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CHAPTER 19
THE ORGANIZATION AND CONTROL OF
EUKARYOTIC GENOMES
=> Larger than prokaryotes
Not all 25,000 genes are
active in all cells
Junk DNA anyone?
• Only 25,000
genes in
humans - 3%
of total DNA in
cell (YIKES!)
• Rest of it (97%)
- junk?????
(noncoding
DNA)
•Prokaryotes- most of the DNA in a
genome codes for protein (or tRNA and
rRNA), with a small amount of noncoding
DNA, primarily regulators.
•Eukaryotes - most of the DNA (about
97% in humans) does not code for protein
or RNA.
96%
similar
to
humans
)
“Noncoding” DNA and what it does in the
eukaryotic genome
1) Some noncoding regions are
regulatory sequences
(these are promotors and
enhancers that can increase
binding of RNA polymerase
to DNA).
2) Other are introns.
3) Finally, even more of it
consists of repetitive DNA,
present in many copies in the
genome.
REPPPPETITIVE DNA IN EUKARYOTES
• Know 2 types of Repetitive
GTTACGTTACGTTAC….
DNA:
repeated 10 to 10 million
times (Satellite DNA)
1)TANDEMLY REPETITIVE
DNA (AKA SATELLITE DNA
- *3 types) - 10 to 15% of
DNA
2) INTERSPERSED
REPETITIVE DNA - 25 - 40%
of DNA
REPPPPETITIVE DNA IN EUKARYOTES
• 1) SATELLITE DNA/TANDEMLY
REPETITIVE DNA
• These sequences (1 to 10 base pairs)
are repeated up to a million times in
series.
• GTTACGTTACGTTAC….
• 3 types:
a) Regular Satellite -100,000 - 10 mill
b) Minisatellite -100 -100,000 repeats
c) Microsatellite - 10 to 100 repeats Very important for forensics - helps
figure out uniqueness of a person’s
DNA
• A number of genetic disorders are
caused by abnormally long stretches of
tandemly repeated nucleotide triplets
within the affected gene.
CAG
Repeat
CAG Repeat Size
Median Age at Onset *
(years)
(95% confidence interval)
39
66 (72-59)
40
59 (61-56)
41
54 (56-52)
42
49 (50-48)
43
44 (45-42)
44
42 (43-40)
45
37 (39-36)
46
36 (37-35)
47
33 (35-31)
48
32 (34-30)
49
28 (32-25)
50
27 (30-24)
You know that antisocial neighbor May be a microsatellite problem!
• Satellite DNA plays a structural role at
telomeres and centromeres. This is
important! You don’t want non-repetitive
DNA in telomeres because?
CSI Lab on this coming upMicrosatellites – more repeats but really
short!
• Are only 1-10
nucleotides long and
are repeated only 10100 times in the
genome
• Used in DNA
fingerprinting
(forensics)
• What, more “junk”?
• (2) About 25-40% of most mammalian genomes
consists of interspersed repetitive DNA.
-One common family of interspersed repetitive
sequences, Alu elements, is transcribed into
RNA molecules with unknown roles in the cell.
-Alu sequences may help alternate RNA splicing
-Transposons are interspersed repetitive DNA
Table 19.1
bottom
Out of 25,000 genes what gets
expressed depends upon:
• Type of cell: Not all genes
are expressed in all cells
(epigenetics controls it)
• Development period:
During embryonic
development certain
genes may be expressed
that are not expressed in
adults (and viceversa)
Gene families - collection of genes
that may be identical/nonidentical
Gene families have evolved by
duplication of ancestral genes
• Most genes are present as a single copy per
haploid set of chromosomes
• Multigene families exist as a collection of
identical or very similar genes
(exceptions).
• These likely evolved from a single ancestral
gene.
• The members of multigene families may be
clustered or dispersed in the genome.
• Identical genes are multigene families that
are clustered tandemly.
• Evolution - first duplicate a gene and then mutate the copy;
result : original copy is still there, mutated gene - could make a
new protein = new function (natural selection acts on it)
• Nonidentical genes have diverged since their initial duplication
event.
PseudogenesDNA segments that have sequences similar to real genes
but that do not yield functional proteins - remnants of evolution or?
Did you know your genome changes
continually in your lifetime?
1) Rare mutations (between 1/106 and 1/105
nucleotides )
2) Gene amplification – selective DNA replication of
some genes to increase protein expression (ex.
after chemotherapy)
3) Transposons/
Retrotransposons(Jumping genes)
50% - Corn
10% Human
(Not inherited)
4)Gene rearrangement
•
1)
2)
3)
4)
Altering genomes during
your lifetime? continued
Rare mutations
Gene amplification temporary increase
(selective loss also
possible) in number of
gene copies
Transposons and
retrotransposons
Gene rearrangement in
Immunoglobin genes
Fig. 19.5
B lymphocytes (WBC) produce immunoglobins, or
antibodies, that specifically recognize and combat
viruses, bacteria, and other invaders.
