Transcript Control of Gene Expression

Control of
Eukaryotic Genes
AP Biology
2007-2008
The BIG Questions…


How are genes turned on & off
in eukaryotes?
How do cells with the same genes
differentiate to perform completely
different, specialized functions?
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Evolution of gene regulation

Prokaryotes
single-celled
 evolved to grow & divide rapidly
 must respond quickly to changes in
external environment



exploit transient resources
Gene regulation

turn genes on & off rapidly


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flexibility & reversibility
adjust levels of enzymes
for synthesis & digestion
Evolution of gene regulation

Eukaryotes
multicellular
 evolved to maintain constant internal
conditions while facing changing
external conditions



homeostasis
regulate body as a whole

growth & development
 long term processes

specialization
 turn on & off large number of genes

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must coordinate the body as a whole rather
than serve the needs of individual cells
Points of control

The control of gene
expression can occur at any
step in the pathway from
gene to functional protein
1. packing/unpacking DNA
2. transcription
3. mRNA processing
4. mRNA transport
5. translation
6. protein processing
7. protein degradation
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1. DNA packing
How do you fit all
that DNA into
nucleus?

DNA coiling &
folding
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
double helix
nucleosomes
chromatin fiber
looped
domains
chromosome
from DNA double helix to
AP Biology chromosome
condensed
Nucleosomes
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8 histone
molecules
“Beads on a string”
1st level of DNA packing
 histone proteins
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8 protein molecules
positively charged amino acids
bind tightly to negatively charged DNA
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DNA
packing movie
DNA packing as gene control
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Degree of packing of DNA regulates transcription

tightly wrapped around histones
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
no transcription
genes turned off
 heterochromatin
darker DNA (H) = tightly packed
 euchromatin
lighter DNA (E) = loosely packed
H
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E
DNA methylation

Methylation of DNA blocks transcription factors


no transcription
 genes turned off
attachment of methyl groups (–CH3) to cytosine


nearly permanent inactivation of genes
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C = cytosine
ex. inactivated mammalian X chromosome = Barr body
Histone acetylation

Acetylation of histones unwinds DNA
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loosely wrapped around histones
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attachment of acetyl groups (–COCH3) to histones
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enables transcription
genes turned on
conformational change in histone proteins
transcription factors have easier access to genes
2. Transcription initiation

Control regions on DNA

promoter
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
enhancer
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
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nearby control sequence on DNA
binding of RNA polymerase & transcription
factors
“base” rate of transcription
distant control
sequences on DNA
binding of activator
proteins
“enhanced” rate (high level)
of transcription
Model for Enhancer action

Enhancer DNA sequences

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Activator proteins
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distant control sequences
bind to enhancer sequence
& stimulates transcription
Silencer proteins

bind to enhancer sequence
& block gene transcription
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Turning
on Gene movie
Transcription complex
Activator Proteins
• regulatory proteins bind to DNA at
Enhancer Sites
distant enhancer sites
• increase the rate of transcription
regulatory sites on DNA
distant from gene
Enhancer
Activator
Activator
Activator
Coactivator
A
E
F
B
TFIID
RNA polymerase II
H
Core promoter
and initiation complex
Initiation Complex at Promoter Site binding site of RNA polymerase
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3. Post-transcriptional control
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Alternative RNA splicing

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variable processing of exons creates a
family of proteins
4. Regulation of mRNA degradation

Life span of mRNA determines amount
of protein synthesis

mRNA can last from hours to weeks
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RNA
processing movie
RNA interference

Small interfering RNAs (siRNA)
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short segments of RNA (21-28 bases)
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bind to mRNA
create sections of double-stranded mRNA
“death” tag for mRNA
 triggers degradation of mRNA
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cause gene “silencing”
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post-transcriptional control
turns off gene = no protein produced
siRNA
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Action of siRNA
dicer
enzyme
mRNA for translation
siRNA
double-stranded
miRNA + siRNA
breakdown
enzyme
(RISC)
mRNA degraded
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functionally
turns gene off
siRNA clip
https://www.youtube.com/watch?v=Fa4sk
YBJHoI
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5. Control of translation

Block initiation of translation stage

regulatory proteins attach to 5' end of mRNA
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prevent attachment of ribosomal subunits &
initiator tRNA
block translation of mRNA to protein
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Control
of translation movie
6-7. Protein processing & degradation
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Protein processing
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folding, cleaving, adding sugar groups,
targeting for transport
Protein degradation
ubiquitin tagging
 proteasome degradation

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Protein
processing movie
1980s | 2004
Ubiquitin

