Download Eukaryotic Gene Expression Heyer 1

Eukaryotic Gene Expression
Eukaryotic Genomes & Gene Expression
Eukaryotic Cells Have Multiple Chromosomes
•  Eukaryotic cells have 5–20x more
DNA per cell than do bacteria.
•  Divided into linear dsDNA +
proteins : chromosomes
•  Typical chromosome averages
~1.5x108 nucleotide pairs.
–  If straight strands would be ~ 4 cm
long.
•  Humans have 46 different
chromosomes
–  So collectively amounts to ~ 2 m long
dsDNA packed into each cell nucleus!
(4 m after replication!)
Plant cell just before division
Stages in gene expression that can be
regulated in eukaryotic cells
Chromatin: nondividing cells
Signal
DNA
RNA
NUCLEUS
Chromatin
Chromatin modification:
DNA unpacking involving
histone acetylation and
DNA demethlation
Gene available
for transcription
Gene
Transcription
Exon
Primary transcript
Intron
RNA processing
Tail
Cap
•  DNA is coiled to pack
into nucleus
mRNA in nucleus
Transport to cytoplasm
CYTOPLASM
mRNA in cytoplasm
Degradation
of mRNA
•  Different degrees of
coiling and packing
Translation
Polypetide
Cleavage
Chemical modification
Transport to cellular
destination
Active protein
Degradation of protein
Degraded protein
Figure 18.6
Levels of chromatin packing
Histone tails and the effect of acetylation
1.  Transcription factors may
catalyze histone acetylation
2.  Acetylated histone tails may
recruit transcription factors
2 nm
DNA double helix
Histones
Histone
tails
Histone H1
Beads of eight histone proteins
10 nm
DNA double helix
Linker DNA
(“string”)
Nucleosome
(“bead”)
chromatin
(a) Nucleosomes (10-nm fiber)
30 nm
Amino acids on tails available
for chemical modification
(a) Histone tails protrude outward from a nucleosome
Dense regions of
heterochromatin
Nucleosome
Unacetylated histones
DNA remains coiled around histone “beads” except during replication.
Figure 18.7
Heyer
Histone tails
Less dense regions of
euchromatin
Acetylated histones
(b) Acetylation of histone tails promotes loose chromatin structure
that permits transcription
Figure 18.7
1
Eukaryotic Gene Expression
DNA in a eukaryotic chromosome from a
developing salamander egg
Chromosomes in the
interphase nucleus
Chromosome
territory
10 µm
Chromatin
loop
Transcription
factory
Figure 18.12
More loosely coiled loops are available for transcription and gene expression.
During cell division
Levels of chromatin packing
30 nm
Nucleosome
chromatin
(b) 30-nm fiber
Protein scaffold
Loops
300 nm
Scaffold
(c) Looped domains (300-nm fiber)
•  chromatin condenses into visible chromosomes
700 nm
chromosome
•  Tightly wound chromosomes segregate without
tangling together
1,400 nm
(d) Metaphase chromosome
Cell Differentiation in Multicellular Organisms
Differential Gene Expression
•  Even though every cell has the same genome,
each cell type only uses a small subset of genes.
– 
– 
– 
– 
~200 cell types in mammals
Each uses only ~20% of total genes
Fewer in more specialized cells
Unused genes may be permanently inactivated
Processing of “primary transcript” RNA
[Review Gene Expression slides!]
•  Histone modification
5ʹ′
–  Methylation of histone residues may condense
associated DNA into non-transcribable heterochromatin
•  DNA methylation
–  Methylation of cytosines related to gene inactivation
–  Methylated DNA may attract/bind histone deacetylation
enzymes
–  Epigenic inheritance — patterns of methylation passed
on to daughter cells
Figure 47.7 Blastulation
Heyer
50 to 250 adenine nucleotides
added to the 3ʹ′ end by
A modified methyl-guanine
nucleotide added to the 5ʹ′ end
poly-A synthetase using ATPs
Protein-coding segment
G P P P
5ʹ′ Cap
Polyadenylation signal
AAUAAA
5ʹ′ UTR
Start
codon
Stop
codon
3ʹ′ UTR
3ʹ′
AAA…AAA
Poly-A tail
• Cap & tail protect mRNA from rapid degradation in the cytoplasm.
• Eukaryotic mRNA stay active for hours, or even days, in the cytoplasm.
• Prokaryotes lack cap & tail; mRNA only lasts for minutes.
Figure 17.9
2
Eukaryotic Gene Expression
A eukaryotic gene and its transcript
Enhancer
Proximal
(distal control elements) control elements
A model for the action of enhancers and
transcription activators
Distal control
element
Poly-A signal Termination
sequence
region
Exon
Intron
Downstream
Poly-A
signal
Cleared 3ʹ′ end
Intron Exon
of primary
RNA processing:
transport
Cap and tail added;
introns excised and
exons spliced together
Transcription
Promoter
Primary RNA
transcript 5ʹ′
(pre-mRNA)
Chromatin changes
Exon
Intron Exon
Transcription
Intron RNA
RNA processing
mRNA
degradation
TATA
box
1 Activator proteins bind
to distal control elements
grouped as an enhancer in
the DNA. This enhancer has
three binding sites.
General
transcription
factors
DNA-bending
protein
2 A DNA-bending protein
brings the bound activators
closer to the promoter.
Other transcription factors,
mediator proteins, and RNA
polymerase are nearby.
Coding segment
Translation
Protein processing
and degradation
Gene
Enhancer
DNA
Upstream
Promoter
Activators
Intron Exon
Exon
Group of
Mediator proteins
RNA
Polymerase II
mRNA G P P P
5ʹ′ Cap 5ʹ′ UTR
(untranslated
region)
Figure 18.8
Start
codon
Stop
codon
3ʹ′ UTR Poly-A tail
(untranslated
region)
•  Enhancer sequences may be several kb upstream or downstream of the gene,
or within an intron.
