Chapter 19 - Eukaryotic Genomes
AIM: What is the effect of differentiated gene expression?
Gene Regulation
(controlling gene expression – turning genes
on/off)
Gene expression = Transcription and Translation of a gene; the cell
Chapter 19 - Eukaryotic Genomes
AIM: What is the effect of differentiated gene expression?
Multicellular organisms are composed of many
different types of cells…
Chapter 19 - Eukaryotic Genomes
AIM: What is the effect of differentiated gene expression?
What makes these cells different from each
other?
The same thing that makes a school different from a bank or a
police station different from a fire house…the workers (proteins)
are different!!
Differential gene expression
(Different cells have different genes turned on/off)
Chapter 19 - Eukaryotic Genomes
AIM: What is the effect of differentiated gene expression?
Chapter 19 - Eukaryotic Genomes
AIM: What is the effect of differentiated gene expression?
Chapter 19 - Eukaryotic Genomes
AIM: What is the effect of differentiated gene expression?
Stem Cells
What’s a stem cell?
- cells that have the ability to differentiate (to turn into / specialize) into a
specific cell type like a neuron or muscle cell. All of their genes have the
potential to be turned on/off.
Stem Cell
Chapter 19 - Eukaryotic Genomes
AIM: What is the effect of differentiated gene expression?
Stem Cells
More Stem Cells
Stem Cell
Stem Cells can divide to
make more stem cells or they
can differentiate.
Differentiated Cells
Chapter 19 - Eukaryotic Genomes
AIM: What is the effect of differentiated gene expression?
Stem Cells
Stem Cells
Stem Cell
Where do you
hypothesize you would
find stem cells?
Differentiated Cells
Chapter 19 - Eukaryotic Genomes
AIM: What is the effect of differentiated gene expression?
Stem Cells
Where do you
hypothesize you would
find stem cells?
1. Embryonic stem cells
- Found in embryos
- Totipotent – means they can become any cell
type (placenta, muscle, neural, epithelial, etc…)
- Pluripotent – means they can become any cell
type except the placenta (muscle, neural,
epithelial, etc…)
2. Adult stem cells
- Found throughout body after embryonic
development
- Multipotent – means they can become ONLY a
limited number of cell types depending on the
type of stem cell.
- ex. (figure to right) hematopoietic stem cells (HSC)
found in bone marrow of femur, hips, sternum, ribs
and other bones (see next slide) and can only become
either red or white blood cells.
Chapter 19 - Eukaryotic Genomes
AIM: What is the effect of differentiated gene expression?
Active
genes
Inactive
genes
Some genes are turned off for the life of the cell:
In differentiated cells, certain genes are
“permanently” shut down by histone packing
like the insulin gene in muscle cells. There is
no reason for muscle to ever make insulin.
Chapter 19 - Eukaryotic Genomes
AIM: What is the effect of differentiated gene expression?
Active
genes
Inactive
genes
Is is possible for a differentiated cell like a neuron or muscle
cell to dedifferentiate back to a stem cell? What would this
require?
Chapter 19 - Eukaryotic Genomes
AIM: What is the effect of differentiated gene expression?
Is is possible for a differentiated cell like a neuron or muscle
cell to dedifferentiate back to a stem cell? What would this
require?
It would require genes that have been “permanently”
turned off, typically by histone packing (more to come
shortly), to be turned back on.
Is this possible? How can we test this?
Active
genes
Inactive
genes
Chapter 19 - Eukaryotic Genomes
AIM: What is the effect of differentiated gene expression?
Is is possible for a differentiated cell like a neuron or muscle
cell to dedifferentiate back to a stem cell? What would this
require?
Let’s try a little experiment:
1. Let’s take an ovum (which is a stem cell of course) from some
multicellular organism like a sheep and remove the nucleus.
Chapter 19 - Eukaryotic Genomes
AIM: What is the effect of differentiated gene expression?
Is is possible for a differentiated cell like a neuron or muscle
cell to dedifferentiate back to a stem cell? What would this
require?
(somatic/differenti
ated cell’s nucleus)
Let’s try a little experiment:
2. Then let’s take the nucleus from a differentiated cell (let’s say a
muscle cell) and put it into the ovum (this is a diploid nucleus of
course).
Chapter 19 - Eukaryotic Genomes
AIM: What is the effect of differentiated gene expression?
Is is possible for a differentiated cell like a neuron or muscle
cell to dedifferentiate back to a stem cell? What would this
require?
