the gene A very large number
of enhancer elements has been identified and
characterized, and each different enhancer has
its own transcription factor that it binds to.
• Enhancers however, do not interact
directly with genes. Instead, they are
bound by specific transcription factors,
co-activators, which in turn bind to
general transcription complex. This
chain of interactions from enhancer to
activator, to coactivator, to general
transcription complex and finally to the
gene itself, increases the activity of
specific genes at specific points in
time.
• Transcription factors can be activated by signals from
other cells in the same organism. Such signals include
hormones and growth factors. Hormones must bind to
a specific receptor on the target cell, and the receptor
mediates the cellular effects of the signal. There are two
basic mechanisms used, one for steroid hormones, and
one for peptide hormones:
– Steroid hormones are lipid (actually cholesterol) derivatives, such
as testosterone and progesterone. These hormones can cross the
cytoplasmic membrane into a cell, where they bind to their
specific receptor. Steroid receptors are transcription factors, and
when they bind to their ligand, they become activated and
initiate transcription of a specific set of genes.
– Peptide hormones cannot cross the cytoplasmic membrane, and
so must bind to a receptor on the cell surface. When bound to its
ligand, these receptors initiate a series of biochemical reactions
inside the cell, with the ultimate result being the activation of a
transcription factor (often by phosphorylation of the transcription
factor), which initiates transcription of a specific set of genes.
• Transcription factors can be activated by
environmental signals. For example, virtually all
organisms have a set of genes called heat
shock genes that encode proteins that help the
organism survive heat stress. These genes are
activated under conditions of heat stress, under
the control of a transcription factor called heat
shock transcription factor. This factor is
always present, but is only activated when
greatly increased temperatures are detected.
• Whereas enhancers help to increase
the transcriptional activity of genes,
other DNA sequences, known as
silencers help to repress the
transcription of genes through a
similar series of interactions.