1. Chapter 15 The Genetic Code The code is degenerate ( 簡併的 ) Three rules govern the genetic code Suppressor mutations can reside in the same or a different gene The code is nearly universal

2. The code is degenerate ( 簡併的 ) Degeneracy ( 簡併性 ) : many amino acids are specified by more than one codon. Synonyms ( 同義密碼子 ): codons specifying the same amino acid.

3. Fig 15-1 codon-anticodon pairing of two tRNA Leu molecules.

5. Perceiving order in the makeup of the code The genetic code might have evolved in a way to minimize deleterious effects of mutations. 1.Codons with pyrimidines in the 2 nd position mostly specify hydrophobic amino acids; while those with purines in the 2 nd position mostly specify hydrophilic amino acids. 2.As the first two positions are both occupied by G or C, any of the four nucleotides in the 3 rd position will specify the same amino acid.

6. Wobble in the anticodon Base in anticodonBase in codon GU or C CG AU UA or G IA, U, or C Table 15-2 pairing combinations with the wobble concept Does a specific tRNA anticodon would exist for every codon? 1.Highly purified tRNA species of known sequences could recognize more than one codon. 2.Inosine (I)

7. Wobble concept ( 擺動概念 ) Francis Crick, 1966 The base at the 5’ end of the anticodon is not as spatially confined as the other two, allowing it to form hydrogen bonds with any of the several bases located at the 3’ end of the codon.

8. Fig. 15-2 wobble base pairing: ribose-ribose distances for all the wobble pairs are close to those of the standard A:U or G:C pairings.

9. Fig 15-3 structure of yeast tRNA phe. The 1 st anticodon base is at the end of the stacking interactions between flat surfaces of the bases, and is perhaps less restricted in its movement than the other two bases.

10. How the code was cracked?

11. Stimulation of amino acid incorporation by synthetic mRNAs Poly-U Poly-C Poly-G

12. Poly-U codes for polyphenylalanine A high magnesium concentration circumvents the need for initiation factors and the special initiator tRNA, allowing chain elongation to occur without proper signals in the mRNA. Poly-U was the first synthetic polyribonucleotide discovered to have mRNA activity. CCC: proline AAA: lysine Poly-G forms strong intra-chain hydrogen bond and does not interact with ribosome.

13. Mixed copolymers allowed additional codon assignments Table 15-3 Such experiments did not reveal the order of nucleotides within a codon. Transfer RNA binding to defined trinucleotide codons Table 15-4 Even in the absence of all the factors required for protein synthesis, specific aminoacyl-tRNA molecules can bind to ribosome-mRNA complexes.

14. Fig 15-5 preparing oligo-ribonucleotides. Organic synthesis and copying by DNA polymerase I.

15. Three rules govern the genetic code 1.Codons are read in a 5’ to 3’ direction. 2.Codons are nonoverlapping and the message contains no gaps. 3.The message is translated in a fixed reading frame, which is set by the initiation codon.

16. Three kinds of point mutations alter the genetic code Missense mutation ( 錯義突變 ): an alteration that changes a codon specific for one amino acid to a codon specific for another amino acid. Nonsense or stop mutation ( 無義突變 ): an alteration causing a chain-termination codon. Frameshift mutation ( 移碼突變 )

17. Genetic proof that the code is read in units of three Bacteriophage T4

18. Suppressor mutations can reside in the same or a different gene Reverse (back) mutations 回復突變 Suppressor mutations 抑制基因突變 Intragenic suppression Intergenic suppression

19. Fig 15-6 (intragenic) suppression of frameshift mutations.

20. Fig 15-7 nonsense suppression (intergenic suppression)

21. The code is nearly universal With the exception of mitochondral DNA Table 15-6

22. Eukaryotic gene regulation at steps after transcription initiation The regulation of alternative mRNA splicing can produce different protein products in different cell types

23. Fig 17-27 early transcriptional regulation of Sxl in male and female flies Pe: promoter for establishment Pm: promoter for maintenance

24. Fig 17-28 a cascade of alternative splicing events determines the sex of a fly. Sxl: splicing repressor Tra: splicing activator

25. Expression of the yeast transcriptional factor Gcn4 is controlled at the level of translation Fig 17-29 translational control of Gcn4 in response to amino acid starvation

26. RNAs in gene regulation RNA interference (RNAi): short RNAs that can repress the expression of those genes with sequence homology to these RNAs. Translational inhibition of that mRNA Destruction of that mRNA Silencing the promoter that directs expression of that mRNA

27. Double-stranded RNA (dsRNA) inhibits expression of genes homologous to that RNA In C. elegans and in plants

28. Short interfering RNAs (siRNAs) are produced from dsRNAs and direct machinery that switches off genes in various ways. RISC: RNA-induced silencing complex

29. Fig 17-30 RNAi silencing

30. MicroRNAs (miRNAs) control the expression of some genes during development miRNAs: 21-22 nts long; arise from large precursors (about 70-90 nts long) transcribed from non-protein encoding sequences. These transcripts contain sequences that form stem loop structures, which are processed by Dicer (or DCL1, Dicer-like 1, in plants). miRNAs lead to the destruction (in plants) or translational repression (in worms) of target mRNAs with homology to the miRNAs.