Upload
lalasa
View
41
Download
1
Tags:
Embed Size (px)
DESCRIPTION
LECTURE 14 RNA: TRANSCRIPTION & PROCESSING. chapter 8 announcements key ideas RNA transcription in Prokaryotes transcription in Eukaryotes. ANNOUNCEMENTS. missed or late for quiz ? same documentation required as for exams no makeup… average of those you write - PowerPoint PPT Presentation
Citation preview
LECTURE 14 RNA: TRANSCRIPTION & PROCESSING
chapter 8 announcements key ideas RNA transcription in Prokaryotes transcription in Eukaryotes
ANNOUNCEMENTS missed or late for quiz ?
same documentation required as for exams no makeup… average of those you write
exam 2 … M 11.13 in class, as listed on calendar extra tutorial help …
M 3:00 – 4:30 & F 11:00 – 12:30 in WHI 111 extra-credit tutoring … in progress, check web page extra-credit seminar WHI AUD F 3:30
Dr. Juli Wade, Michigan State U. Hormonal and Genetic Influences on Sexual
Differentiation of the Zebra Finch Song System
CH8 KEY IDEAS 3 processes of information transfer in genetics
CH8
CH7
CH9
CH8 KEY IDEAS
translation requires transfer RNAs & ribosomes information transfer by non-overlapping triplet code special DNA sequences signal initiation & termination
of transcription & translation in Eukaryotes only... mRNA transcripts are processed prior to translation noncoding introns interrupt coding exon sequences introns are spliced out of 1° mRNA final mRNA
DNA transcription RNA translation protein
CH8 KEY IDEAS transcription & translation
RNA DNA protein... RNA intermediate? pulse-chase experiments with RNA precursors... pulse
radioactive, chase nonradioactive, autoradiograph
synthesis of RNA in nucleus protein in cytoplasm
RNA
RNA
RNA
RNA
usually single-stranded, double-stranded, complex 3D structures antiparallel
ribose sugar deoxyribose C2
H
C2
OH
RNA
AU AT
RNA DNA
uracil pyrimidine thymine
classes
1. informational mRNA: translated polypeptides
2. functional tRNA: bind & transport amino acids rRNA: components of ribosomes snRNA: participate in modifying rRNA
components of spliceosomes
RNA
TRANSCRIPTION IN PROKARYOTES
T2 bacteriophage infection of E. coli pulse-chase with radiolabeled uracil (RNA-precursor) labeled RNA recovered only immediately after pulse...
rapid turnover phage-induced
RNA similar to T2 DNA... DNA RNA
TRANSCRIPTION IN PROKARYOTES in vitro synthesis shows RNA DNA...
DNA template & complementary RNA bases
similar to DNA replication, with DNA polymerase... transcription enzyme... RNA
polymerase ?
TRANSCRIPTION RNA synthesized from 1
or both DNA strands? hybridization experiment complementary DNA
strands have different densities
each RNA hybridizes to only 1 DNA strand
transcription is asymmetrical
TRANSCRIPTION IN PROKARYOTES DNA template for mRNA transcription
note directional nature of event
TRANSCRIPTION IN PROKARYOTES
DNA coding strand = nontemplate strand sequence homologous to transcribed mRNA DNA has A, RNA has U
DNA template strand mRNA
TRANSCRIPTION IN PROKARYOTES
new mRNA grows 5' 3' RNA pol moves along DNA template strand 3' 5'
5'3'
5'3'
error!error!error!
genes transcribed from one but either DNA strand transcription by complementary pairing of bases catalyzed by RNA polymerase (RNA pol)
TRANSCRIPTION IN PROKARYOTES
RNA pol moves along DNA template strand 3' 5'
new mRNA grows 5' 3' normally illustrate in other direction for convenience
TRANSCRIPTION IN PROKARYOTES
transcription of rRNA genes in Triturus nucleus
TRANSCRIPTION IN PROKARYOTES
3 distinct stages of transcription
1. initiation
2. elongation
3. termination
TRANSCRIPTION IN PROKARYOTES
initiation
2 types of sequences in prokaryotic genes
1. promoter sequences signal initiation
2. coding sequences are transcribed
TRANSCRIPTION IN PROKARYOTES initiation 2 regions of homology among promoter sequences
consensus sequences = RNA pol binding sites
TRANSCRIPTION IN PROKARYOTES initiation factor binds to –10 and –35 consensus regions initiates melting or denaturing of DNA transcription begins when subunit dissociates
open promoter complex
different factors recognize different DNA sequences
closed promoter complex
TRANSCRIPTION IN PROKARYOTES elongation new mRNA grows 5' 3' RNA pol and transcription bubble moves 3' 5'
along the DNA template strand
5'3'
TRANSCRIPTION
elongation
new mRNA grows 5' 3'
RNA pol moves along DNA
template strand 3' 5'
ribonucleoside triphosphate
added to 3' end of nth base
3'
5'
5'
3'
elongation
new mRNA grows 5' 3'
RNA pol moves along DNA
template strand 3' 5'
pyrophosphate ion
released
3'
3'
5'
5'
TRANSCRIPTION IN PROKARYOTES
termination RNA pol recognizes signal for chain termination 2 mechanisms for termination in prokaryotes
1. direct termination - termination sequence
TRANSCRIPTION IN PROKARYOTES
mRNA 3' UTR DNA CG rich + AAA...(6+)
5'3'
termination
direct termination, RNA pol recognizes ~40 bp terminator sequence on template