“Death tag”
mark unwanted proteins with a label
 76 amino acid polypeptide, ubiquitin
 labeled proteins are broken down
rapidly in "waste disposers"

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AP
proteasomes
Aaron Ciechanover
Biology Israel
Avram Hershko
Israel
Irwin Rose
UC Riverside
Proteasome
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Protein-degrading “machine”
cell’s waste disposer
 breaks down any proteins
into 7-9 amino acid fragments
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cellular recycling
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play
Nobel animation
CENTRAL DOGMA
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Genetic information always goes
from DNA to RNA to protein
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Gene regulation has been well
studied in E. coli
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When a bacterial cell encounters a
potential food source it will
manufacture the enzymes
necessary to metabolize that food
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CENTRAL DOGMA

Genetic information always goes
from DNA to RNA to protein

Gene regulation has been well
studied in E. coli

When a bacterial cell encounters a
potential food source it will
manufacture the enzymes
necessary to metabolize that food
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Gene Regulation
In addition to sugars like glucose and
lactose E. coli cells also require
amino acids
 One essential aa is tryptophan.

When E. coli is swimming in
tryptophan (milk & poultry) it will
absorb the amino acids from the
media
 When tryptophan is not present in the
media then the cell must manufacture
its’ own amino acids

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Trp Operon

E. coli uses several proteins encoded by
a cluster of 5 genes to manufacture the
amino acid tryptophan
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All 5 genes are transcribed together as a
unit called an operon, which produces a
single long piece of mRNA for all the
genes

RNA polymerase binds to a promoter
located at the beginning of the first gene
and proceeds down the DNA transcribing
the genes in sequence
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Fig. 16.6
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GENE REGULATION

In addition to amino acids, E.
coli cells also metabolize
sugars in their environment
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In 1959 Jacques Monod and
Fracois Jacob looked at the
ability of E. coli cells to digest
the sugar lactose
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GENE REGULATION
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In the presence of the sugar lactose,
E. coli makes an enzyme called beta
galactosidase
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Beta galactosidase breaks down the
sugar lactose so the E. coli can
digest it for food
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It is the LAC Z gene in E coli that
codes for the enzyme beta
galactosidase
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Lac Z Gene
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The tryptophan gene is turned on
when there is no tryptophan in the
media
That is when the cell wants to make
its’ own tryptophan
E. coli cells can not make the sugar
lactose
They can only have lactose when it
is present in their environment
Then they turn on genes to beak
down lactose
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GENE REGULATION
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The E. coli bacteria only needs beta
galactosidase if there is lactose in the
environment to digest
There is no point in making the
enzyme if there is no lactose sugar to
break down
It is the combination of the promoter
and the DNA that regulate when a
gene will be transcribed
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6
7
Gene Regulation
protein
processing &
degradation
1 & 2. transcription
- DNA packing
- transcription factors
5
4
initiation of
translation
mRNA
processing
3 & 4. post-transcription
- mRNA processing
- splicing
- 5’ cap & poly-A tail
- breakdown by siRNA
5. translation
- block start of
translation
1 2
initiation of
transcription
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3
6 & 7. post-translation
- protein processing
- protein degradation
4
mRNA
protection
GENE REGULATION
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This combination of a promoter
and a gene is called an
OPERON
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Operon is a cluster of genes
encoding related enzymes that
are regulated together
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GENE REGULATION
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Operon consists of
A promoter site where RNA polyerase
binds and begins transcribing the
message
 A region that makes a repressor
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Repressor sits on the DNA at a spot
between the promoter and the gene to
be transcribed
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This site is called the operator
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LAC Z GENE
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EE.coli regulate the production of
Beta Galactocidase by using a
regulatory protein called a repressor
The repressor binds to the lac Z gene
at a site between the promotor and
the start of the coding sequence
The site the repressor binds to is
called the operator
.
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LAC Z GENE

Normally the repressor sits on
the operator repressing
transcription of the lac Z gene

In the presence of lactose the
repressor binds to the sugar
and this allows the polymerase
to move down the lac Z gene
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LAC Z GENE
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This results in the production
of beta galactosidase which
breaks down the sugar

When there is no sugar left the
repressor will return to its spot
on the chromosome and stop
the transcription of the lac Z
gene
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Concept 18.5: Cancer results from
genetic changes that affect cell cycle
control