(somatic/differenti
ated cell’s nucleus)
Let’s try a little experiment:
A. What do you predict should happen if differentiated cells cannot turn back
on the silenced (off) genes?
B. What if the genes can be turned back on?
Chapter 19 - Eukaryotic Genomes
AIM: What is the effect of differentiated gene expression?
Is is possible for a differentiated cell like a neuron or muscle
cell to dedifferentiate back to a stem cell? What would this
require?
Embryo = time between conception
(fertilization) until eight weeks old
Let’s try a little experiment:
3. It turns out that the genes can be reactivated (they are not permanently
turned off) and the “zygote” divides to become an embryo.
What would you try next?
Chapter 19 - Eukaryotic Genomes
AIM: What is the effect of differentiated gene expression?
Is is possible for a differentiated cell like a neuron or muscle
cell to dedifferentiate back to a stem cell? What would this
require?
Let’s try a little experiment:
4. We can try to implant the early embryo (blastocyst) into the uterus of a
surrogate mother (a black face ewe in this case) and see what happens…
Chapter 19 - Eukaryotic Genomes
AIM: What is the effect of differentiated gene expression?
Is is possible for a differentiated cell like a neuron or muscle
cell to dedifferentiate back to a stem cell? What would this
require?
Let’s try a little experiment:
5. Amazingly, the embryo develops and the lamb is born.
Is this lamb, a clone, genetically identical to the ovum donor, surrogate
nucleusdonor?
donor as the nucleus contained the DNA
mother or theThe
nucleus
Chapter 19 - Eukaryotic Genomes
AIM: What is the effect of differentiated gene expression?
Is is possible for a differentiated cell like a neuron or muscle
cell to dedifferentiate back to a stem cell? What would this
require?
This process is called REPRODUCTIVE CLONING.
Does this answer the above question?
This indicates that genes in a differentiated nucleus have the “potential” to
reactivate and therefore differentiated cells IN THEORY can dedifferentiate.
Chapter 19 - Eukaryotic Genomes
AIM: What is the effect of differentiated gene expression?
Is is possible for a differentiated cell to
dedifferentiate back to a stem cell?
REPRODUCTIVE CLONING
Dolly (left) and her
surrogate mother. A black
face sheep cannot give
birth to a white face sheep
Chapter 19 - Eukaryotic Genomes
AIM: What is the effect of differentiated gene expression?
What could reproductive cloning be used for?
1. Repopulating endangered species…is there a
They are all genetically identical and therefore equally
problem?
susceptible to the same environmental changes…
2. Clone drug-producing animals (pharm animals)
3. Clone genetically-unique animals, etc…
Should we do this with humans?
What if you had a
reproductive clone. One
day you fell ill and needed
part of a liver or a kidney or
bone
marrow?...
There
are
arguments on both
Chapter 19 - Eukaryotic Genomes
AIM: What is the effect of differentiated gene expression?
How many different animals have been cloned thus
far?
At least 20 ranging from camels, cats, dogs, a horse
all the way to fish, frogs and fruit flies.
Cloned cats…
Chapter 19 - Eukaryotic Genomes
AIM: What is the effect of differentiated gene expression?
How many different animals have been cloned thus
far?
At least 20 ranging from camels, cats, dogs, a horse
all the way to fish, frogs and fruit flies.
Cloned cats…that have been
genetically modified (next chapter)
Chapter 19 - Eukaryotic Genomes
AIM: What is the effect of differentiated gene expression?
?
What else could we do with this embryo?
Chapter 19 - Eukaryotic Genomes
AIM: How are stem cells generated and used?
We can grow them in a dish (culture them) and then treat the
cells with different hormones to get them to differentiate into
Chapter 19 - Eukaryotic Genomes
AIM: How are stem cells generated and used?
What can we use these differentiated cells for?
One could make any cell type they want:
1. Skin cells for burn victims
2. Organs for transplant patients
3. Neurons for a person with a
spinal cord injury
4. Basic scientific research, etc…
What is the advantage of these cells over other neurons
or organs in terms of transplants?
These transplanted cells will not be rejected (destroyed by the immune
system) because they are genetically identical to the patient (their
antibodies will not bind to them).
Chapter 19 - Eukaryotic Genomes
AIM: How are stem cells generated and used?
This form of cloning is called Therapeutic Cloning.
The nucleus would obviously be one of your nuclei and the ovum would come from a donor…
Chapter 19 - Eukaryotic Genomes
AIM: How are stem cells generated and used?