RNA forms hairpin loop
poly-A tail bonds weak
signal to release RNA pol
ATP-independent
TRANSCRIPTION IN PROKARYOTES
termination
RNA pol recognizes signal for chain termination
2 mechanisms for termination in prokaryotes
1. direct termination - termination sequence
2. rho-dependent
TRANSCRIPTION IN PROKARYOTES
termination
rho-dependent termination
no U-residues formed
no hairpin loop
rho binds to rut site
on RNA
TRANSCRIPTION IN PROKARYOTES
TRANSCRIPTION IN PROKARYOTES
termination
rho-dependent termination
no U-residues formed
no hairpin loop
rho binds to rut site
on RNA
rho “pulls” RNA from
RNA pol
TRANSCRIPTION IN PROKARYOTES
termination
rho-dependent termination
no U-residues formed
no hairpin loop
rho binds to rut site
on RNA
rho “pulls” RNA from
RNA pol
TRANSCRIPTION IN EUKARYOTES
differences because of complexity in Eukaryotes1. RNA synthesis
TRANSCRIPTION IN EUKARYOTES
Prokaryotes:
RNA pol all types of RNA
polycistronic mRNA
Eukaryotes:
RNA pol I rRNA (except 5S rRNA)
RNA pol II monocistronic mRNA, some snRNA
RNA pol III tRNA, 5S rRNA, some snRNA
TRANSCRIPTION IN EUKARYOTES
Prokaryotes:
RNA pol only
Eukaryotes:
RNA pol II + general transcription factors (GTFs) mRNA
DNA GTFs
TRANSCRIPTION IN EUKARYOTES
differences because of complexity in Eukaryotes1. RNA synthesis2. RNA processing
TRANSCRIPTION IN EUKARYOTES
Prokaryotes:
no processing
Eukaryotes:
processed before being transported to the cytoplasm
TRANSCRIPTION IN EUKARYOTES
differences because of complexity in Eukaryotes1. RNA synthesis2. RNA processing3. chromosome organization4. split genes
TRANSCRIPTION IN EUKARYOTES
Prokaryotes:
naked DNA (nearly)
Eukaryotes:
chromatin – euchromatin & heterochromatin
TRANSCRIPTION IN EUKARYOTES
3 distinct stages of transcription
1. initiation
2. elongation
3. termination
TRANSCRIPTION IN EUKARYOTES initiation GTFs TATA sequence
before RNA pol II binding attract RNA pol II positions complex
GTFs added after preinitiation complex transcription bubble
TRANSCRIPTION IN EUKARYOTES elongation GTFs added after
preinitiation complex transcription bubble RNA pol II carboxyl
tail domain (CTD) phosphorylated...
TRANSCRIPTION IN EUKARYOTES 2 types of processing
1. cotranscriptional
TRANSCRIPTION IN EUKARYOTES cotranscriptional processing – CTD dependend
guanyltransferase adds 7'-methylguanosine “cap” to 5' end of mRNA
protects single stranded RNA from degradation
TRANSCRIPTION IN EUKARYOTES cotranscriptional processing – CTD dependend
splicing by spliceosomes (... stay tuned)
TRANSCRIPTION IN EUKARYOTES termination
AAUAAA sequence near 3' end initiates cleavage
... by endonuclease ~ 20 bp downstream
TRANSCRIPTION IN EUKARYOTES 2 types of processing
1. cotranscriptional
2. posttranscriptional
TRANSCRIPTION IN EUKARYOTES posttranscriptional processing
poly(A) polymerase adds poly(A) tail of 150-200 adenosine residues to 3' end cleavage site
complete °1 mRNA
EUKARYOTIC RNA
°1 mRNA shortened before transport to cytoplasm
chicken ovalbumin DNA/mRNA hybrid
coding sequences... exons
EUKARYOTIC RNA
°1 mRNA shortened before transport to cytoplasm
chicken ovalbumin DNA/mRNA hybrid
coding sequences... exons intervening sequences...
introns
EUKARYOTIC RNA
°1 mRNA shortened before transport to cytoplasm
chicken ovalbumin DNA/mRNA hybrid
coding sequences... exons intervening sequences...
introns introns spliced from mRNA genomic DNA RNA
exons + introns
summary... genomic DNA
RNA exons + introns transcribed 1°
mRNA transcript processed cap &
polyadenylation spliced splicing
intermediate spliced mature
mRNA
TRANSCRIPTION IN EUKARYOTES
1 gene multiple functions by alternative splicing
multiple gene functions in different... tissues developmental stages
different mRNAs from same 1° mRNA transcript... e.g., -tropomyocin gene...
TRANSCRIPTION IN EUKARYOTES
1 gene multiple functions by alternative splicing e.g., -tropomyocin gene...
TRANSCRIPTION IN EUKARYOTES
gene splicing mechanism... sequence homologies at exon-intron splice junctions = consensus sequences for splicing enzymes...
TRANSCRIPTION IN EUKARYOTES
spliceosome snRNA aligns sequences correctly for splicing
TRANSCRIPTION IN EUKARYOTES
splicing reaction intron lariat structure introns excised by 2
transesterification reactions
TRANSCRIPTION IN EUKARYOTES
splicing reaction intron lariat structure introns excised by 2
transesterification reactions snRNA + proteins small
ribonuclear particles (snRNPs) snRNP + 1° mRNA transcript
spliceosome spliceosome catalyzes splicing
transesterification reactions
TRANSCRIPTION IN EUKARYOTES
self splicing mechanisms, mRNA catalyzes reaction (called ribozymes)
TRANSCRIPTION IN EUKARYOTES
self splicing mechanisms, mRNA catalyzes reaction spliceosome-catalyzed mechanism
TRANSCRIPTION IN EUKARYOTES
TUTORIAL 6
QUESTIONS – CHAPTER 8