The gene regulation systems that go wrong
during cancer are the very same systems
involved in embryonic development
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© 2011 Pearson Education, Inc.
Types of Genes Associated with
Cancer
 Cancer can be caused by mutations to genes

that regulate cell growth and division
Tumor viruses can cause cancer in animals
including humans
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© 2011 Pearson Education, Inc.
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Oncogenes are cancer-causing genes
Proto-oncogenes are the corresponding
normal cellular genes that are responsible for
normal cell growth and division
Conversion of a proto-oncogene to an
oncogene can lead to abnormal stimulation of
the cell cycle
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© 2011 Pearson Education, Inc.
Figure 18.23
Proto-oncogene
DNA
Translocation or
transposition: gene
moved to new locus,
under new controls
Gene amplification:
multiple copies of
the gene
New
promoter
Normal growthstimulating
protein in excess
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Point mutation:
within a control
within
element
the gene
Oncogene
Normal growth-stimulating
protein in excess
Normal growthstimulating
protein in
excess
Oncogene
Hyperactive or
degradationresistant
protein

Proto-oncogenes can be converted to
oncogenes by
Movement of DNA within the genome: if it
ends up near an active promoter, transcription
may increase
 Amplification of a proto-oncogene: increases
the number of copies of the gene
 Point mutations in the proto-oncogene or its
control elements: cause an increase in gene
expression

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© 2011 Pearson Education, Inc.
Tumor-Suppressor Genes

Tumor-suppressor genes help prevent
uncontrolled cell growth

Mutations that decrease protein products of
tumor-suppressor genes may contribute to
cancer onset

Tumor-suppressor proteins

Repair damaged DNA

Control cell adhesion

Inhibit the cell cycle in the cell-signaling
pathway
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© 2011 Pearson Education, Inc.
Interference with Normal CellSignaling Pathways
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Mutations in the ras proto-oncogene and p53
tumor-suppressor gene are common in human
cancers
Mutations in the ras gene can lead to
production of a hyperactive Ras protein and
increased cell division
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© 2011 Pearson Education, Inc.
Figure 18.24
MUTATION
1 Growth
factor
Ras
GTP
3 G protein
Ras
P
P
P
P
P
P
2 Receptor
4
2 Protein kinases
Hyperactive Ras protein
(product of oncogene)
issues signals on its
own.
GTP
MUTATION
3 Active
form
of p53
UV
light
Protein kinases
(phosphorylation
cascade)
5
1 DNA damage
in genome
Defective or missing
transcription factor,
such as
p53, cannot
activate
transcription.
DNA
NUCLEUS
Transcription
factor (activator)
Protein that
inhibits
the cell cycle
DNA
Gene expression
(b) Cell cycle–inhibiting pathway
Protein that
stimulates
the cell cycle
EFFECTS OF MUTATIONS
Protein
overexpressed
Protein absent
(a) Cell cycle–stimulating pathway
Cell cycle
overstimulated
(c) Effects of mutations
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Increased cell
division
Cell cycle not
inhibited
Figure 18.24a
MUTATION
1 Growth
factor
Ras
3 G protein
GTP
Ras
P
P
P
2 Receptor
P
P
P
Hyperactive Ras protein
(product of oncogene)
issues signals on its
own.
GTP
4 Protein kinases
(phosphorylation
cascade)
5
NUCLEUS
Transcription
factor (activator)
DNA
Gene expression
Protein that
stimulates
the cell cycle
(a) Cell cycle–stimulating pathway
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Figure 18.24b
2 Protein kinases
3 Active
form
of p53
UV
light
1 DNA damage
in genome
DNA
Protein that
inhibits
the cell cycle
(b) Cell cycle–inhibiting pathway
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MUTATION
Defective or missing
transcription factor,
such as
p53, cannot
activate
transcription.

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Suppression of the cell cycle can be important
in the case of damage to a cell’s DNA; p53
prevents a cell from passing on mutations due
to DNA damage
Mutations in the p53 gene prevent suppression
of the cell cycle
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© 2011 Pearson Education, Inc.
Figure 18.24c
EFFECTS OF MUTATIONS
Protein
overexpressed
Cell cycle
overstimulated
(c) Effects of mutations
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Protein absent
Increased cell
division
Cell cycle not
inhibited
The Multistep Model of Cancer
Development