Ethics
Should we be allowed to generate embryos for
the sake of using the embryonic stem cells for
research/medicine?
Chapter 19 - Eukaryotic Genomes
AIM: How are stem cells generated and used?
Recent advances:
In 2008, scientists at UCLA figured out how to turn skin
cells into embryonic stem cells, alleviating the need for
cloning and embryo destruction
Kathrin Plath, UCLA stem cell scientists
http://www.sciencedaily.com/releases/2008/02/080211172631.htm
Chapter 19 - Eukaryotic Genomes
AIM: How are stem cells generated and used?
Bone Marrow Transplant is Stem Cell Treatment
Ex. Patient with Leukemia (white
blood cell [leukocyte] cancer).
Destroy all white blood cells of
patient using
radiation/chemotherapy.
Take bone marrow from
matching donor and infuse
patient with hematopeotic stem
cells.
Chapter 19 - Eukaryotic Genomes
AIM: Do differentiated cells retain their genetic potential?
Bone Marrow Transplant Cures HIV (aside):
http://www.nature.com/nm/journal/v15/n4/full/nm0409-371.html
Chapter 19 - Eukaryotic Genomes
AIM: Do differentiated cells retain their genetic potential?
Using Stem Cells with Gene Therapy
Gene Therapy involves replacing a mutated gene within an already developed
organism with a functional gene (somatic gene therapy) or replacing a gene in a
germ line cell (sperm of egg) resulting in a heritable change (germ line gene
therapy).
Chapter 19 - Eukaryotic Genomes
AIM: Do differentiated cells retain their genetic potential?
Where else do we observe already differentiated
cells dedifferentiating and becoming other cells
types?
Regeneration
- Regrowth of a lost of damaged body
part
Chapter 19 - Eukaryotic Genomes
AIM: Do differentiated cells retain their genetic potential?
Can differentiated cells dedifferentiate into stem cells in
plants?
Chapter 19 - Eukaryotic Genomes
AIM: Do differentiated cells retain their genetic potential?
Fig. 11.3A
Chapter 19 - Eukaryotic Genomes
AIM: How are stem cells generated and used?
Review
1. Stem Cells
- Embryonic vs. adult stem cells
(toti/pluripotent)
(multipotent)
- Therapeutic vs. Reproductive cloning
Chapter 19 - Eukaryotic Genomes
AIM: How are stem cells generated and used?
Not all genes are going to be silenced for the life
of the cell/organism, many will be turned on/off
as needed…
Ex. The genes coding for enzymes that make
glycogen in the liver…
If the blood glucose concentration is low, the liver
will be releasing glucose, not building glycogen
from it. Therefore, the genes should be off.
Likewise the genes whose protein products are
involved in secreting glucose should be on.
Gene are CONSTANTLY being turned on and off
in Let’s
yourlook
cells
at how this is accomplished in eukaryotes.
Chapter 19 - Eukaryotic Genomes
NEW AIM: How are genes regulated in eukaryotes?
How are eukaryotic genes
regulated?
Chapter 19 - Eukaryotic Genomes
AIM: How are genes regulated in eukaryotes?
1.
2.
3.
4.
5.
6.
7.
8.
DNA/Chromatin packing
Transcription initiation
Splicing (RNA processing)
Transport to Cytoplasm
mRNA degradation
Translation initiation
Protein modification/activation
Protein Breakdown
Fig. 19.3
Chapter 19 - Eukaryotic Genomes
AIM: How are genes regulated in eukaryotes?
Eukaryotic gene regulation
1. DNA/Chromatin Packing
Histones (“beads”) can pack genes or entire
segments of DNA (“string”) tightly such that
transcription factors and RNA polymerases
cannot access the DNA. These gene are
typically turned off for the life of the cell.
Fig. 11.6
Chapter 19 - Eukaryotic Genomes
AIM: How are genes regulated in eukaryotes?
Eukaryotic gene regulation
1. DNA/Chromatin Packing
What is a nucleosome’s
structure?
1. The core is composed of 8 proteins (H2A,
H2B, H3 and H4 – two of each) known as
histones. DNA wraps twice around the core.
The N-terminal tails of the histones hang out
from the nucleosome.
2. Another histone (H1), not technically part
of the nucleosome, clamps the DNA to the
core.
Fig. 11.6
Chapter 19 - Eukaryotic Genomes
AIM: How are genes regulated in eukaryotes?
