Chapter 13

Regulation of Gene
Expression
The biochemistry and molecular
biology department of CMU

Section 1
Principles and Concepts

1.1 Concepts
Gene: A DNA segment that contains
the all genetic information required to
encodes RNA and protein molecules.
Genome: A complete set of genes of a
given species.
Gene expression: A process of gene
transcription and translation.

Specificity of gene expression

Temporal specificity (also called
stage specificity): why in the infant
not in the aged ones?
Spatial specificity (also called tissue
specificity): why in liver not in brain?

Type of gene expression

a. Constitutive expression
Some genes are essential and
necessary for life, and therefore are
continuously expressed, such as
those enzymes involved in TAC.
These genes are called housekeeping
genes.

b. Induction and repression

The expression levels of some genes
fluctuate in response to the external
signals.

Some genes demonstrate higher

expression level once being activated.
It is called induced expression.
On the other hand, some genes are
repressed and their expression levels
are lower. It is called repressed
expression.

1.2 Regulatory Elements

Gene expression is a multiple-level
process.
Transcription initiation is a key point
of controlling gene expression.

Basic elements that regulate the

transcription include:
a. Special DNA sequences
b. Regulatory proteins
c. DNA-protein interaction and
protein-protein interaction
d. RNA polymerase

a. Special DNA sequence

For prokaryotic systems:
Operon is composed of structural
genes, promoter, operator, and other
regulatory sequences.
Promoter
Other requlatory
sequence

Operator

Sturctural genes

Promoter
The DNA sequence that RNA-pol can
bind to and initiate the transcription.

Operator
The DNA sequence adjacent to the
structural genes that the repressor
protein can bind to and prevent the
transcription of structural genes.

promoter
RNA
pol

operator
repressor

structural gene

For eukaryotic systems:

Cis-acting elements is the special DNA
sequence that can affect the expression
of its own gene.
start
B

structural gene

b. Regulatory proteins
For prokaryotic systems:
Specific factor: It facilitates the
binding of RNA-pol to particular DNA
sequence.
Repressor: It binds to the operator
and prevent the transcription, known
as negative regulation.

 Activator: It associates with DNA

near the initiation point, resulting in
the increase of RNA-pol binding
affinity and the enhancement of the
transcription efficiency.

For eukaryotic systems:

The regulatory proteins are called
transcription factors (TF).
After expression, TF will interact with
the cis-acting elements to activate
another genes. Therefore, they are
referred to as trans-acting factors.

c. DNA-protein interactions
The regulation is implemented
through numerous interactions
between cis-acting elements and
trans-acting factors.
They are non-covalent bond.

Protein-protein interactions
Proteins may have to interact with
each other prior to the DNA binding.
Proteins can form a homo or heterodimer form to function properly.
Present in prokaryotes as well as
eukaryotes.

Section 2
Gene Regulation of
Prokaryotic Systems

Common features
Prokaryotic genes are polycistron
systems, that is, several relevant
genes are organized together to form
a transcription unit --- operon.
The majority of gene regulation is
negative. Inducers are used to
remove the repression.

2.1 Regulation of Transcription

Operon is a coordinate unit for the
regulation.
Transcription initiation is the key
point for regulation. Translation can
also be regulated.

Structure of lac operon

regulatory site
I

structural gene
Z

O
operator

promoter

CAP-binding site
regulatory gene

transacetylase
permease

-galactosidase

Metabolism of
lactose

Inducible expression
Bacteria do not express these three
enzymes when glucose is available.
However, bacteria produce those
enzymes if lactose is present and
glucose is absent.

Sequence of lac operon

lac operon (TTTACA/TATGTT) is a weak

promoter, and has a basal expression
level.
CAP (Catabolite gene activator protein)
binding site is at -60 region.
CAP is a homodimer with binding ability
to DNA and cAMP.

 Glucose inhibits the formation of

cAMP.
When glucose is present, [cAMP] is
lower. Only after glucose is
exhausted, [cAMP] becomes higher.
The CAP-cAMP complex is formed,
and this complex binds to the CAP
binding site on lac operon.

Situation 1
When lactose is absent, no lac gene is
expressed.
I

mRNA

RNA
P pol

 lacI gene has its own promoter, and its

expression can produce LacI
repressor.
The tetrameric Lac repressor binds to
the lac operator site Olac.
The binding blocks the RNA-pol
moving on DNA template, and no lacZ,
lacY, and lacA are expressed.

Situation 2
When lactose is present, lacZ, lacY,
and lacA genes are expressed.
I

RNA
pol
A
Y

mRNA

repressor

galactosidase

allolactose

lactose

 The galactosidase is weakly expressed

(at the basal level).
When lactose is present, it is
converted to allolactose or galactose
that binds to the repressor.
The repressor can no longer bind to
the operator, and lac gene can be
expressed.
Galactose , Allolactose and IPTG are
referred to as inducer.

