Regulation of gene expression in eukaryotes

I. Overview of Eukaryotic gene
regulation
• Mechanisms similar to
those found in bacteriamost
genes controlled
at the transcriptional
level
• Much more complex
than prokaryotic
– chromatin
– TFs
– Enhancers
– Activators
A. Prokaryotes vs. Eukaryotes
In eukaryotes,
(in bacteria, one mRNA can be polycistronic, or code for
several proteins).
DNA in eukaryotes forms a stable, compacted
complex with histones. In bacteria, the chromatin is
not in a permanently condensed state.
Eukaryotic DNA contains large regions of
Eukaryotic genes are divided into exons and introns;
in bacteria, genes are almost never divided.
In eukaryotes, mRNA is synthesized in the nucleus
and then processed and exported to the cytoplasm; in
bacteria, transcription and translation can take place
simultaneously off the same piece of DNA.
B. Eukaryote gene expression
is regulated at 6 levels:
II. Transcriptional Control
A. Control factors
1) cis-acting “next to” elements
• Promoter region: TATA box (-30), CAAT box
(-80) GC box (-110)
• Alternate promoters
– The level of transcription initiation can vary between
alternative promoters
– the translation efficiency of mRNA isoforms with
different leader exons can differ
– alternative promoters can have different tissue
specificity and react differently to some signals
• Enhancers & Silencers far away from promoter
2) trans-acting “across from” factors
• Transcription factors
• Activators, Coactivators
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Control factors continued:
3) DNA methylation (add methyl to C)
• Occurs at 5’ position, usually in CG doublets
• 5’- m CpG-3’
– Transcriptionally active genes possess
significantly lower levels of methylated DNA
than inactive genes.
• A gene for methylation is essential for development in
mice (turning off a gene also can be important).
• Methylation results in a human disease called fragile
X syndrome; FMR-1 gene is silenced by methylation.
Control factors continued:
4) Chromatin conformation (remodelling)
a. Antirepressors & nucleosome positioning.
b. Histone acetylation – (acetyl groups on lysines),
histone acetyltransferase enzyme catalyzes the
addition of lysine, targeted to genes by specific
TFs.
c. Heterochromatin – highly condensed,
transcriptionally inert (off).
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B. Eukaryotic Promoters
Usually located within 100 bp upstream
‣ Recognized byRNA Pol II (transcribes mRNA)
‣ Require the binding of several protein factors to initiate
transcription (DNA binding domains on TFs – ‘motifs’)
‣ May be positively or negatively regulated
C. Transcription Factors –the
transcription complex
1) TFIIA, TFIIB,
TFIID, TFIIE,
TFIIH
2) TATA binding
protein (TBP)
3) TBP associated
factors (TAFs)
‣Assembly of the basal
transcription apparatus -
involves stepwise binding
of various transcription
factor proteins.
Commitment Stage &
Clearance Stage…
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Enhancers
Cis regulators that interact
with regulatory proteins &
TFs to increase the
efficiency of transcription
initiation.
Silencers – cis-acting,
bound by repressors, or
cause the chromatin to
condense and become
inactive.
D. An example of transcriptional control:
Galactose metabolism in yeast
‣ GAL1, GAL7, GAL10 genes… products
required for conversion of galactose into
glucose
Activators - Proteins that
function by contacting basal
transcription factors and
stimulating the assembly of
pre-initiation complexes at
promoters.
Galactose metabolizing
pathway of yeast.
Controlling GAL
GAL80 encodes a protein that negatively regulates
transcription. The repressor protein binds to an
Activator protein, rendering it inactive.
GAL4 encodes an activator w/zinc finger motif that
activates transcription of the three GAL genes
individually.
Galactose = Inducer, that binds to Gal80, causing
it to release Gal4
‣ Although this looks similar to Lac Operon, there
are different molecular mechanisms…
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Activation model of GAL genes in yeast.
III. Post-transcriptional control
Alternative polyadenylation and splicing of the human CACL gene in
thyroid and neuronal cells.
A. Alternative splicing - Some messages undergo
alternate splicing depending on what tissue they
are located in. The regulation is at the level of
snRNP production.
– Some pre-mRNAs can be
– Controlled by RNA binding splicing factors that commit
splicing in a particular way
Calcitonin
gene-related
peptide
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Post-transcriptional control cont.
B. The stability of a class of mRNA can be controlled.
‣ Some short-lived mRNAs have multiple copies of the
sequence AUUUA which may act as a target for
degradation.
‣ the hormone prolactin enhances the stability of the
mRNA for the milk protein casein
‣ high levels of iron decrease the stability of the mRNA
for the receptor that brings iron into cells
C. RNA interference – poorly understood, but
appears to be widespread in fungi, plants and
animals as a regulatory mechanism
‣ miRNAs & siRNAS (small RNA molecules) pair with
proteins to form an RNA-induced silencing complex
(RISC)
‣ RISC pairs w/complentary base sequences of specific
mRNAs and causes:
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