Eukaryotic gene regulation
1. DNA/Chromatin Packing
Euchromatin
-DNA wrapped around nucleosomes.
- Nucleosomes not bound to each other
- This is the form of an active gene
(a gene that can be transcribed if
desired by RNA polymerase)
Heterochromatin
- Nucleosomes binds to each other
with help of additional histone
called H1 condensing the DNA.
- These genes are silenced and
cannot be transcribed.
Ex. Gene for insulin in cells other
than pancreatic beta cells.
Chapter 19 - Eukaryotic Genomes
AIM: How are genes regulated in eukaryotes?
Eukaryotic gene regulation
1. DNA/Chromatin Packing
Euchromatin
How does the chromatin stay in this
“loose” euchromatin conformation?
Histone Acetylation
The N-terminal tails have the amino
acid lysine to which an acetyl group
is added preventing the
nucleosomes from packing.
= acetyl (memorize it)
Chapter 19 - Eukaryotic Genomes
AIM: How are genes regulated in eukaryotes?
Eukaryotic gene regulation
1. DNA/Chromatin Packing
Histones Regulation
In addition to acetylation,
histones can be modified on
their N-termini a number of
other ways as shown in this
figure.
For example, methylation
appears to promote
condensation.
-CH3 = methyl (memorize it)
Chapter 19 - Eukaryotic Genomes
AIM: How are genes regulated in eukaryotes?
Eukaryotic gene regulation
1. DNA/Chromatin Packing
These additional levels of
condensing require non-histone
proteins and occur only during
prophase
Chapter 19 - Eukaryotic Genomes
AIM: How are genes regulated in eukaryotes?
Eukaryotic gene regulation
Fig. 11.6
Ex. One of the X chromosomes in XX females (humans
included) is randomly silenced by histones. Females, like
males, only have one active X chromosome. The other is
Chapter 19 - Eukaryotic Genomes
AIM: How are genes regulated in eukaryotes?
Eukaryotic gene regulation
Fig. 11.6
Ex. One of the X chromosomes in XX females (humans
included) is randomly silenced by histones. Females, like
males, only have one active X chromosome. The other is
Chapter 19 - Eukaryotic Genomes
AIM: How are genes regulated in eukaryotes?
Eukaryotic gene regulation
DNA methylation (an aside)
In addition to histone methylation, most plants and animals use methylation
of the DNA itself on the base of cytosine (see below) to regulate gene
expression:
DNA methyltransferase
Excessive methylation of a gene appears to be associated with turning a
gene off…
Chapter 19 - Eukaryotic Genomes
AIM: How are genes regulated in eukaryotes?
Eukaryotic gene regulation
DNA methylation as a mode of epigenetic inheritance
What is epigenetics?
Any modification to the
genome that results in a
change in function, but does
not change the DNA sequence
Ex. Environmental chemicals,
DNA methylation, histone
acetylation.
Epigenetic inheritance - Some
of these modifications can be
inherited (i.e. DNA methylation
patterns) making them
significant in terms of
diversity and evolution.
http://classic.the-scientist.com/blog/display/55342/
Chapter 19 - Eukaryotic Genomes
AIM: How are genes regulated in eukaryotes?
1.
2.
3.
4.
5.
6.
7.
8.
DNA/Chromatin packing
Transcription initiation
Splicing (RNA processing)
Transport to Cytoplasm
mRNA degradation
Translation initiation
Protein modification/activation
Protein Breakdown
Fig. 19.3
Recall Transcription
Chapter 19 - Eukaryotic Genomes
AIM: How are genes regulated in eukaryotes?
Eukaryotic gene regulation
2. Transcription Initiation
Recall the structure of a
eukaryotic gene:
Fig. 11.8
Chapter 19 - Eukaryotic Genomes
AIM: How are genes regulated in eukaryotes?
Eukaryotic gene regulation
Control elements
2. Transcription Initiation
Transcription factors (TF’s) are
required to start transcription.
A. General transcription factors are
required for the transcription of all
genes. These are the ones that bind at
the promoter and interact with RNA
polymerase II.
NO TF’s, NO Transcription
Enhancer
proteins
General
Transcriptio
n factors
Fig. 11.8
Chapter 19 - Eukaryotic Genomes
AIM: How are genes regulated in eukaryotes?
Eukaryotic gene regulation
2. Transcription Initiation
Transcription factors (TF’s) are
required to start transcription.
All of the TF’s in this diagram are general
TF’s needed by every gene to be
transcribed.
