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In prokaryotes, transcription produces a nearly exact mRNA copy of the DNA, and the transcript is immediately translated into protein. In eukaryotes, a series of modifications occur to mRNA during and after transcription.
在原核生物中，转录产生的 mRNA 几乎是 DNA 的准确拷贝，并且这一转录产物会立即被转译成蛋白质。在真核生物中，转录时以及转录后会对 mRNA 进行一系列修饰。
Chapter 5 mRNA Modifications in Eukaryotes
第 5 章 真核生物 mRNA 的修饰
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5.1 Capping
5.2 Polyadenylation
5.3 Splicing
5.4 mRNA Editing
5.5 Experiments
5.1 加帽
5.2 聚腺苷酸化
5.3 剪接
5.4 mRNA 编辑
5.5 实验研究
Chapter 5 mRNA Modifications in Eukaryotes
第 5 章 真核生物 mRNA 的修饰
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5.1 Capping / 加帽An mRNA that has been transcribed but is not yet ready for translation is called a pre-mRNA, or a primary transcript.
Capping is the process of adding a derivative of guanine nucleotide to the 5’ end of the pre-mRNA.
一条已经转录出来但还没有准备好用于转译的 mRNA 称为前体 mRNA 或初级转录本。
加帽是在前体 mRNA的 5’ 末端加上一个鸟嘌呤核苷酸衍生物的过程。
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Structure of the cap / 帽的结构5’-3’ phospho-diester bond
5’-5’ triphosphate bond
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RNA polymerase II
Discussed in Chapter 4
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Capping takes place quite early.
5’-Cap
RNA polymerase II
RNA
DNA
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Capping process / 加帽过程
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CTD: C-terminal domain
RNA polymerase II
DNA
Methyl transferase
Guanylyltransferase
RNA triphosphatase
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Functions of the cap structure帽结构的功能
1. Helps prevent degradation 帮助防止降解
2. Helps transport into cytoplasm 帮助转运到细胞质中
3. Enhances translation / 增强转译
4. Helps remove the first intron 帮助去除第一个内含子
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1. Helps prevent degradation帮助防止降解
RNase RNase
5’-5’ triphosphate bond
5’-3’ phosphodiester bond
5’-3’ phosphodiester bond
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2. Helps transport into cytoplasm帮助转运到细胞质中
Discussed in Chapter 7
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3. Enhances translation / 增强转译
Cap-binding protein
No translation occurs.
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RNA polymerase II
RNA
DNA
4. Helps remove the first intron帮助去除第一个内含子
The first intron
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AAAAAAA - - - - - - AAAA
5.2 Polyadenylation / 聚腺苷酸化
Poly(A) tail
Polyadenylation
Pre-mRNA
Polyadenylation is a modification process in which a string of about 250 adenine nucleotides is added to the 3’ end of the transcript.
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Polyadenylation does not occur at the natural end
AAAAAAA - - - - - - AAAA
Poly(A) tail
Polyadenylation
Pre-mRNA
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The polyadenylation signal / 聚腺苷酸化信号
AAAAAAA - - - - - - AAAA
Poly(A) tail
Polyadenylation
Pre-mRNA
Polyadenylation signal has a typical typical sequence of AAUAAA. This sequence gives an instruction as “to cut the mRNA about 20 nucleotides downstream, near a GU-rich sequence”.
AAUAAA GU
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The cleavage complex / 切割复合体
Cleavagecomplex
CPSF (cleavage andpolyadenylationspecificity factor)
CstF (cleavagestimulation factor)
CFI (cleavage factor I)
CFII (cleavage factors II)
Poly(A)-Binding Protein
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CTD: C-terminal domain
RNA polymerase II
DNA
Methyl transferase
Guanylyltransferase
RNA triphosphatase
Proteinsfor cappingat CTD
Proteins forpolyadenylationat CTD
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Functions of the poly(A) tailpoly(A) 尾的功能
AAAAAAA - - - - - - AAAA
AAAAAAA - - -
AA
A
AA
A
The main function of poly(A) tail is to protect the mRNA from degradation by ribonucleases.
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Functions of the poly(A) tailpoly(A) 尾的功能
AAAAAAA - - - - - - AAAA
There is also some evidence that the poly-A tail
is involved in splicing and enhances translation
of mRNAs.
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5.3 Splicing / 剪接Exons: Parts of a gene that are expressed as protein. Introns: Sequences that do not code for protein and interrupt the coding regions.
Splicing: The process of removing introns from a pre-mRNA.
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5.3.1 The Basic Splicing Reaction基本的剪接反应
5’ AG/GUAUGU…body of intron…UACUAAC-YAG / 3’
Splice sites in yeast
Splice sites: Sequences that mark the beginning and ends of introns and exons.
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The basic splicing reaction
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5.3.2 Proteins involved in Splicing在剪接中发挥作用的蛋白质
Spliceosome: The col
lection of factors, esp
ecially snRNPs, that h
elp with the splicing o
f introns.
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snRNPs: small nuclear ribonucleoproteins
snRNPs are small particles found in the nucleus and contain both protein and RNA.
snRNPs functioning in splicing: U1, U2, U4, U5 and U6.
U
A
C
G
C
G
A
U
U
A
A
U
C
G
A
U5’ 3’
Exon Intron
RNA in snRNP
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Now showing
mRNA splicing
File: biophoto7 \ Lehninger Principles of Biochemistry \ mRNA splicing
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5.3.3 Self-Splicing / 自我剪接
Tetrahymena thermophilia嗜热四膜虫
Splicing that occurs without the help of proteins or snRNPs is called self-splicing.
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Self-splicing of group I intron
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Self-splicing of group II intronII 类内含子的自我剪接
With helpof proteins Without help
of proteins
Self-splicingNormalsplicing
U2 A
U5U4
U1U6
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5.3.4 Trans-Splicing / 反式剪接
Mature mRNA
ProteinRNA piece of
separate origin
DNA
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Trans-Splicing / 反式剪接
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5.3.5 Reasons for Introns内含子存在的原因
G
G
U
A
5’ 3’Exon 1 Exon 2
Intron
Introns-early theory
Introns-late theory
Intron: “Why am I here?”
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Intron-early theory / 内含子早现说
E. coli
“I heard somebody say E. coli once had introns. Is that true?”
“Couldbe. Who knows?!”
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Intron-late theory / 内含子后现说
E. coli
“We are the oneshaving introns!
We are proud of it.”
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Alternative splicing / 可变剪接Alternative splicing: a kind of splicing that can produce various proteins from one gene.
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Drosophila Dscam gene
Exon 3 Exon 54.1 4.2 4.3 4.4 4.5 4.6 4.7 4.8 4.9 4.10 4.11 4.12
Exon 4 Exon 6 Exon 9
Alternative Splicing Goes Mainstream可变剪接成为主流
The Scientist, Volume 17(24): 28, Dec. 15, 2003
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Alternative splicing / 可变剪接
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5.4 mRNA Editing / mRNA 编辑Sleeping sickness is one of the most neglected
human diseases, and mainly affects the most deprived peoples of some African countries.
Trypanosome 锥虫
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RNA editing in trypanosome
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CU editing / CU 编辑
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In the liver … / 在肝脏中…
Translation
APOB (large)
APOB mRNA-CGA-
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In the small intenstine …
C U editong
APOB (small)
APOB mRNA
-UGA-
-CGA-
Translation
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5.5 Experiments / 实验研究
Hybridization between mRNA with DNA
mRNA 与 DNA 之间的杂交
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Detection of introns with DNase
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Hybridization between pre-mRNA and DNA前体 mRNA 与 DNA 之间的杂交
