Part 3
Gene expression (Ⅰ)transcription
Http : //web.umr.edu/.../ 331figures.html
Summary for Transcription
the Recognization of template ↓
Initiation↓
Pass the promoter↓
Elongation↓
Termination
Ⅰ. Transcription in Prokaryote
Ⅱ. Transcription in Eucaryote
Ⅲ. Products of transcription
Ⅰ. transcription in ProkaryoteIntroduction:• RNA polymerase• Promoter • Initiation and
termination of transcription
RNA polymerase
核心酶
σ 因子
Function of subunit:
α : combine to the promoter, unwind the double-strand
β : for the nucleotide’s combining in initiation and elongation process
β’ : recognize and combine to the template ,unspecifically
σ : recognize and combine to the template specifically
ω : unkown
Promoter
• -10 signal (TATA box,Pribnow box)
• -35 signal
• transcription start site
TATAATTGACA
-35 box
+1
-10 box
-40 ~-30
转录起点
上游
upstream下游
downstream
原核生物的启动子
17+1bp
-10 ~ -7
Initiation
The process of transcription initiation, with every subunit of the RNA polymerase making function orderly ,is from the template being recognized and bound to the first nucleotide residue being transcribed on the start site.
+1
+1
+1
+1
-10
-10
-10
-10
-35
-35
-35
-35
β’
σ
α
β
脱落的 σ ATP or GTP
1
2
3
4
Termination
Two different pattern: ρ- independent termination
ρ –dependent termination
1.reverse repeat with GC rich
2.poly (A) following the GC rich se
quence directly in template
ρ factor:
(1) moves along the nascent RNA towards the transcription complex
(2) Hydrolyzes the ATP
(3) Helicase
不依赖ρ因子的终止子
依赖ρ因子的终止子
Ⅱ. Transcription in Eucaryote
Introduction:• RNA polymerase• Promoter and transcription initiation• Termination of transcription• Post-transcriptional Processing
RNA polymerase
Multi-subunit enzyme
Contain: two large subunits whose MW are more than 1*105 several small subunits which may be the same in 2 o
r 3 kinds of RNA polymerase
different RNA polymerase is with responsibility for different gene
Polymerase
location Respondent gene
RNA polymerase Ⅰ
Nucleoli ( 核仁 )
rRNA gene cluster (except 5s rRNA gene)
RNA polymerase Ⅱ
nucleoplasm (核质)
all genes encoding protein, some encoding U-RNA
RNA polymerase Ⅲ
nucleoplasm (核质)
5s rRNA gene, tRNA gene
RNA polymerase and its respondent gene
rRNA gene cluster (Pol gene )Ⅰ
18s-5.8s-28s
18s-5.8s-28s
18s-5.8s-28s
18s-5.8s-28s
promoterPromoterrepeat
Promoter repeat
60/81bp repeatNTS
Typical mRNA gene (pol gene)Ⅱ
Coding region5’- 侧
翼 3’ - 侧翼
5’UTR
3’UTR
EXON EXON EXON EXON
Pol geneⅢ
+1
内启动子
A box
+50~+65
C box
+81~+99
5s rRNA gene
tRNA gene
+1
内启动子
A box B box
D-loop TψC-loop
Promoter & transcriptional initiation
• Promoter : A sequence region which promote the initiation of transcription
• Transcriptional initiation
RNA pol promoterⅠ
+1
+6-31-100
UCE:50~80bp Core promoter Pre-rRNA gene
Enhance transcription
Essential for transcription
Initiation of pol gene’s transcriptionⅠUBFUBF
SL1POL Ⅰ
NOTE: UBF----upstream binding factor
SL1----selectivity factor
RNA Pol PromoterⅡ
Consist of:1) Transcription start-site: A or G
2) TATA box: determine the exact starting
3) UAS (UPE): determine the frequency of initiation
4) Enhancer : enhance the frequency of initiation
ATATAACCAATGCCACACCC
或
GGGCGGG
+1
转录起点TATA boxCAAT boxGC box增强子
UPE or UAS
Py A Py
promoter model
-35 ~ -25-80 ~ -70-110 ~ -80
Enhancer in SV40
72bp ,分 A 、 B 、 C三区
A C B
-200-220-232-250 -248-271
两个核心 A
或 核心 A+ 部分A
核心 C+TCⅡ
Sph +SphⅡ Ⅰ
三个主区相互作用发挥增强转录的作用
The enhancer’s architecture in SV40
Transcription initiation of pol geneⅡ
Transcription factors recognize and bind to the promoter to assemble an initiation complex(basal transcriptional initiation complex or activated transcriptional initiation complex)
Basal transcriptional initiation complex
Activated transcriptional initiation complex
TATA
ⅡAⅡB
ⅡF
ⅡE
ⅡH
TBPTAFs
PolⅡCBPUSA
Mediator
AAAAAATAAA
3’
加尾信号 Poly(A)
真核生物中与转录终止有关的序列
Termination
终止
0.5~2 kb15~30 bp
Termination of transcription
In eucaryote, the transcription termination is different from that in prokaryote.