•  One gene may have several enhancers.
Chromatin changes
3 The activators bind to
certain general transcription
factors and mediator
proteins, helping them form
an active transcription
initiation complex on the promoter.
Transcription
RNA processing
mRNA
degradation
RNA
Polymerase II
Translation
Protein processing
and degradation
Transcription
Initiation complex
RNA synthesis
Figure 18.10
Cell type–specific transcription
An Activator Protein and DNA-binding
Promoter
Enhancer
Albumin
gene
Control
elements
Crystallin
gene
Lens cell
nucleus
Liver cell
nucleus
Activation
domain
DNA-binding
domain
Available
activators
Available
activators
DNA
Albumin
gene not
expressed
Albumin
gene
expressed
Crystallin gene
Crystallin gene
not expressed
expressed
(a) Liver cell
(b) Lens cell
Figure 18.9
Sequential Regulation of Gene Expression During Cellular Differentiation
Nucleus
Embryonic
precursor cell
Master regulatory
gene myoD
Other muscle-specific genes
Alternative RNA splicing
•  Esp. in vertebrate animals
•  >1 possible polypeptide/gene!
DNA
OFF
OFF
Figure 18.11
Chromatin changes
Transcription
RNA processing
OFF
mRNA
Myoblast
(determined)
mRNA
degradation
MyoD protein
(transcription
factor)
Translation
Protein processing
and degradation
Exons
DNA
mRNA
Part of a muscle fiber
(fully differentiated cell)
Heyer
MyoD
mRNA
Another
transcription
factor
mRNA
mRNA
Myosin, other
muscle proteins,
and cell cycle–
blocking proteins
Figure 18.18
Primary
RNA
transcript
RNA splicing
or
mRNA
Figure 18.13
3
Eukaryotic Gene Expression
Regulation of gene expression by microRNAs
(miRNAs)
1 The microRNA (miRNA)
precursor folds
back on itself,
held together
by hydrogen
bonds.
2 An enzyme
called Dicer moves
along the doublestranded RNA,
cutting it into
shorter segments.
3 One strand of
each short doublestranded RNA is
degraded; the other
strand (miRNA) then
associates with a
complex of proteins.
4 The bound
miRNA can base-pair
with any target
mRNA that contains
the complementary
sequence.
Degradation of a protein by a proteasome
5 The miRNA-protein
complex prevents gene
expression either by
degrading the target
mRNA or by blocking
its translation.
Chromatin changes
Transcription
Chromatin changes
1 Multiple ubiquitin molecules are attached to a protein
by enzymes in the cytosol.
2 The ubiquitin-tagged protein
is recognized by a proteasome,
which unfolds the protein and
sequesters it within a central cavity.
2 Enzymatic components of the
proteasome cut the protein into
small peptides, which can be
further degraded by other
enzymes in the cytosol.
Transcription
RNA processing
RNA processing
mRNA
degradation
Translation
Protein processing
and degradation
Protein
complex
mRNA
degradation
Proteasome
and ubiquitin
to be recycled
Ubiquitin
Translation
Proteasome
Protein processing
and degradation
Dicer
Degradation of mRNA
OR
miRNA
Protein to
be degraded
Ubiquinated
protein
Protein entering a
proteasome
Target mRNA
Protein
fragments
(peptides)
Blockage of translation
Hydrogen
bond
Figure 18.14
See Figure 18.15
Movement of eukaryotic transposable elements
The effect of transposable elements
on corn kernel color
New copy of
transposon
Transposon
DNA of genome
Transposon
is copied
Insertion
Mobile transposon
(a) Transposon movement (“copy-and-paste” mechanism)
New copy of
Retrotransposon
retrotransposon
Figure 21.9
DNA of genome
RNA
Reverse
transcriptase
Insertion
(b) Retrotransposon movement
Figure 21.10
Types of DNA sequences in the human genome
Exons (1.5%)
Alu elements
(10%)
Encode Project: Rethinking ‘Junk’ DNA
Introns (5%)
GENES Each human cell contains about 10 feet of DNA, coiled into a dense tangle. But only a very small percentage of DNA encodes genes, which control inherited traits like eye color, blood type and so on Unique
noncoding
DNA (15%)
Repetitive
DNA
unrelated to
transposable
elements
(14%)
Simple sequence
DNA (3%)
Heyer
Figure 21.8
Regulatory
sequences
(∼20%)
Repetitive
DNA that
includes
transposable
elements
and related
sequences
(44%)
L1
sequences
(17%)
Barbara McClintock’s breeding experiments with Indian corn
Large-segment
duplications (5-6%)
Figure 21.7
“JUNK” DNA [“DARK MATTER”] Stretches of DNA around and between genes seemed to do nothing, and were called junk DNA. But now researchers think that the junk DNA contains a large number of *ny gene*c switches, controlling how genes func*on within the cell. Over half of the
human genome is
gene switches.
REGULATION DISEASE The many gene*c regulators seem to be arranged in a complex and redundant hierarchy. Scien*sts are only beginning to map and understand this network, which regulates how cells, organs and *ssues behave. Errors or muta*ons in gene*c switches can disrupt the network and lead to a range of diseases. The new findings will spur further research and may lead to new drugs and treatments. Bits of Mystery DNA, Far From ‘Junk,’ Play Crucial Role
By GINA KOLATA; September 5, 2012, NY Times
National Human Genome Research Institute
Encyclopedia Of DNA Elements
4