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Conclusions (last lecture)
• Conserved structure of RNA pol II
• RNA pol II must navigate a chromatin
template in a highly dynamic
environment
Outline
• Mechanism of Transcription Activation
– RNA pol II structure and function
• Coordination of RNA processing
– Transcription Regulatory elements
• Identification & characterization in vivo
• Examples of mammalian transcriptional control regions
– Transcription Factors
Central
Dogma
Why aren’t tRNAs and
rRNAs processed like
mRNAs?
What is different about
how mRNAs are
made?
Fig. 1-9 Lodish 2013
Only RNA Polymerase II has a C-Terminal Domain
CTD is Phosphorylated Prior to Transcription Initiation
CTD
Red arrow indicates location where unique C-terminal
domain extends from in RNA pol II
Fig. 7-10. Lodish 2013
The RNA Pol II C-Terminal Domain (CTD) facilitates
capping, splicing and polyadenylation by recruitment
of RNA processing factors.
Hsin J , and Manley J L Genes Dev.
2012;26:2119-2137
©2012 by Cold Spring Harbor Laboratory Press
RNA Pol II CTD phosphorylation coordinates the transcription cycle
with pre-mRNA processing and histone modification.
5’ Capping
Splicing
3’ Processing
Hirose Y , Ohkuma Y J Biochem 2007;141:601-608
© 2007 The Japanese Biochemical Society.
Drosophila
chromosome
stained with
antibody to
phosphorylated
Pol II (Red) or
unphosphorylated
Pol II (Green)
What do you
notice?
Fig. 7-13 Lodish 2013
Central
Dogma
Fig. 1-9 Lodish 2013
Alternative Splicing
Simple
Complex
Which promoter is regulating mRNA2 transcription?
Pax6: Master regulator of eye development
Ectopic Expression of Eyeless (PAX 6 homolog)
in Drosophila results in aberrant formation of eye structures
Antenna
Lower leg
Halder et al. Science 267: 1788 (1995)
Different Pax 6 enhancers are active in different tissues:
Results in different sites of transcription initiation and
alternative splicing in different tissues
Transcript a is detected in lense
and pancreas. Lacks exons 1 and α
Fig. 7-6 Lodish et al. 2008
How can we study gene
expression in vivo?
One method: transgenic mouse
Introduce “reporter gene” controlled by
potential regulatory elements into a
mouse
transgenic mouse animation
Distinct from reporter gene assay which
is performed in cell culture
Answer Pax6 gene questions
in your course manual
Alternative Pax 6 promoters & enhancers are used to generate
different transcripts in different tissues
Transcript c:
detected in retina
lacks exons 0-4
contains exon α
Fig. 7-6 Lodish et al. 2008
Conclusions
• Unique Pol II CTD provides a mechanism for
coupling mRNA-specific processing with
transcription
• Different enhancers/promoters can control
transcription of the same gene in different
cell types
• Different subsets of transcription factors bind
to enhancers of the same gene in different
cell types
• Enhancers can be located far from
transcription start sites