-Millions of types of Antibodies can
be produced depending on what the
infectious agent is - how?
-Immunoglobins have constant and
variable region
-100s of gene segments code for the
variable region of the antibody.
-DNA segments are put together to
create an endless combination of
constant and variable regions - gene
rearrangement occurs in your
lifetime!
TRANSLATION
TRANSCRIPTION
Promotor
RNA Polymerase makes
premRNA using the
‘elves’ - transcription
factors (proteins)
Many protein ‘factors’ are involved in
translation as well
How is gene expression controlled?
•That is = if/what protein is made? How can you
control this?
Levels of control –goals…..
• 1) Changing DNA physically =>’ mRNA making’
affected
• 2) Changing access to DNA Promotor
• 3) If mRNA is made; How long mRNA hangs
around; change which protein is made from one
mRNA - (splicing); don’t use the mRNA
• 4) Change/destroy the protein after its made
How is gene expression controlled in
you? (IMPORTANT)
•When is the gene active (on or off)? That is what
protein is made? How can you control this?
• Gene expression control = which genes are “on”
• Levels of control –
• 1) chromatin (DNA) packing and chromatin modification change access sites on DNA for RNA Polymerase so that its
binding decreases/increases (epigenetics - layer of control
above the genome - NOVA Video)
• 2) Transcription - when DNA makes mRNA
• 3) Post-transcriptional - RNA processing, translation
• 4) Post-translational - various alterations to the protein product.
Fig. 19.7
1a) Level of packing is one way that gene
expression is regulated.
– Densely packed areas are inactivated.
(Heterochromatin)
- during mitosis
– Loosely packed areas are being actively
transcribed. (Euchromatin) -- during Interphase
Chromatin structure is based on
successive levels of DNA packing
INTERPHASE
• Interphase - chromatin fibers highly
extended
• Mitosis - chromatin coils and
condenses to form short, thick
chromosomes.
MITOSIS
Which stage
do you seeproteins
‘beads on are
a string’?
(Interphase)
• Histone
responsible
for the
Are genes first
active?
- Yes
transcribed
into mRNA!
level
of DNA
packaging.
• Their positively charged amino acids
bind tightly to negatively charged DNA.
Beads on a string = a
nucleosome, in
which DNA winds
around a core of
histone proteins
• Next level of packing - ‘30 nm solenoid fiber’ –
nucleosome fiber
• Has (DNA + HISTONES) with 6 nucleosomes
per turn
Which stage do you see ‘30 nm fiber’? (Mitosis)
Are genes active? - Yes transcribed into mRNA!
• The 30 nm fiber
forms looped
domains attached to
a scaffold of
nonhistone proteins.
Which stage do you see
‘looped domains’?
(Mitosis)
Are genes active? -No
1b) Chromatin modifications
(epigenetics)
• Chemical modifications of DNA bases:
• A) DNA methylation is the attachment by
specific enzymes of methyl groups (-CH3)
Inactive DNA is highly methylated compared to
DNA that is actively transcribed.
– Genomic imprinting is related to DNA methylation
DNA Methylation - add a methyl group to
make DNA less accessible
to RNA Polymerase
1b) Chromatin modifications
B) Histone acetylation (addition of an acetyl
group -COCH3) and deacetylation
Acetylated histones grip DNA less tightly = ?
More access to RNA Polymerase! SO,….
Epigenetics - DNA methylation and histone acetylation
may be responsible for a lot of traits that are not just
related to whether you have the gene/not. Example: If
your gene is methylated you may never express the trait!
2) Control of Transcription – very
important - to make or not make mRNA
Control elements - noncoding DNA segments that
regulate transcription by binding transcription
factors that are needed for RNA Polymerase
binding. (TATA Box -Promotor, Activators in
bacteria - ‘Enhancers’ in Eukaryotes, Repressors
in bacteria - ‘Silencers’ in Eukaryotes)
• How can a DNA control element 100s of basepairs
upstream of a gene regulate the access to RNA
Polymerase?
• Bending of DNA enables transcription factors,
activators (like steroid hormones), bound to
enhancers to contact the complex at the promoter.
Mostly positive gene
regulation in eukaryotes!
Fig. 19.9
• The hundreds of eukaryotic transcription
factors follow only a few basic structural
principles.
– Each protein generally has a DNA-binding
domain that binds to DNA and a proteinbinding domain that recognizes other
transcription factors.
Fig. 19.10
3) Post-transcriptional mechanisms - so
mRNA is made, what next?