Don’t memorize this level of detail
unless you have nothing else to
do. First email me though and I
will find you something else to
Chapter 19 - Eukaryotic Genomes
AIM: How are genes regulated in eukaryotes?
Eukaryotic gene regulation
Control elements
2. Transcription Initiation
Transcription factors (TF’s) are
required to start transcription.
A. General transcription factors are
required for the transcription of all
genes. These are the ones that bind at
the promoter and interact with RNA
polymerase II.
B. Specific transcription factors (either
activators or repressors) will bind at
DNA sequences called control
elements (enhancer regions or
repressor regions)distant from the
gene itself and turn the gene on or off.
Enhancer
proteins
General
Transcriptio
n factors
Fig. 11.8
Chapter 19 - Eukaryotic Genomes
AIM: How are genes regulated in eukaryotes?
Eukaryotic gene regulation
2. Transcription Initiation
Recap:
Control element = enhancer or repressor
region
Activator TF’s bind enhancer regions
and promote gene expressions
Repressors TF’s bind to repressor
regions and inhibit gene expression
Mediator Proteins
Mediate (bridge) the interaction
between activators and general
TF’s
Fig. 11.8
Chapter 19 - Eukaryotic Genomes
AIM: How are genes regulated in eukaryotes?
Eukaryotic gene regulation
2. Transcription Initiation
Combinatorial Gene Activation
Genes are typically regulated by a number of
different enahancer/repressor regions.
Therefore, activation requires a
number of different activators to be
present at the same time:
Chapter 19 - Eukaryotic Genomes
AIM: How are genes regulated in eukaryotes?
Eukaryotic gene regulation
2. Transcription Initiation
EXAMPLE:
a. A signal molecule (ligand) like growth
factor will bind to a surface receptor.
b. Signal transduction occurs (a story you
should know well…) and a TF is activated
usually via phosphorylation.
c. This TF, assuming it to be an activator,
will undergo a conformation change
resulting in exposure of the nuclear
localization signal allowing entrance to the
nucleus. It will then bind to specific
enhancer control elements to promote
expression of these genes.
Chapter 19 - Eukaryotic Genomes
AIM: How are genes regulated in eukaryotes?
1.
2.
3.
4.
5.
6.
7.
8.
DNA/Chromatin packing
Transcription initiation
Splicing (RNA processing)
Transport to Cytoplasm
mRNA degradation
Translation initiation
Protein modification/activation
Protein Breakdown
Fig. 19.3
Chapter 19 - Eukaryotic Genomes
AIM: How are genes regulated in eukaryotes?
Eukaryotic gene regulation
3. Alternative RNA splicing
- Alternative splicing
can control how much
mRNA is synthesized
of each alternative
transcript.
Fig. 11.9
Chapter 19 - Eukaryotic Genomes
AIM: How are genes regulated in eukaryotes?
Eukaryotic gene regulation
3. Alternative RNA splicing
Many proteins are involved
in regulating which splice
variant is formed…
Don’t memorize…
Fig. 11.9
Chapter 19 - Eukaryotic Genomes
AIM: How are genes regulated in eukaryotes?
1.
2.
3.
4.
5.
6.
7.
8.
DNA/Chromatin packing
Transcription initiation
Splicing (RNA processing)
Transport to Cytoplasm
mRNA degradation
Translation initiation
Protein modification/activation
Protein Breakdown
Fig. 19.3
Chapter 19 - Eukaryotic Genomes
AIM: How are genes regulated in eukaryotes?
Eukaryotic gene regulation
5. mRNA degradation
A. Generic degradation
Mediated by enzymatic
removale of tail and cap
followed by nuclease digestion.
Chapter 19 - Eukaryotic Genomes
AIM: How are genes regulated in eukaryotes?
Eukaryotic gene regulation
5. mRNA degradation
B. RNA interference (RNAi)
In the case of RNAi, a gene
codes for a special type of
RNA called a small
interfering RNA (siRNA)
that is complementary to
the target mRNA.
In the end, a enzyme called RISC
(RNA-induced silencing complex)
binds to the siRNA, which anneals
to the mRNA. RISC then cuts the
mRNA or prevents translation…
Chapter 19 - Eukaryotic Genomes
AIM: How are genes regulated in eukaryotes?
Eukaryotic gene regulation
5. mRNA degradation
B. RNA interference (RNAi)
Figure from book:
DICER is an enzyme that cuts the initial RNA transcript resulting in smaller
pieces that will function as siRNA (or micro RNA, miRNA).
