1 The trp Operon Contains 5 genes coding for proteins enzymes required for the synthesis of the amino acid tryptophan Also contains a promoter and operator When tryptophan levels in the cell are low the expression of the ID: 910714
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Slide1
Regulation of gene Expression in Prokaryotes & Eukaryotes
1
Slide2The trp OperonContains 5 genes coding for proteins (enzymes) required for the synthesis of the amino acid tryptophan. Also contains a promoter and operator.
When tryptophan levels in the cell are low, the expression of the trp
operon
genes is relatively high.There are trp repressor proteins in the cell, but they are unable to bind to the trp operon operator all on their own.
Slide3The trp Operon
When tryptophan levels in the cell are high:Tryptophan binds to the repressor and activates it.
The activated
trp
repressor can now bind to the trp operator and block transcription of the trp operon genes.
Slide4The lac and trp Operons
Slide5Positive Control of TranscriptionOccurs when a regulatory protein (an activator) binds to DNA and increases the rate of transcription of downstream genes.
Let’s look at two examples of activator proteins at work:
AraC
proteins can increase transcription of the
ara operon genes.CAP proteins can increase transcription of the lac
operon
genes.
Slide6Positive Control of the ara OperonE. coli can also utilize arabinose
, a pentose found in plant cell walls, as an energy source.The ara operon
includes:
3 genes required for
arabinose metabolism.A promoter to which RNA polymerase can bind.An initiator sequence to which an activator protein (AraC) can bind to stimulate transcription.
Slide7Positive Control of the ara OperonExpression of the ara
operon genes is high only when arabinose levels are
high
:
(If you were E. coli, you wouldn’t want to waste precious resources expressing these genes unless there was arabinose around to break down!)How does the presence of arabinose stimulate expression of the
ara
operon
genes?
Slide8Positive Control of the ara Operon
If arabinose levels are relatively high…Arabinose binds to
AraC
proteins in the cell…
….which allows the AraC proteins to bind to the initiator region of the ara operon….….which helps RNA polymerase to bind to the promoter region of the ara operon
successfully…
…which increases transcription rate of
ara
operon
genes.
Slide9Positive Control of the ara Operon
Thus, AraC proteins, when bound to arabinose, can act as
activators
of gene expression at the
ara operon:Example of positive control of gene expression.Let’s look at one more example of positive control of gene expression, this time going back to the lac
operon
.
Gene expression at the
lac
operon
is under both:
Negative control (by the
lacI
repressor protein)
Positive control (by a protein called CAP)
Slide10GENE REGULATION IN EUKARYOTES
10
Slide11Eukaryotic cells have a much larger genomeEukaryotes have much greater cell specializationThus eukaryotic cells contain an enormous amount of DNA that does not program the synthesis of RNA or proteinThis requires complex organizationIn eukaryotes expression of gene into proteins can be controlled at various locations
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Slide12Control of gene Expression
Mammalian cells possess about 1000 times more genetic information than does the bacterium
Escherichia coli.
Much of this additional genetic information is probably involved in regulation of gene expression during the differentiation of tissues and biologic processes in the
multicellular organism and in ensuring that the organism can respond to complex environmental challenges.
12
Slide13Synthesis of proteins is controlled right from the chromatin stage.
Expression of gene is controlled at many steps during the process of transcription and translation
.
SEVERAL STEPS OF EUKARYOTIC GENE REGULATION
1
. Transcriptional control.
2. RNA processing control.
3
. RNA transport &
localisation
control.
4
. Translation
control.
5.
mRNA degradation
control.
6. Protein activator control.
Check POINTS FOR GENE Expression in EUKARYOTES
Slide141.Chromatin Structure
Two forms of chromatin
Euchromatin
– A
lesser
coiled, less
methylation
transcriptionally active region which can be easily accessed by the RNA
polymerases.
Heterochromatin
–
Large regions
highly
condensed
methylated
transcriptionally inactive region. The genes in this region cannot be accessed by the RNA polymerases for active transcription.
14
Slide15Heterochromatin change can switchgene on or off
Slide1616
Slide17Mechanism of regulation of gene expression- Details
1) Chromatin Remodeling
Chromatin structure provides an important level of control of gene transcription.
With few exceptions, each cell contains the same complement of genes (
antibody-producing cells are a notable exception).
The development of specialized organs, tissues, and cells and their function in the intact organism depend upon the differential expression of genes.
Some of this
differential expression
is achieved by having different regions of chromatin available for transcription in cells from various tissues.
17
Slide181) Chromatin Remodeling
Large regions of chromatin are
transcriptionally
inactive in some cells while they are either active or potentially active in other specialized cells
For example, the DNA containing the -globin gene cluster is in "active" chromatin in the reticulocytes but in "inactive" chromatin
in muscle cells.
