Chapter 5 mRNA Modifications in Eukaryotes 第 5 章 真核生物 mRNA 的修饰

1. Chapter 5 mRNA Modifications in Eukaryotes第5章 真核生物mRNA的修饰 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。 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.

2. Chapter 5 mRNA Modifications in Eukaryotes第5章 真核生物mRNA的修饰 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

3. 5.1 Capping / 加帽 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。 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.

4. Structure of the cap / 帽的结构 5’-3’ phospho- diester bond 5’-5’ triphosphate bond

5. RNA polymerase II Discussed in Chapter 4

6. Capping takes place quite early. 5’-Cap RNA DNA RNA polymerase II

7. Capping process / 加帽过程

8. CTD: C-terminal domain RNA triphosphatase Methyl transferase Guanylyl transferase DNA RNA polymerase II

9. 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

10. 1. Helps prevent degradation帮助防止降解 RNase RNase 5’-3’ phospho diester bond 5’-3’ phospho diester bond 5’-5’ triphosphate bond

11. 2. Helps transport into cytoplasm帮助转运到细胞质中 Discussed in Chapter 7

12. 3. Enhances translation / 增强转译 Cap-binding protein No translation occurs.

13. 4. Helps remove the first intron帮助去除第一个内含子 The first intron RNA DNA RNA polymerase II

14. 5.2 Polyadenylation / 聚腺苷酸化 Polyadenylationis a modification process in which a string of about 250 adenine nucleotides is added to the 3’ end of the transcript. Pre-mRNA Polyadenylation AAAAAAA - - - - - - AAAA Poly(A) tail

15. Polyadenylation does not occur at the natural end

16. The polyadenylation signal / 聚腺苷酸化信号 Polyadenylation signalhas 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”. Pre-mRNA AAUAAA GU Polyadenylation AAAAAAA - - - - - - AAAA Poly(A) tail

17. The cleavage complex / 切割复合体 CPSF (cleavage and polyadenylation specificity factor) Cleavage complex CstF (cleavage stimulation factor) CFI (cleavage factor I) CFII (cleavage factors II)

18. Poly(A)-Binding Protein

19. CTD: C-terminal domain Proteins for capping at CTD Proteins for polyadenylation at CTD

20. Functions of the poly(A) tailpoly(A)尾的功能 The main function of poly(A) tail is to protect the mRNA from degradation by ribonucleases. AAAAAAA - - - - - - AAAA A AAAAAAA - - - A A A A A

21. Functions of the poly(A) tailpoly(A)尾的功能 There is also some evidence that the poly-A tail is involved insplicing and enhances translation of mRNAs. AAAAAAA - - - - - - AAAA

22. 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.

23. 5.3.1 The Basic Splicing Reaction基本的剪接反应 Splice sites: Sequences that mark the beginning and ends of introns and exons. Splice sites in yeast 5’AG/GUAUGU…body of intron…UACUAAC-YAG/ 3’

24. The basic splicing reaction

25. 5.3.2 Proteins involved in Splicing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. snRNPs: small nuclearribonucleoproteins snRNPs are small particles found in the nucleus and contain both protein and RNA. snRNPs functioning in splicing: U1, U2, U4, U5 and U6.

29. Now showingmRNA splicing File: biophoto7 \Lehninger Principles of Biochemistry \ mRNA splicing

30. 5.3.3 Self-Splicing / 自我剪接 Splicing that occurs without the help of proteins or snRNPs is called self-splicing. Tetrahymena thermophilia 嗜热四膜虫

31. Self-splicing of group I intron

32. Self-splicing of group II intronII 类内含子的自我剪接 Normal splicing Self-splicing With help of proteins Without help of proteins

33. 5.3.4 Trans-Splicing / 反式剪接 DNA Mature mRNA Protein RNA piece of separate origin

34. Trans-Splicing / 反式剪接

35. 5.3.5 Reasons for Introns内含子存在的原因 Introns-early theory Intron: “Why am I here?” Introns-late theory

36. Intron-early theory / 内含子早现说 E. coli “Could be. Who knows?!” “I heard somebody say E. coli once had introns. Is that true?”

37. Intron-late theory / 内含子后现说 E. coli “We are the ones having introns! We are proud of it.”

38. Alternative splicing / 可变剪接 Alternative splicing: a kind of splicing that can produce various proteins from one gene.

39. Drosophila Dscam gene Exon 4 Exon 6 Exon 9 Exon 3 Exon 5 4.1 4.2 4.3 4.4 4.5 4.6 4.7 4.8 4.9 4.10 4.11 4.12 Alternative Splicing Goes Mainstream 可变剪接成为主流 The Scientist, Volume 17(24): 28, Dec. 15, 2003

40. Alternative splicing / 可变剪接

41. 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

42. RNA editing in trypanosome

43. CU editing/ CU 编辑

44. In the liver …/ 在肝脏中…

45. In the small intenstine …

46. 5.5 Experiments / 实验研究 Hybridization between mRNA with DNA mRNA与DNA之间的杂交

47. Detection of introns with DNase

48. Hybridization between pre-mRNA and DNA前体mRNA与DNA之间的杂交