Chapter 19 - Eukaryotic Genomes
AIM: How are genes regulated in eukaryotes?
Eukaryotic gene regulation
5. mRNA degradation
B. RNA interference (RNAi) as a
tool
This means we can shut
down any mRNA we want
by sending in a
complementary RNA that
can be recognized by
DICER.
http://www.nature.com/nrg/multimedia/rnai/index.html
Chapter 19 - Eukaryotic Genomes
AIM: How are genes regulated in eukaryotes?
1.
2.
3.
4.
5.
6.
7.
8.
DNA/Chromatin packing
Transcription initiation
Splicing (RNA processing)
Transport to Cytoplasm
mRNA degradation
Translation initiation
Protein modification/activation
Protein Breakdown
Fig. 19.3
Chapter 19 - Eukaryotic Genomes
AIM: How are genes regulated in eukaryotes?
Eukaryotic gene regulation
6. Translation Initiation
Like transcription, translation also requires
other proteins to start called initiation
factors (IF’s - prokaryotes) or elongation
factors (elF’s – eukaryotes (that is what
the “e” is for).
NO IF’s, NO Translation
Chapter 19 - Eukaryotic Genomes
AIM: How are genes regulated in eukaryotes?
Eukaryotic gene regulation
6. Translation Initiation
Like transcription, translation also requires
other proteins to start called initiation
factors (IF’s) or elongation factors (elF’s).
Eukaryotes of course are more complicated…
Don’t memorize, just understand concept
Chapter 19 - Eukaryotic Genomes
AIM: How are genes regulated in eukaryotes?
1.
2.
3.
4.
5.
6.
7.
8.
DNA/Chromatin packing
Transcription initiation
Splicing (RNA processing)
Transport to Cytoplasm
mRNA degradation
Translation initiation
Protein modification/activation
Protein Breakdown
Fig. 19.3
Chapter 19 - Eukaryotic Genomes
AIM: How are genes regulated in eukaryotes?
Eukaryotic gene regulation
Fig. 11.10
7. Protein activation
(pre-insulin)
A. Activation by Proteolysis (cutting the protein)
Insulin is made as a single polypeptide, which then fold into its
inactive form. An enzyme will cut (cleave) the polypeptide
forming the active protein form of insulin.
Chapter 19 - Eukaryotic Genomes
AIM: How are genes regulated in eukaryotes?
Eukaryotic gene regulation
Fig. 11.10
7. Protein activation
(pre-insulin)
B. Phosphorylation
Activation/Inactivation through addition of a phosphate, which
you should be very familiar with at this point.
Chapter 19 - Eukaryotic Genomes
AIM: How are genes regulated in eukaryotes?
1.
2.
3.
4.
5.
6.
7.
8.
DNA/Chromatin packing
Transcription initiation
Splicing (RNA processing)
Transport to Cytoplasm
mRNA degradation
Translation initiation
Protein modification/activation
Protein Breakdown
Fig. 19.3
Chapter 19 - Eukaryotic Genomes
AIM: How are genes regulated in eukaryotes?
Eukaryotic gene regulation
8. Protein Degradation
When a protein is no longer
needed (the cell has enough
product of a certain enzyme) it can
broken
down –into its amino
be degraded
acids, which are then recycled
into new polypeptides.
This is accomplished by a large
assembly (complex) of proteins called
the proteosome.
It is really a “polypeptide shredder”.
Chapter 19 - Eukaryotic Genomes
AIM: How are genes regulated in eukaryotes?
Eukaryotic gene regulation
A single ubiquitin protein
8. Protein Degradation
What is the signal to degrade a particular protein?
Ubiquitination
The protein to be degraded is tagged. It is marked by the enzymatic addition of
ubiquitin (a small protein itself). Not just one, but many in a row shown as green
spheres below. Ubiquitin in the mark of death. If a chain of them are attached to
you, you will be shredded.
AIM: How are genes regulated (controlled) in eukaryotes?
Chapter 19 - Eukaryotic Genomes
AIM: How are genes regulated in eukaryotes?
1.
2.
3.
4.
5.
6.
7.
8.
DNA/Chromatin packing
Transcription initiation
Splicing (RNA processing)
Transport to Cytoplasm
mRNA degradation
Translation initiation
Protein modification/activation
Protein Breakdown
Clearly more complex than prokaryotes…
Fig. 19.3
Chapter 19 - Eukaryotic Genomes
NEW AIM: What is the genetic basis of cancer?