• A) RNA processing – alternative splicing - controls
which protein is made from one mRNA - mix-n-match
introns/exons
3) Post-transcriptional
mechanisms
B) Life span of a mRNA molecule
Prokaryotic mRNA molecules degraded by
enzymes after only a few minutes.
Eukaryotic mRNAs endure typically for hours or
even days or weeks.
G L
5’ Cap Leader
T
Trailer
AAAAA
Poly A tail
3) Post-transcriptional
mechanisms
C) Translation - can be blocked by regulatory
proteins that bind to 5’ leader region of mRNA.
(prevents attachment of mRNA to ribosomes)
Protein factors required to initiate translation =
simultaneous control of translation of all the mRNA
in a cell.
G L
5’ Cap Leader
T
Trailer
AAAAA
Poly A tail
4) Post-translational mechanisms
• Processing of polypeptides to yield functional
proteins.
– This may include cleavage, chemical modifications,
and transport to the appropriate destination.
• Regulation may occur at any of these steps.
• The cell limits the lifetimes of normal proteins
by selective degradation.
• Proteins intended for degradation are marked
by the attachment of ubiquitin proteins.
• Giant proteosomes recognize the ubiquitin and
degrade the tagged protein.
Fig. 19.12
CANCER REVIEW- read on
your own - use these animations
Cancer results from genetic
changes that affect the cell cycle
• Cell cycle CONTROL events don’t work
• Spontaneous mutations or environmental
influences (carcinogens)
• Cancer-causing genes – oncogenes
(retroviruses), proto-oncogenes (in other
organisms).
• What happens when protooncogenes/oncogenes are turned ‘ON’?
(Ras gene)
• Cell will divide without stopping
• Malignant cells often have significant
changes in chromosomes
Fig. 19.13
Are there genes that prevent
cancer?
• Tumor-suppressor genes -normal
products inhibit cell division, repair DNA,
control adhesion (p53).
• Mutations to these tumor suppressor genes
= cancer
Oncogene proteins and
faulty tumor-suppressor
proteins
• The p53 gene, named for its 53,000-dalton
protein product, is often called the
“guardian angel of the genome”.
• Damage to the cell’s DNA acts as a signal
that leads to expression of the p53 gene.
• The p53 protein is a transcription factor for
several genes.
– It can activate the p21 gene, which halts the
cell cycle.
– It can turn on genes involved in DNA repair.
– When DNA damage is irreparable, the p53
protein can activate “suicide genes” whose
protein products cause cell death by apoptosis.
3. Multiple mutations underlie
the development of cancer
• More than one somatic mutation is generally
needed to produce the changes
characteristic of a full-fledged cancer cell.
• If cancer results from an accumulation of
mutations, and if mutations occur throughout
life, then the longer we live, the more likely
we are to develop cancer.
• Colorectal cancer, with 135,000 new cases
in the U.S. each year, illustrates a multistep cancer path.
• The first sign is often a polyp, a small
benign growth in the colon lining with fast
dividing cells.
• Through gradual accumulation of mutations
that activate oncogenes and knock out
tumor-suppressor genes, the polyp can
develop into a malignant tumor.
Fig. 19.15
• About a half dozen DNA changes must occur
for a cell to become fully cancerous.
• These usually include the appearance of at
least one active oncogene and the mutation or
loss of several tumor-suppressor genes.
– Since mutant tumor-suppressor alleles are usually
recessive, mutations must knock out both alleles.
– Most oncogenes behave as dominant alleles.
• In many malignant tumors, the gene for
telomerase is activated, removing a natural limit
on the number of times the cell can divide.
• Viruses, especially retroviruses, play a role
is about 15% of human cancer cases
worldwide.
– These include some types of leukemia, liver
cancer, and cancer of the cervix.
• Viruses promote cancer development by
integrating their DNA into that of infected
cells.
• By this process, a retrovirus may donate an
oncogene to the cell.
• Alternatively, insertion of viral DNA may
disrupt a tumor-suppressor gene or convert
a proto-oncogene to an oncogene.
• The fact that multiple genetic changes are
required to produce a cancer cell helps
explain the predispositions to cancer that
run in some families.
– An individual inheriting an oncogene or a
mutant allele of a tumor-suppressor gene will
be one step closer to accumulating the
necessary mutations for cancer to develop.
• Geneticists are devoting much effort to
finding inherited cancer alleles so that
predisposition to certain cancers can be
detected early in life.
– About 15% of colorectal cancers involve
inherited mutations, especially to DNA repair
genes or to the tumor-suppressor gene APC.
• Normal functions of the APC gene include regulation
of cell migration and adhesion.
– Between 5-10% of breast cancer cases, the 2nd
most common U.S. cancer, show an inherited
predisposition.
• Mutations to one of two tumor-suppressor genes,
BRCA1 and BRCA2, increases the risk of breast
and ovarian cancer.