Transcript Foundations of Biology

Hebrews 1:1-2
1 God, who at sundry times and in
divers manners spake in time
past unto the fathers by the
prophets,
2 Hath in these last days spoken
unto us by his Son, whom he
hath appointed heir of all things,
by whom also he made the
worlds;
©2000 Timothy G. Standish
Gene Regulation Eukaryotes
Timothy G. Standish, Ph. D.
©2000 Timothy G. Standish
Control Of Expression In
Eukaryotes
Some of the general methods used to control
expression in prokaryotes are used in
eukaryotes, but nothing resembling operons
is known
Eukaryotic genes are controlled individually
and each gene has specific control sequences
preceding the transcription start site
In addition to controlling transcription, there
are additional ways in which expression can
be controlled in eukaryotes
©2000 Timothy G. Standish
Eukaryotes Have Large
Complex Genomes
The human genome is about 3 x 109
base pairs or ≈ 1 m of DNA
Because humans are diploid, each
nucleus contains 6 3 x 109 base pairs
or ≈ 2 m of DNA
That is a lot to pack into a little
nucleus!
©2000 Timothy G. Standish
Only a Subset of Genes is
Expressed at any Given Time
It takes lots of energy to express genes
Thus it would be wasteful to express all
genes all the time
By differential expression of genes, cells
can respond to changes in the environment
Differential expression, allows cells to
specialize in multicelled organisms.
Differential expression also allows
organisms to develop over time.
©2000 Timothy G. Standish
Control of Gene Expression
Cytoplasm
Packaging
Degradation
DNA
Transcription
Transportation
Modification
RNA
RNA
Processing
mRNA G
G
AAAAAA
Nucleus
Export
Degradation etc.
AAAAAA
Translation
©2000 Timothy G. Standish
Logical Expression Control Points
Increasing cost
DNA packaging
Transcription
RNA processing
mRNA export
mRNA masking/unmasking
and/or modification
mRNA degradation
Translation
Protein modification
Protein transport
Protein degradation
The logical
place to
control
expression is
before the
gene is
transcribed
©2000 Timothy G. Standish
Eukaryotic DNA Must be
Packaged
Eukaryotic DNA exhibits many
levels of packaging
The fundamental unit is the
nucleosome, DNA wound around
histone proteins
Nucleosomes arrange themselves
together to form higher and higher
levels of packaging.
©2000 Timothy G. Standish
Packaging DNA
Nucleosomes
Metaphase
Chromosome
Tight helical fiber
B DNA Helix
Looped Domains
Protein scaffold
©2000 Timothy G. Standish
Highly Packaged DNA Cannot
be Expressed
The most highly packaged form of
DNA is “heterochromatin”
Heterochromatin cannot be transcribed,
therefore expression of genes is
prevented
Chromosome puffs on some insect
chomosomes illustrate where active
gene expression is going on
©2000 Timothy G. Standish
Eukaryotic RNA Polymerase II
RNA polymerase is a very fancy
enzyme that does many tasks in
conjunction with other proteins
RNA polymerase II is a protein
complex of over 500 kD with
more than 10 subunits:
©2000 Timothy G. Standish
Eukaryotic RNA Polymerase II
Promoters
Several sequence elements spread over about
200 bp upstream from the transcription start
site make up RNA Pol II promoters
Enhancers, in addition to promoters,
influence the expression of genes
Eukaryotic expression control involves many
more factors than control in prokaryotes
This allows much finer control of gene
expression
©2000 Timothy G. Standish
Initiation
T. F.
Promoter
T. F.
RNA
Pol. II
RNA
Pol. II
mRNA
5’
©2000 Timothy G. Standish
Eukaryotic RNA Polymerase II
Promoters
Eukaryotic promoters are made up of a number
of sequence elements spread over about 200 bp
upstream from the transcription start site
In addition to promoters, enhancers also
influence the expression of genes
Control of gene expression in eukaryotes
involves many more factors than control in
prokaryotes
This allows much finer control of gene
expression
©2000 Timothy G. Standish
A “Simple” Eukaryotic Gene
Transcription
Start Site
5’
5’ Untranslated Region
Introns
Exon 1 Int. 1
Promoter/
Control Region
3’ Untranslated Region
Exon 2
3’
Int. 2 Exon 3
Exons
Terminator
Sequence
RNA Transcript
©2000 Timothy G. Standish
Eukaryotic Promoters
Promoter
5’
Exon 1
Sequence elements
TATA
~200 bp
“TATA Box”
Initiator
Transcription
start site
SSTATAAAASSSSSNNNNNNNNNNNNNNNNNYYCAYYYYYNN
(Template strand)
~-25
-1+1
S = C or G
Y = C or T
N = A, T, G or C
©2000 Timothy G. Standish
Response Elements
Response elements are short sequences
found either within about 200 bp of the
transcription start site, or as part of
enhancers
Different genes have different response
elements
Binding of transcription factors to response
elements determines which genes will be
expressed in any cell type under any set of
conditions
©2000 Timothy G. Standish
Initiation
TFIID Binding
TFIID
“TATA Box”
Transcription
start site
TBP Associated
Factors (TAFs)
-1+1
TATA Binding
Protein (TBP)
©2000 Timothy G. Standish
Initiation
TFIID Binding
Transcription
start site
TFIID
-1+1
80o Bend
©2000 Timothy G. Standish
Initiation
TFIIA and B Binding
TFIID
TFIIB
Transcription
start site
-1+1
TFIIA
©2000 Timothy G. Standish
Initiation
TFIIF and RNA Polymerase Binding
TFIID
TFIIB
Transcription
start site
-1+1
TFIIA
TFIIF
RNA Polymerase
©2000 Timothy G. Standish
Initiation
TFIIE Binding
TFIIF TFIIB
RNA Polymerase
-1+1
TFIIA
TFIIE
TFIID
Transcription
start site
TFIIE has some
helicase activity and
may be involved in
unwinding DNA so
that transcription can
start
©2000 Timothy G. Standish
Initiation
TFIIH and TFIIJ Binding
TFIIJ
TFIIH
TFIIF TFIIB
P
TFIIA
PP
RNA Polymerase
-1+1
TFIIE
TFIID
Transcription
start site
TFIIH has some
helicase activity and
may be involved in
unwinding DNA so
that transcription can
start
©2000 Timothy G. Standish
Initiation
TFIIH and TFIIJ Binding
TFIIJ
TFIIH
TFIIF TFIIB
P
PP
-1+1
TFIIE
TFIID
Transcription
start site
RNA Polymerase
TFIIA
©2000 Timothy G. Standish
Initiation
TFIIH and TFIIJ Binding
Transcription
start site
P
-1+1
PP
RNA Polymerase
©2000 Timothy G. Standish
Enhancers
DNA
Many bases
5’
3’
Enhancer
5’
Promoter
TF
Transcribed Region
3’
TF
5’
TF TF RNA
RNA
Pol.
Pol.
5’
3’
RNA
©2000 Timothy G. Standish
©2000 Timothy G. Standish