18
Slide19Formation and disruption of nucleosome structure
The presence of
nucleosomes
and of complexes of
histones and DNA provide a barrier against the ready association of transcription factors with specific DNA regions. The disruption of nucleosome structure is therefore an important part of eukaryotic gene regulation and the processes involved are as follows :
19
Slide20Formation and disruption of nucleosome structure (contd.)
A)
Histone
acetylation and deacetylationAcetylation is known to occur on lysine residues in the amino terminal tails of histone
molecules.
This modification
reduces the positive charge
of these tails and decreases the binding affinity of
histone
for the negatively charged DNA.
Accordingly, the
acetylation
of
histones
could result in disruption of
nucleosomal
structure and allow readier access of transcription factors of the same regulatory DNA elements.
20
Slide21A) Histone Acetylation and deacetylation
The amino-terminal tail of
histone
H3 extends into a pocket in which a lysine side chain can accept an acetyl group from acetyl
CoA bound in an adjacent site.
21
Slide22Histone AcetylationAcetylation of the lysine makes RNA polymerase and transcription factors easier to access the promoter
region. Therefore, in most cases, histone acetylation enhances transcription while histone deacetylation represses transcription.
Histone acetylation is catalyzed by
histone
acetyltransferases (HATs) and histone deacetylation is catalyzed by histone deacetylases (denoted by HDs or HDACs
).
Slide23Acetylation :
Methylation:
by HDACs and corepressors leads to heterochromatin formation
by HATs and coactivators leads to
euchromatin
formation
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Slide24Modifications amino acids ofhistones
Acetylation
Methylation
Slide25Methylation of deoxycytidine residues in DNA may effect gross changes in chromatin so as to prevent
its active transcription. Acute demethylation of
deoxycytidine
residues in a specific region of the tyrosine
aminotransferase gene—in response to glucocorticoid hormones—has been associated with an increased rate of transcription of the gene.25
B) Modification of DNA
Slide26C
) DNA Binding Proteins
The binding of specific transcription factors to certain DNA elements may result in disruption of nucleosomal structure.
Many eukaryotic genes have multiple protein-binding DNA elements.
The serial binding of transcription factors to these elements may either directly disrupt the structure of the
nucleosome
or prevent its re-formation.
These reactions result in chromatin-level structural changes that in the end increase DNA accessibility to other factors and the transcription machinery.
26
Slide27Mechanisms which affect the chromatin structure and hence the expression of gene are:
Histone modifications
– These modifications make a region of gene either transcriptionally active or inactive.
Acetylation
↑Acetylation
↓
Condensation of DNA
↑ Transcription of genes in that region
Ubiquitination
Ubiquitination
of H2A – Transcriptional inactivation
Ubiquitination
of H2B - Transcriptional activation
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Slide28DNA methylation
: is
the addition or removal of a methyl group
predominately
where cytosine bases occur consecutively.
Methylation occurs most often in symmetrical
CG
sequences.
METHYLATION
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Slide29DNA methylation
Slide30DNA methylation is inheritedby daughter cells
Slide31DNA methylation in gene controlregion inactivates gene expression
CpG Island: DNA region with many CG sequences, of which C can become
methylated
.
In mammals, 70% to 80% of CpG cytosines are methylated.
Slide32Effects of DNA methylation-
Inactivation of a single gene expression- Inactivation of genes from part of chromosome- Inactivation of genes of whole chromosome (X)
Slide33Phosphorylation
Slide34Most important histone modifications
AcetylationMethylationPhosphorylation
•
Ubiquitination
For all these modifications, special enzymes are needed!
Slide35Histone-modification sites
Slide36Effects of histone-modifications on geneexpression (histone H3)
Slide37Humans have Hox genes in four clusters:
Genes
Chromosome
Cluster
HOXA1
,
HOXA2
,
HOXA3
,
HOXA4
,
HOXA5
,
HOXA6
,
HOXA7
,
HOXA9
,
HOXA10
,
HOXA11
,
HOXA13
chromosome 7
HOXA
HOXB1
,
HOXB2
,
HOXB3
,
HOXB4
,
HOXB5
,
HOXB6, HOXB7, HOXB8, HOXB9, HOXB13chromosome 17HOXBHOXC4, HOXC5, HOXC6, HOXC8, HOXC9, HOXC10, HOXC11, HOXC12, HOXC13chromosome 12HOXCHOXD1, HOXD3, HOXD4, HOXD8, HOXD9, HOXD10, HOXD11, HOXD12, HOXD13chromosome 2HOXD
Slide38Epigenetics ( above genetics)
Science studying heritable changes in gene expression, not resulting from changes in nucleotide sequence of DNA• Reversible modifications of chromatin organization
- DNA
- Histones
• mRNA modifications
Slide39Genetics vs. epigenetics
Slide40END Part II