分子生物学第五章

1、1 / 48In 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，转录产在原核生物中，转录产生的生的mRNA几乎是几乎是DNA的准确拷贝，并且的准确拷贝，并且这一转录产物会立即被这一转录产物会立即

2、被转译成蛋白质。在真核转译成蛋白质。在真核生物中，转录时以及转生物中，转录时以及转录后会对录后会对mRNA进行进行一一系列修饰系列修饰。Chapter 5 mRNA Modifications in Eukaryotes第第5章章 真核生物真核生物mRNA的修饰的修饰2 / 485.1 Capping5.2 Polyadenylation5.3 Splicing5.4 mRNA Editing5.5 Experiments5.1 加帽加帽5.2 聚腺苷酸化聚腺苷酸化5.3 剪接剪接5.4 mRNA编辑编辑5.5 实验研究实验研究Chapter 5 mRNA Modifications in E

3、ukaryotes第第5章章 真核生物真核生物mRNA的修饰的修饰3 / 485.1 Capping / 加帽加帽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一条已经转录出来但还没有准备好用于转还没

4、有准备好用于转译的译的mRNA称为称为前体前体mRNA或或初级转录本初级转录本。加帽加帽是在前体是在前体mRNA的的5末端加上一个鸟末端加上一个鸟嘌呤核苷酸衍生物的嘌呤核苷酸衍生物的过程。过程。4 / 48Structure of the cap / 帽的结构帽的结构5-3 phospho-diester bond5-5 triphosphate bond5 / 48RNA polymerase IIDiscussed in Chapter 46 / 48Capping takes place quite early.5-CapRNA polymerase IIRNADNA7 / 48Capp

5、ing process / 加帽过程加帽过程8 / 48CTD: C-terminal domainRNA polymerase IIDNAMethyl transferaseGuanylyltransferaseRNA triphosphatase9 / 48Functions of the cap structure帽结构的功能帽结构的功能1. Helps prevent degradation 帮助防止降解帮助防止降解2. Helps transport into cytoplasm 帮助转运到细胞质中帮助转运到细胞质中3. Enhances translation / 增强转译增强转译

6、4. Helps remove the first intron 帮助去除第一个内含子帮助去除第一个内含子10 / 481. Helps prevent degradation帮助防止降解帮助防止降解RNaseRNase5-5 triphosphate bond5-3 phosphodiester bond5-3 phosphodiester bond11 / 482. Helps transport into cytoplasm帮助转运到细胞质中帮助转运到细胞质中Discussed in Chapter 712 / 483. Enhances translation / 增强转译增强转译Ca

7、p-binding proteinNo translation occurs.13 / 48RNA polymerase IIRNADNA4. Helps remove the first intron帮助去除第一个内含子帮助去除第一个内含子The first intron14 / 48AAAAAAA - - - - - - AAAA5.2 Polyadenylation / 聚腺苷酸化聚腺苷酸化Poly(A) tailPolyadenylationPre-mRNAPolyadenylation is a modification process in which a string of ab

8、out 250 adenine nucleotides is added to the 3 end of the transcript.15 / 48Polyadenylation does not occur at the natural end 16 / 48The polyadenylation signal / 聚腺苷酸化信号聚腺苷酸化信号AAAAAAA - - - - - - AAAAPoly(A) tailPolyadenylationPre-mRNAPolyadenylation signal has a typical typical sequence of AAUAAA. T

9、his sequence gives an instruction as “to cut the mRNA about 20 nucleotides downstream, near a GU-rich sequence”. AAUAAAGU17 / 48The cleavage complex / 切割复合体切割复合体CleavagecomplexCPSF (cleavage andpolyadenylationspecificity factor)CstF (cleavagestimulation factor)CFI (cleavage factor I)CFII (cleavage f

10、actors II)Poly(A)-Binding Protein19 / 48CTD: C-terminal domainProteinsfor cappingat CTDProteins forpolyadenylationat CTD20 / 48Functions of the poly(A) tailpoly(A)尾的功能尾的功能AAAAAAA - - - - - - AAAAAAAAAAA - - -AAAAAAThe main function of poly(A) tail is to protect the mRNA from degradation by ribonucle

11、ases.21 / 48Functions of the poly(A) tailpoly(A)尾的功能尾的功能AAAAAAA - - - - - - AAAAThere is also some evidence that the poly-A tail is involved in splicing and enhances translation of mRNAs. 22 / 485.3 Splicing / 剪接剪接Exons: Parts of a gene that are expressed as protein. Introns: Sequences that do not c

12、ode for protein and interrupt the coding regions. Splicing: The process of removing introns from a pre-mRNA.23 / 485.3.1 The Basic Splicing Reaction基本的剪接反应基本的剪接反应5 AG/GUAUGUbody of intronUACUAAC-YAG / 3Splice sites in yeastSplice sites: Sequences that mark the beginning and ends of introns and exons

13、.24 / 48The basic splicing reaction25 / 485.3.2 Proteins involved in Splicing在剪接中发挥作用的蛋白质在剪接中发挥作用的蛋白质Spliceosome: The collection of factors, especially snRNPs, that help with the splicing of introns. 26 / 48snRNPs: small nuclear ribonucleoproteinssnRNPs are small particles found in the nucleus and c

14、ontain both protein and RNA. snRNPs functioning in splicing: U1, U2, U4, U5 and U6.29 / 48Now showingmRNA splicingFile: biophoto7 Lehninger Principles of Biochemistry mRNA splicing30 / 485.3.3 Self-Splicing / 自我剪接自我剪接Tetrahymena thermophilia嗜热四膜虫嗜热四膜虫 Splicing that occurs without the help of protein

15、s or snRNPs is called self-splicing. 31 / 48Self-splicing of group I intron32 / 48Self-splicing of group II intronII 类内含子类内含子的自我剪接的自我剪接With helpof proteinsWithout helpof proteinsSelf-splicingNormalsplicing33 / 485.3.4 Trans-Splicing / 反式剪接反式剪接Mature mRNAProteinRNA piece ofseparate origin DNA34 / 48T

16、rans-Splicing / 反式剪接反式剪接35 / 485.3.5 Reasons for Introns内含子存在的原因内含子存在的原因Introns-early theoryIntrons-late theoryIntron: “Why am I here?”36 / 48Intron-early theory / 内含子早现说内含子早现说E. coli“I heard somebody say E. coli once had introns. Is that true?”“Couldbe. Who knows?!”37 / 48Intron-late theory / 内含子后现

17、说内含子后现说E. coli“We are the oneshaving introns! We are proud of it.”38 / 48Alternative splicing / 可变剪接可变剪接Alternative splicing: a kind of splicing that can produce various proteins from one gene.39 / 48Drosophila Dscam geneExon 3Exon 54.1 4.2 4.3 4.4 4.5 4.6 4.7 4.8 4.9 4.10 4.11 4.12Exon 4 Exon 6 Exo

18、n 9Alternative Splicing Goes Mainstream可变剪接成为主流可变剪接成为主流The Scientist, Volume 17(24): 28, Dec. 15, 2003 40 / 48Alternative splicing / 可变剪接可变剪接41 / 485.4 mRNA Editing / mRNA编辑编辑Sleeping sickness is one of the most neglected human diseases, and mainly affects the most deprived peoples of some African countries. Trypanosome 锥虫锥虫42 / 48RNA editing in trypanosome43 / 48CU editing / CU 编辑编辑