Multiple mutations are generally needed for
full-fledged cancer; thus the incidence
increases with age
At the DNA level, a cancerous cell is usually
characterized by at least one active oncogene
and the mutation of several tumor-suppressor
genes
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© 2011 Pearson Education, Inc.
Figure 18.25
Colon
1 Loss
of tumorsuppressor
gene APC
(or other)
2 Activation
of ras
oncogene
4 Loss
of tumorsuppressor
gene p53
3 Loss
5 Additional
Colon wall
mutations
of tumorSmall benign suppressor
Larger
Normal colon
Malignant
growth
epithelial cells
tumor
gene DCC benign growth
(polyp)
(adenoma)
(carcinoma)
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Figure 18.25a
Colon
Colon wall
Normal colon
epithelial cells
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Figure 18.25b
1 Loss of tumorsuppressor gene
APC (or other)
Small benign
growth (polyp)
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Figure 18.25c
2 Activation of
ras oncogene
3
Loss of
tumor-suppressor
gene DCC
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Larger benign
growth (adenoma)
Figure 18.25d
4
Loss of
tumor-suppressor
gene p53
5 Additional
mutations
Malignant tumor
(carcinoma)
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Inherited Predisposition and Other
Factors Contributing to Cancer



Individuals can inherit oncogenes or mutant
alleles of tumor-suppressor genes
Inherited mutations in the tumor-suppressor
gene adenomatous polyposis coli are common
in individuals with colorectal cancer
Mutations in the BRCA1 or BRCA2 gene are
found in at least half of inherited breast
cancers, and tests using DNA sequencing can
detect these mutations
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© 2011 Pearson Education, Inc.
Figure 18.26
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Figure 18.UN01
Operon
Promoter
Genes
A
B
C
Operator
RNA
polymerase
A
B
C
Polypeptides
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Figure 18.UN02
Genes expressed
Genes not expressed
Promoter
Genes
Operator
Inactive repressor:
no corepressor present
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Active repressor:
corepressor bound
Corepressor
Figure 18.UN03
Genes not expressed
Promoter
Operator
Genes
Active repressor:
no inducer present
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Genes expressed
Inactive repressor:
inducer bound
Figure 18.UN04
Transcription
Chromatin modification
• Genes in highly compacted
chromatin are generally not
transcribed.
• Histone acetylation seems
to loosen chromatin structure,
enhancing transcription.
• DNA methylation generally
reduces transcription.
• Regulation of transcription initiation:
DNA control elements in enhancers bind
specific transcription factors.
Bending of the DNA enables activators to
contact proteins at the promoter, initiating
transcription.
• Coordinate regulation:
Enhancer for
Enhancer for
liver-specific genes
lens-specific genes
Chromatin modification
Transcription
RNA processing
RNA processing
• Alternative RNA splicing:
Primary RNA
transcript
mRNA
degradation
Translation
Protein processing
and degradation
mRNA
or
Translation
• Initiation of translation can be controlled
via regulation of initiation factors.
mRNA degradation
• Each mRNA has a
characteristic life span,
determined in part by
sequences in the 5 and
3 UTRs.
AP Biology
Protein processing and degradation
• Protein processing and
degradation by proteasomes
are subject to regulation.
Figure 18.UN04a
Chromatin modification
• Genes in highly compacted
chromatin are generally not
transcribed.
• Histone acetylation seems
to loosen chromatin structure,
enhancing transcription.
• DNA methylation generally
reduces transcription.
Transcription
• Regulation of transcription initiation:
DNA control elements in enhancers bind
specific transcription factors.
Bending of the DNA enables activators to
contact proteins at the promoter, initiating
transcription.
• Coordinate regulation:
Enhancer for
Enhancer for
lens-specific genes
liver-specific genes
Chromatin modification
Transcription
RNA processing
RNA processing
• Alternative RNA splicing:
Primary RNA
transcript
mRNA
degradation
Translation
Protein processing
and degradation
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mRNA
or
Figure 18.UN04b
Chromatin modification
Transcription
RNA processing
mRNA
degradation
Translation
Protein processing
and degradation
Translation
• Initiation of translation can be controlled
via regulation of initiation factors.
mRNA degradation
• Each mRNA has a
characteristic life span,
determined in part by
sequences in the 5 and
3 UTRs.
AP Biology
Protein processing and degradation
• Protein processing and
degradation by proteasomes
are subject to regulation.
Figure 18.UN05
Chromatin modification
Chromatin modification
• Small or large noncoding RNAs can
promote the formation of heterochromatin
in certain regions, blocking transcription.
Transcription
RNA processing
mRNA
degradation
Translation
• miRNA or siRNA can block the translation
of specific mRNAs.
Translation
Protein processing
and degradation
mRNA degradation
• miRNA or siRNA can target specific
mRNAs for destruction.
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Figure 18.UN06
Enhancer
Promoter
Gene 1
Gene 2
Gene 3
Gene 4
Gene 5
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Figure 18.UN07
Enhancer
Promoter
Gene 1
Gene 2
Gene 3
Gene 4
Gene 5
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Figure 18.UN08
Enhancer
Promoter
Gene 1
Gene 2
Gene 3
Gene 4
Gene 5
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