How does one get cancer?
Genetic change falling into one of three
categories:
1. Mutation
2. Movement of DNA within the
genome
3.
Amplification of a gene
Let’s look at what this means…
Chapter 19 - Eukaryotic
Genomes
AIM: What is the genetic
basis of cancer?
Signal transduction pathway
- process by which the cell
converts one signal into
another
In this case (to the right) an
external signal is converted
into an internal signal
through relay proteins.
Fig. 11.15A
Chapter 19 - Eukaryotic
Genomes
growth factor (GF)
AIM: What is the genetic
basis of cancer?
Let’s say the signal
molecule (ligand) is a
growth factor (GF) and the
new proteins being made
activates cell division
(instructs the cell to
produce cyclin and proceed
past the G1 checkpoint).
Activate
division
Fig. 11.15A
Chapter 19 - Eukaryotic
Genomes
AIM: What is the genetic
basis of cancer?
If there were no growth
factor there should be no…
Activate
division
Fig. 11.15A
Chapter 19 - Eukaryotic
Genomes
AIM: What is the genetic
basis of cancer?
If there were no growth
factor there should be no…
…new protein being made
and cell division should….
be off.
Q. What if there is a
mutation in the gene of the
receptor or one of the relay
proteins that changes its
shape so that it is always
on?
Chapter 19 - Eukaryotic
Genomes
AIM: What is the genetic
basis of cancer?
The transduction pathway will
always be on regardless of
growth factor…
This can lead to uncontrolled
cell division…cancer.
Fig. 11.16A
Chapter 19 - Eukaryotic
Genomes
AIM: What is the genetic
basis of cancer?
Proto-oncogene
A normal gene that when modified
causes cancer is called a protooncogene.
Oncogene
The modified form of the gene that
causes cancer. Does not NEED to be a
mutation…
Proto = “before”
oncos = “tumor” or cancer
Gene = gene
Chapter 19 - Eukaryotic Genomes
AIM: What is the genetic basis of cancer?
Fig. 19.11
Amplification (overproduction)
How a proto-oncogene can become an oncoge
Let’s look at some specific examples…
Chapter 19 - Eukaryotic Genomes
AIM: What is the genetic basis of cancer?
Ras
Fig. 19.11
Ras is a G protein normally activated by an RTK pathway.
Ras is commonly mutated in many cancers resulting in a
Chapter 19 - Eukaryotic Genomes
AIM: What is the genetic basis of cancer?
Fig. 11.15A
If you get a mutation in one proto-oncogene like
Ras, does that mean you get cancer?
No, it takes more than one mutation in one gene to
cause cancer…read on about tumor suppressor
Chapter 19 - Eukaryotic
Genomes
AIM: What is the genetic
basis of cancer?
Cells have genes that code for
proteins that inhibit cell
division called tumor
suppressor genes.
They can be:
1. TF’s that activate proteins, which
prevent cell division or cause
apoptosis like p53.
Let’s look at p53 in more detail…
Chapter 19 - Eukaryotic Genomes
AIM: What is the genetic basis of cancer?
p53
“The guardian angel of the genome”
p53 (53 for 53,000-dalton mw) is a TF that is activated in
response to excessive DNA damage through a phosphorylation
cascade
(see above).
- p53 turns on genes involved in inhibiting the cell cycle and if DNA
damage is too great, apoptosis genes as well.
Chapter 19 - Eukaryotic
Genomes
AIM: What is the genetic
basis of cancer?
Cells have genes that code for
proteins that inhibit cell
division called tumor
suppressor genes.
They can be:
1. TF’s that activate proteins, which
prevent cell division or cause
apoptosis like p53.
OR
2. DNA repair proteins like BRCA-1
and BRCA-2, which prevent
mutations obviously.
Chapter 19 - Eukaryotic Genomes
AIM: What is the genetic basis of cancer?
Both BRCA1 and BRCA2 are
DNA repair proteins – fix DNA
breaks.
Mutations in the BRCA1 gene
increase the risk breast,
ovarian, Fallopian tube,
prostate and colon cancers.
Over 600 different mutations have
been identified
Among breast cancer patients of
Jewish ancestry, 10% had mutations
in one of these two genes.
Fig. 11.16B
Chapter 19 - Eukaryotic
Genomes
AIM: What is the genetic
basis of cancer?
Based on what you have learned
thus far, what genetic changes are
necessary to cause cancer?
