1. RNA and Gene Expression BIO 224 Intro to Molecular and Cell Biology

2. RNA Molecules Three major classes – Ribosomal (rRNA) make up parts of ribosomes – Messenger (mRNA) provide RNA copies of genes – Transfer (tRNA) smallest of RNAs, bring AAs to site of protein synthesis

3. Role of RNA DNA does not directly dictate protein synthesis A molecule is needed to take information from DNA to the site of protein synthesis RNA can be made from a DNA template – SS molecule, uses ribose as sugar, has pyrimidine U instead of T – Characteristics suggested the central dogma of the flow of genetic information in molecular biology DNA  RNA  Protein – mRNA molecules transcribed from DNA, serve as template for translation of proteins

4. Transcription Similar to DNA replication, but different process Makes a complementary RNA copy of the DNA strand Entire genome never transcribed at once – Only transcribe certain genes/gene groups at certain times RNA transcription enzymes need to recognize which gene to transcribe and where to begin – RNA polymerase enzyme transcribes RNA from DNA template: mRNA RNA polymerase creates nucleic acid polymer of ribonucleotides – Discriminates between DNA and RNA nucleotides – Only adds ribonucleotides to polymer, creates only RNA molecule

5. 4.9 Synthesis of RNA from DNA

6. RNA Polymerase Recognizes start and stop points of DNA molecule Morphologically different from DNA polymerase – Both add nucleotides to 3’ OH of polymers One recognized in E coli, several identified in eukaryotes Made of subunits, recognizes beginning of gene Binds over region of 60 bp or so, causes DNA to locally unwind for initiation of transcription Promoter is region upstream or at beginning of gene that is bound – Specialized short DNA sequence – If mutated, can’t be bound, doesn’t function

7. 7.1 E. coli RNA polymerase

8. 7.5 Structure of bacterial RNA polymerase

9. Initiation Promoter sequences are upstream and downstream of start site Near promoter is closest to transcription beginning, has conserved sequence: TATAATA (TATA box) Far promoter has conserved sequence: TTGACA RNA subunit σ allows specific binding to promoter region sequences

10. 7.2 Sequences of E. Coli promoters

11. Elongation After RNA polymerase binds to promoter DNA is unwound to provide template Unwinds near beginning of gene and provides 3’OH for template 2 RNTPs are added for transcription to begin After addition of 10 nucleotides, σ dissociates and allows continuation of elongation

12. 7.4 Transcription by E. coli RNA polymerase (Part 1)

13. 7.4 Transcription by E. coli RNA polymerase (Part 2)

14. Termination RNA polymerase recognizes termination signal to end transcription Inverted repeat of GC rich area followed by poly-A tail transcribed to form stem-loop hairpin structure Structure disrupts RNA association with DNA template and terminates transcription Other method involves protein Rho that binds extended SS segments of RNA to cause termination

15. 7.6 Transcription termination

16. Eukaryote Transcription Similar to that in prokaryotes Have different RNA polymerases divided into classes Class I transcribes rRNAs Class II transcribes mRNAs Class III transcribes tRNAs Mitochondria and chloroplasts have different RNA polymerases

18. 7.11 Structure of yeast RNA polymerase II

19. Eukaryote Initiation RNA polymerase recognizes promoter sequence Upstream promoter sequence TATAA similar to bacterial TATA box for initiation Also have downstream promoter element; some genes use only this for initiating transcription along with Inr Elongation occurs in similar manner to prokaryotes

20. 7.19 A eukaryotic promoter

21. 7.12 Formation of a polymerase II transcription initiation complex (Part 1)

22. 7.12 Formation of a polymerase II transcription initiation complex (Part 2)

23. 7.13 Model of the polymerase II transcription initiation complex

24. 7.14 RNA polymerase II/Mediator complexes

25. 7.17 Transcription of RNA polymerase III genes

26. Messenger RNA Vary in size small to very large RNA copy of information in DNA Prokaryote mRNA is translated by ribosomes in the cytoplasm while still being transcribed Prokaryotic mRNAs do not exist for long Often degraded within minutes

27. Eukaryote mRNA Transcribed as pre-mRNA in nucleus and processed before export Introns removed, 5’ end capped with 7- methylguanosine, 3’ end polyadenylated with poly-A tail Polyadenylation leads to termination of transcription, important in regulation of translation

28. 7.44 Processing of eukaryotic messenger RNAs

29. 7.45 Formation of the 3’ ends of eukaryotic mRNAs

30. 7.47 Splicing of pre-mRNA

31. 7.50 Self-splicing introns

32. 7.52 Alternative splicing in Drosophila sex determination

33. 7.54 Editing of apolipoprotein B mRNA

34. 7.55 Regulation of transferrin receptor mRNA stability

35. mRNA Translation All mRNAs translated in 5’ to 3’ direction Polypeptide chains assembled from amino to carboxy terminus Each AA specified by an mRNA codon dictated by genetic code Translation occurs on ribosomes, needs all three types of RNAs plus proteins

36. Expression of Genetic Information DNA  RNA  Protein Genes determine protein structure Proteins direct cell metabolism via enzymatic activity Genetic information specified by arrangement of DNA bases Proteins are polymers of 20 AAs determined by sequence that dictates structure and function Mutation in DNA sequence leads to AA sequence change: colinearity

37. 4.8 Colinearity of genes and proteins

38. The Genetic Code Genetic code allows understanding of how sequence of 4 nucleotides is converted to 20 AAs tRNAs act as adaptor between AAs & mRNAs during translation tRNA anticodons pairs in complementary fashion with mRNA codons for attachment of AA to polypeptide chain Three nucleotides specify each AA 64 codons in genetic code: 61 code for AAs, 3 stop codons – Some AAs specified by more than one codon Nearly all organisms use same genetic code

40. 4.11 Genetic evidence for a triplet code

41. 4.12 The triplet UUU encodes phenylalanine