polyadenylation signal & Poly(A) addition site: the terminus signal but not the terminator
Terminate downstream
TF CⅢ
Pol promoter & transcriptioⅢn initiation
+1内启动子
A box
+50~+65
C box
+81~+99
5s rRNA gene
TF BⅢ
Pol Ⅲ
TF AⅢ
内启动子
A box B box
tRNA gene
D-loop TψC-loop
+1
TF CⅢTF BⅢ
Pol Ⅲ
Post-transcriptional Processing
• tRNA processing
• rRNA processing
• mRNA processing
rRNA processing
ITS1 ITS2ETS1 ETS2
RNase
RNase
RNase
tRNA processing
mRNA processing
• 5’ capping• 3’ polyadenylation• splicing : primary product: hnRNA ( intron, exon)
• RNA editing and modification
mRNA
POSTTRANSCRIPTIONAL PROCESS
5’ capping
m7G 5’-p-p-p-5’AN
m7G ppp p-p-p-5’ A N
腺苷酸转移酶甲基转移酶
Cap 0
Cap 1
Cap 2
5’ , 5’ – 磷酸二酯键
m7G 5’-p-p-p-5’Am2’N
m7G 5’-p-p-p-5’Am2’Nm2’
GTP 尿苷酸 -7- 甲基转移酶
3’ polyadenylation
AAUAAA CASTOP15~30 bases
0.5~2kb
polyadenylation
Cleavage site
Poly(A)addition site
Pre-mRNA
splicing Interrupted gene: exon & intron Classes of intron:
» 主要的内含子 : 5 ’GU……AG 3’» 次要的内含子 : 5’ AU……AC 3’» Ⅰ 类内含子» Ⅱ 类内含子» Ⅲ 类内含子» 双内含子» tRNA 前体中的内含子
Introns are spliced in posttranscription processing :见课本 P92 、 94 、 95
Alternative splicing: Reference: “regulation of eucaryote gene expression”
主要的内含子
exon exon
AGGUAAGU PyPyPyPyPyPyNCAG
5’ 剪接位点
3’剪接位点
多聚嘧啶系列
CURAY
分支点序列
RNA modification
Modification:specific chemical modification. 甲基化 去氨基 硫代 积极的同分异构化 二价键的饱和化 核苷酸的替代
Methylation guided by snoRNA
RNA editing
Reference: “regulation of eucaryote gene expression”
Editing: to alter the coding information by change the mRNA sequence.
Products of transcription
• In prokaryote short half-life:Multi-cistronStarting codon & SD sequence• In eucaryote 5’ cap3’ tailPrimary products must be peocessed to pro
duce mature mRNA
SD sequence
AUG~10bases
UAAGGAGGU:complementary with 16s rRNA
5’
The site for ribosome binding
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