©2001 Timothy G. Standish Initiation of Initiation of Transcript Transcript ion: ion: In Eukaryotes In Eukaryotes Timothy G. Standish, Ph. D.
©2001 Timothy G. Standish
Initiation ofInitiation of Transcription:Transcription:
In EukaryotesIn EukaryotesTimothy G. Standish, Ph. D.
©2001 Timothy G. Standish
Expression Control In EukaryotesExpression Control 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
©2001 Timothy G. Standish
Eukaryotes Have Large Eukaryotes Have Large Complex GeneomesComplex Geneomes
The human genome is about 3 x 109 base pairs or ≈ 1 m of DNA
Because humans are diploid, each nucleus contains 6 x 109 base pairs or ≈ 2 m of DNA
Some gene families are located close to one another on the same chromosome
Genes with related functions appear to be distributed almost at random throughout the the genome
©2001 Timothy G. Standish
Highly Packaged DNA Cannot Highly Packaged DNA Cannot be Expressedbe Expressed
Because of its size, eukaryotic DNA must be packaged
Heterochromatin, the most highly packaged form of DNA, cannot be transcribed, therefore expression of genes is prevented
Chromosome puffs on some insect chomosomes illustrate areas of active gene expression
©2001 Timothy G. Standish
Only a Subset of Genes is Only a Subset of Genes is Expressed at any Given TimeExpressed 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.
©2001 Timothy G. Standish
DNA
Cytoplasm
Nucleus
G AAAAAA
Export
Degradation etc.G AAAAAA
Control of Gene ExpressionControl of Gene Expression
G AAAAAA
RNAProcessing
mRNA
RNA
Transcription
Nuclear pores
Ribosom
e
Translation
Packaging
Modification
Transportation
Degradation
©2001 Timothy G. Standish
Logical Expression Control PointsLogical Expression Control Points DNA packaging Transcription RNA processing mRNA Export mRNA masking/unmasking and/or
modification mRNA degradation Translation Protein modification Protein transport Protein degradation
Increasing costIncreasing cost
The logical place to control
expression is before the
gene is transcribed
The logical place to control
expression is before the
gene is transcribed
©2001 Timothy G. Standish
Three Eukaryotic Three Eukaryotic RNA PolymerasesRNA Polymerases
1 RNA Polymerase I - Produces rRNA in the nucleolus, accounts for 50 - 70 % of transcription
2 RNA Polymerase II - Produces mRNA in the nucleoplasm - 20 - 40 % of transcription
3 RNA Polymerase III - Produces tRNA in the nucleoplasm - 10 % of transcription
©2001 Timothy G. Standish
A “Simple” Eukaryotic GeneA “Simple” Eukaryotic Gene
Terminator Sequence
Promoter/Control Region
Transcription Start Site 5’ Untranslated Region 3’ Untranslated Region
Exons
Introns
3’5’ Exon 2 Exon 3Int. 2Exon 1 Int. 1
RNA Transcript
©2001 Timothy G. Standish
5’DNA
3’
EnhancersEnhancers
Enhancer Transcribed Region
3’5’ TF TFTF
3’5’ TF TFTF
5’RNA
RNAPol.
RNAPol.
Many bases
Promoter
©2001 Timothy G. Standish
Eukaryotic RNA Polymerase IIEukaryotic 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:
©2001 Timothy G. Standish
Eukaryotic RNA Polymerase II Eukaryotic RNA Polymerase II PromotersPromoters
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
©2001 Timothy G. Standish
RNA Pol. II
InitiationInitiation
T. F.
RNA Pol. II
5’mRNA
Promoter
T. F.
T. F.
©2001 Timothy G. Standish
Eukaryotic PromotersEukaryotic Promoters
5’ Exon 1Promoter
Sequence elements
~200 bp
TATA
~-25
InitiatorInitiator“TATA Box”“TATA Box”
Transcription start site
(Template strand)-1+1
SSTATAAAASSSSSNNNNNNNNNNNNNNNNNYYCAYYYYYNN
S = C or G Y = C or T N = A, T, G or C
©2001 Timothy G. Standish
InitiationInitiationTFIID BindingTFIID Binding
-1+1
Transcription start site
TFIID
“TATA Box”
TBP Associated Factors (TAFs)
TATA Binding Protein (TBP)
©2001 Timothy G. Standish
InitiationInitiationTFIID BindingTFIID Binding
TFIID
80o Bend
-1+1
Transcription start site
©2001 Timothy G. Standish
InitiationInitiationTFIIA and B BindingTFIIA and B Binding
TFIID
TFIIA
-1+1
Transcription start site
TFIIB
©2001 Timothy G. Standish
InitiationInitiationTFIIF and RNA Polymerase BindingTFIIF and RNA Polymerase Binding
TFIID
TFIIA
-1+1
Transcription start site
TFIIB
RNA PolymeraseTFIIF
©2001 Timothy G. Standish
InitiationInitiationTFIIE BindingTFIIE Binding
TFIID
TFIIA
-1+1
Transcription start site
RNA PolymeraseTFIIBTFIIF
TF
IIE
TFIIE has some helicase activity and may by involved in unwinding DNA so that transcription can start
©2001 Timothy G. Standish
InitiationInitiationTFIIH and TFIIJ BindingTFIIH and TFIIJ Binding
TFIID
TFIIA
-1+1
Transcription start site
RNA PolymeraseTFIIBTFIIF
TF
IIE
TFIIH has some helicase activity and may by involved in unwinding DNA so that transcription can start
TFIIH
P PP
TFIIJ
©2001 Timothy G. Standish
InitiationInitiationTFIIH and TFIIJ BindingTFIIH and TFIIJ Binding
TFIID
TFIIA
-1+1
Transcription start site
RNA PolymeraseTFIIBTFIIF
TF
IIETFIIH
P PP
TFIIJ
©2001 Timothy G. Standish
InitiationInitiationTFIIH and TFIIJ BindingTFIIH and TFIIJ Binding
-1+1
Transcription start site
RNA PolymeraseP P
P
©2001 Timothy G. Standish