Inducers

Situation 3
When glucose is present, the [cAMP] is
low, no CAP-cAMP is formed and the
expression of the lac operon is still low.
CAP

RNA
P pol

Situation 4
When glucose is absent and lactose is
present, the CAP-cAMP complex binds
to the CAP site to activate the lac gene.
cAMP

CAP

RNA
P pol

Coordinate expression
no lactose

lactose

RNA pol O

No glucose
RNA pol

Glucose
RNA pol O

RNA pol

2.2 Transcription Attenuation

Trp operon
The trp operon is one of the
constitutive genes expressed at the
basal level.
The structural gene of trp operon
encodes 5 enzymes used for the
synthesis of Trp.

 The trp repressor gene can be

expressed, but it does not bind to the
operator.
When Trp is more than enough, the
repressor will form a complex with
Trp. The complex binds to the
operator, blocking the synthesis of
Trp.

Trp operon

structural
gene

regulatory
regions

trp L

attenuator

Trp mRNA
attenuated mRNA

Trp

Attenuation mechanism
In addition to the repressor regulation,
trp gene has a fine tuning mechanism
called attenuation.
The trp operon is regulated using
attenuation mechanism at the
translation level.

Leader sequence

Possible hairpins

1/2 and 3/4

hairpin
structure

2/3 hairpin
structure

High Trp concentration

Low Trp concentration

2.3 Protein Degradation

Under the normal conditions, the LexA
gene expressed to repressor proteins
that bind to promoters of other genes
and block their expressions.
Once the repressors are degraded, the
repressed genes will be expressed.
At the basal level, the normal cell
contains about 1000 copies of RecA
protein.

SOS response

LexA digestion

 When DNA is extensively damaged,

DNA replication is halted and the
number of ssDNA gaps increases.
The RecA protein binds to this
damaged ssDNA, which activates the
proteins coprotease activity.
While bound to ssDNA, the RecA
protein facilitates the cleavage of
LexA repressor as well as the
inactivation of the LexA repressor.

Section 3
Regulation of
Eukaryotic Transcription

Structural features
Large genome: 3 x 109 bps, 35 000
genes
Monocistron
Repeated sequences: different
lengths and different frequencies.
Often inverted repeats
Splite genes: separated by introns
and exons alternatively

Regulation features
1. RNA-pol: 3 forms (I, II, and III) for
different RNAs
2. Changes of chromosomal structure
Hypersensitive site
Base modification
Isomer-conversion
Histone changes

3. Positive regulation
4. Transcription and translation are
separated
5. Post-transcriptional modification
6. Regulation through intercellular and
intracellular signals

3.1 Cis-acting elements

They are specific DNA sequences,
each of which regulates transcription
of one or more genes. They usually
have consensus sequences.
Promoter: TATA box, CAAT box, and
GC box,

Promoter of eukaryote
cis-acting element
structural gene
GCGC

CAAT

TATA

exon

intron

exon

start
TATA box

enhancer

CAAT box

(Hogness box)

-25 region

GC box

The promoter consensus sequence of eukaryote is TATA which

locate on the -25 region, also called Hogness box or TATA box.
There are other promoters upstream the transcriptive start site,
such as Inr box(YYCARR) CAAT box and GC box .

TATA box
Sequence: TATAAAA
Location:

- 25 ~ - 30 bp

Function: It is the binding site for

TFII D, which is required for RNA
polymerase binding. It controls the
veracity and frequency of
transcriptional initiation.

CAAT box
Sequence: GCCAAT
Location:

~ -70 bp

Function: It is the binding site for

CTF1 (CAAT-binding transcription
factor) and C/EBP (enhancer binding
protein).

GC box
Sequence: GGGCGG
Location:

-30 ~ -110 bp

Function: It is the binding site for

a protein called Sp1.

enhancer
It is a DNA sequence that can
determine the temporal and spatial
specificities of expression and
increase the promoter activity.

Silencer
It is a negative regulation element.
It will repress the transcription once
interacted with specific proteins.

3.2. Trans-acting factors

They are the proteins that bind
indirectly to cis-acting elements and
then regulate the transcription
initiation.
The trans-acting factors can be
transcription factors (TF).

transcription factors
General transcription factors
Special transcription factors
Transcription activators
EBP (enhancer binding protein)
Transcription inhibitors

General structure of TF
DNA-binding domain
Activation domain
Protein-protein interaction domain

Gene activation from distance

Fig. 8-25

3.3 DNA-protein interactions

Regulatory proteins have discrete
DNA-binding domains of particular
structure, i.e., binding motif.
The AA side chains of regulatory
proteins interact with bases of DNA
through H bonds.

Leucine zipper
Yeast activator
protein GCN4

Zinc finger

Steroid hormone receptor

Zinc finger
Mouse
regulatory
protein Zif268

Helix-loop-helix

Helix-loop-helix
Human
transcription
factor MAX

Helix-turn-helix
Lac repressor

Helix-turn-helix

Trp repressor