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Transcription
(CHAPTER 12- Brooker Text)
Sept 18 & 23, 2008
BIO 184
Dr. Tom Peavy
• Bacterial mRNA may be polycistronic, which
means it encodes two or more polypeptides
The Stages of Transcription
• Transcription occurs in three stages
– Initiation
– Elongation
– Termination
• These steps involve protein-DNA interactions
– Proteins such as RNA polymerase interact with DNA
sequences
Initiation
• promoter = recognition site for transcription factors
• transcription factors + RNA polymerase to bind to the promoter
(initial phase) = closed promoter complex
• then DNA is denatured (bubble) = open promoter complex
Prokaryotic Transcription
• E. coli RNA polymerase = holoenzyme
– Core enzyme (Four subunits = a2bb’)
– Sigma factor (One subunit = s)
Initiation stages involve RNA pol holoenzyme
• Binding loosely to the DNA
• Scaning for promoter region
• Forming Open promoter complex
• Synthesizing Short stretch of RNA
• Releasing Sigma factor
Bacterial Promoter
Sequence elements that
play a key role in
transcription
(pribnow box)
• The RNA transcript is synthesized during ELONGATION step
• The DNA strand used as a template for RNA synthesis is
termed the template or noncoding strand
• The opposite DNA strand is called the coding strand
– It has the same base sequence as the RNA transcript
• Except that T in DNA corresponds to U in RNA
Termination of Bacterial
Transcription
• short RNA-DNA hybrid is forced to separate
= release of newly made RNA
• E. coli has two different mechanisms for termination
– 1. rho-dependent termination
• Requires a protein known as r (rho)
– 2. rho-independent termination
• Does not require r but uses stem loop structure and U–rich
sequence region
rho utilization site
Rho protein is
a helicase
Figure 12.8
r-dependent termination
Figure 12.8
r-dependent termination
Types of RNA
mRNA = messenger RNA encodes for the sequence of
amino acids within a polypeptide
tRNA = transfer RNA carries amino acids at the amino
acyl terminus for incorporation into growing
polpeptides during translation at ribosome
rRNA = ribosomal RNA is a major component of the
ribosomal complex essential for coordinating
protein sythesis using the mRNA as a template
(translation)
TRANSCRIPTION IN EUKARYOTES
• Similar but more complex
– Larger organisms
– Cellular complexity
– Multicellularity
•
Three Different RNA polymerase needed
1) RNA pol I = rRNA genes (except 5S)
2) RNA pol II = mRNAs (structural genes)
3) RNA pol III = tRNAs and 5S rRNA
Figure 12.11
• The core promoter is relatively short = TATA box
- determines start point for transcription
- basal transcription by itself (low level)
• Regulatory Elements (e.g. GC and CAAT boxes)
- Enhancers (stimulate transcription)
- Silencers (inhibit transcription)
* often found nearby (-50 to -100) but can also be
found great distances away in either direction
Factors that Control Gene Expression
• cis-acting elements
• DNA sequences that exert their effect only on nearby
genes
• Example: TATA box, enhancers and silencers
• trans-acting elements
• Regulatory proteins that bind to such DNA sequences
• In Bacteria:
– DNA coding strand = mRNA sequence directly
– Corresponds to amino acid sequence in the polypeptide
= colinearity of gene expression
• In Eukaryotes, RNA is modified:
- coding sequences, called exons, are interrupted by
intervening sequences or introns (which are removed)
= RNA splicing
• In eukaryotes, the transcription of
structural genes, produces a long
transcript known as pre-mRNA
– Also as heterogeneous
nuclear RNA (hnRNA)
• RNA is altered by splicing and
other modifications, before it
leaves the nucleus
• Spliceosome required for splicing
= multicomponent structure
Figure 12.16
Capping
• Most mature mRNAs have a 7-methyl guanosine covalently
attached at their 5’ end = capping
• Occurs as pre-mRNA is being synthesized (RNA pol II)
• Cap structure is recognized by cap-binding proteins
Role of Cap-binding proteins
– Movement of some RNAs into the cytoplasm
– Early stages of translation
– Splicing of introns
Addition of poly A tail
• Most mature mRNAs have a string of adenine
nucleotides at their 3’ ends = polyA tail
• polyA tail is not encoded in the gene sequence
– added enzymatically after the gene is completely
transcribed
Figure 12.20
Consensus sequence in
higher eukaryotes
Appears to be important in the
stability of mRNA and the
translation of the polypeptide
Length varies between species
From a few dozen adenines
to several hundred
Intron Advantage?
Alternative splicing:
• pre-mRNA with multiple introns can be spliced in
different ways
• Generates mature mRNAs with different
combinations of exons = different polypeptides
• Organism can carry fewer genes within genome
• Variation in splicing can occur in different cell types
or during different stages of development