Chapter 19 - Eukaryotic
Genomes
AIM: What is the genetic
basis of cancer?
You would need a mutation in BOTH
tumor suppressor genes and one
oncogene…
why both tumor supressor genes?
Just because you knocked out one, the other
can still function and stop the division (two hit
hypothesis).
Why don’t both protooncogenes need to be
These
proteins activate and you only need one
modified/mutated?
oncogene to activate the pathway.
Chapter 19 - Eukaryotic
Genomes
AIM: What is the genetic
basis of cancer?
Additional genetic changes are
typically required like activation of
telomerase and genes involved in
cell migration.
Explain.
Telomerase is needed to maintain the length of
the ends of chormosomes (telomeres) since
they shorten with each division thanks to the
lagging strand, and in order to be cancerous
the cells need to be able to migrate.
Chapter 19 - Eukaryotic Genomes
AIM: What is the genetic basis of cancer?
Fig. 11.17A
Chapter 19 - Eukaryotic Genomes
AIM: What is the genetic basis of cancer?
Predisposition to Cancer
Reminder: BRCA1 (BReast CAncer) and
BRCA2 are DNA repair proteins – fix
DNA breaks.
Breast Cancer is the second
most common type of cancer
next to Prostate Cancer.
~230,000 new cases a year in
females
Inheriting one mutated BRCA1 allele
gives you a 60% chance of
developing breast cancer before the
age of 50 compared to 2% normally.
Fig. 11.16B
Chapter 19 - Eukaryotic Genomes
AIM: What is the genetic basis of cancer?
Fig. 11.17A
Chapter 19 - Eukaryotic Genomes
AIM: What is the genetic basis of cancer?
Conclusion:
1. Multiple mutations are required for cancer to occur
a. A proto-oncogene must be modified by one of the
methods discussed to an oncogene promoting cell
growth
b. Tumor suppressor genes must be rendered inactive so
they don’t inhibit division or cause apoptosis.
c. Additional genetic changes must occur like activation
of telomerase and/or genes involved in motility.
Chapter 19 - Eukaryotic Genomes
AIM: What is the genetic basis of cancer?
Old chart, prostate
now higher than
breast…
Chapter 19 - Eukaryotic Genomes
AIM: Eukaryotic non-coding regions
Eukaryotic genomes consist
mostly of non-coding regions
in addition to genes called
“junk DNA”…is it really junk
though??
A. 98.5% of our genome does
NOT code for mRNA, tRNA or
rRNA!!!
B. Most of this DNA is repetitive
DNA (DNA sequence of various
length that just keep repeating over
and over)
C. ~44% of human genome
consists of transposable
elements!
Chapter 19 - Eukaryotic Genomes
AIM: Eukaryotic non-coding regions
Transposable Elements
Two types in Eukaryotes
1. Transposons
2. Retrotransposons
-most transposable elements
in eukaryotes are retro
Chapter 19 - Eukaryotic Genomes
AIM: Eukaryotic non-coding regions
Not all of our genes exist as isolated
islands in the genome:…
A. 50% of our genes are arranged in
multigene families – collections of similar or
identical genes.
B. The genes coding for the three pieces
or rRNA (18S, 5.8S and 28S) are
grouped and this group is repeated 100’s
to 1000’s of times so that ribosomes can
be made super quickly and efficientyl.
The three genes are made as a single
transcript and then cleaved apart.
C. The globin genes (code for hemoglobin
subunits) are clustered together as well.
Hemoglobin is composed of two alpha and
two beta subunits. The alpha subunits are
clustered on chromosome 16 and the betas
on chromosome 11.
Chapter 19 - Eukaryotic Genomes
AIM: Eukaryotic non-coding regions
Gene duplication followed by mutation is key in evolution
Ex. The hemoglobin subunits
Duplication of an ancestral
globin gene freed one up to be
mutated resulting in two alleles
(alpha and beta). This was
followed by transposition as they
are on different chromosomes,
more duplication and then more
mutation…
Different exons (colored boxes)
Chapter 19 - Eukaryotic Genomes
AIM: Eukaryotic non-coding regions
Exon Shuffling within the genome
can lead to the evolution of new
genes:
Shown to the right are a series of genes in
humans each composed of numerous exons as
indicated by colored boxes.
Notice how the many different genes have
similar exons (same color box) and many of the
exons are repeated in a given gene.
It is clear that exons are being moved around
and duplicated through time resulting in the
evolution of new genes…
genes