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Transcript
Chapter 13 The Genetic Code and Transcription Lecture Presentation by Dr. Cindy Malone, California State University Northridge © 2015 Pearson Education, Inc. Section 13.1: The Genetic Code The genetic code: General features – Written in linear form using ribonucleotide bases that compose mRNA – Each “word” consists of three ribonucleotide letters, or a triplet code Codon: Every three ribonucleotides – Unambiguous – each triplet specifies only one amino acid continued © 2015 Pearson Education, Inc. Section 13.1: The Genetic Code continued The genetic code – Degenerate: A given amino acid can be specified by more than one triplet codon – Contains “start” and “stop” signals: triplets that initiate and terminate translation © 2015 Pearson Education, Inc. Section 13.1: The Genetic Code continued – Nonoverlapping: Any single ribonucleotide within mRNA is part of one triplet – Nearly universal: A single coding dictionary is used by viruses, prokaryotes, archaea, and eukaryotes © 2015 Pearson Education, Inc. Section 13.2: Operational Patterns mRNA – messenger RNA – Serves as intermediate in transferring genetic information from DNA to proteins – Genetic information is stored in DNA – Code that translates it to protein is in RNA © 2015 Pearson Education, Inc. Section 13.2: The Triplet Code Triplet code – Provides 64 codons to specify 20 amino acids © 2015 Pearson Education, Inc. Section 13.2: Frameshift Mutations Reading frame – Contiguous sequence of nucleotides – Insertions or deletions shift reading frame and change codons downstream frameshift mutation Triplet nature code was revealed by frameshift mutations (Figure 13-2) © 2015 Pearson Education, Inc. © 2015 Pearson Education, Inc. Figure 13-2 © 2015 Pearson Education, Inc. Figure 13-2a © 2015 Pearson Education, Inc. Figure 13-2b Section 13.3: Triplet Binding Assay Triplet binding assay – Developed by Nirenberg and Leder to determine other specific codon assignments – Ribosomes bind to single codon of three nucleotides – Complementary amino acid-charged tRNA can bind (Figure 13-5 and Table 13.2) © 2015 Pearson Education, Inc. © 2015 Pearson Education, Inc. Table 13.2 Section 13.3: Repeating Copolymers Chemically synthesized long RNAs – Short repeating sequences enzymatically joined short sequences together, which made long RNAs – Used for in vitro translation to determine more codon assignments – Figure 13-6 and Table 13.3 © 2015 Pearson Education, Inc. Section 13.4: Degeneracy of the Genetic Code The genetic code is degenerate – Many amino acids specified by more than one codon – Only tryptophan and methionine are encoded by single codon (Figure 13-7) – Genetic code shows order: chemically similar amino acids share one or two middle bases in triplets encoding them © 2015 Pearson Education, Inc. © 2015 Pearson Education, Inc. Figure 13-7 Section 13.4: The Wobble Hypothesis The wobble hypothesis – The initial two ribonucleotides of triplet codes are often more critical than the third – Third position Less spatially constrained Need not adhere as strictly to established base-pairing rules (Table 13.4) © 2015 Pearson Education, Inc. © 2015 Pearson Education, Inc. Table 13.4 Section 13.4: Initiation Methionine (AUG) – initiator codon – Initial amino acid incorporated into all proteins – In bacteria: modified form of methionine N-formylmethionine (fmet) AUG: Only codon to encode methionine – Appears internally in mRNA unformylated © 2015 Pearson Education, Inc. Section 13.4: Termination Termination codons: UAG, UAA, UGA – Do not code for any amino acid – Are not recognized by tRNA – Translation terminates when these codons are encountered © 2015 Pearson Education, Inc. Section 13.4: Nonsense Mutations Nonsense mutations – Mutations that produce a stop codon internally in gene – Translation is terminated – Partial polypeptide is produced © 2015 Pearson Education, Inc. Section 13.7: Overlapping Genes Overlapping genes – Single mRNA has multiple initiation points – Creates different reading frames – Specifies more than one polypeptide © 2015 Pearson Education, Inc. Section 13.7: Open Reading Frames ORF: Open reading frame (overlapping genes) – DNA sequence produces RNA with start and stop – Series of triplet codons specify amino acids to make polypeptide In some viruses, initiation at different AUG positions out of frame with another leads to distinct polypeptides (Figure 13-8) © 2015 Pearson Education, Inc. © 2015 Pearson Education, Inc. Figure 13-8 © 2015 Pearson Education, Inc. Figure 13-8a Section 13.8: Transcription Transcription – RNA synthesized on DNA template – Genetic information stored in DNA is transferred to RNA – Serves as intermediate molecule between DNA and proteins – Each triplet codon is complementary to anticodon of tRNA © 2015 Pearson Education, Inc. VIDEO - TRANSCRIPCION © 2015 Pearson Education, Inc. Section 13.10: RNA Polymerase RNA polymerase – Enzyme directs synthesis of RNA using DNA template – Nucleotides contain ribose, not deoxyribose – No primer required for initiation n(NTP) DNA RNA polymerase © 2015 Pearson Education, Inc. (NMP)n + n(PPi) Section 13.10: Promoters Transcription results in ssRNA – Template strand is transcribed – Transcription begins with template binding by RNA polymerase at promoter (Figure 13-9) – Promoters: Specific DNA sequences in 5 region upstream of initial transcription point – subunit responsible for promoter recognition (initiation of transcription) © 2015 Pearson Education, Inc. VIDEO https://www.youtube.com/watch?v=SMtWvDbfHL o&nohtml5=False © 2015 Pearson Education, Inc. © 2015 Pearson Education, Inc. Figure 13-9 © 2015 Pearson Education, Inc. Figure 13-9a © 2015 Pearson Education, Inc. Figure 13-9b © 2015 Pearson Education, Inc. Figure 13-9c Section 13.10: Transcription Start Site Transcription start site – DNA double helix is denatured: unwound to make template strand accessible for RNA polymerase – Interaction of promoters and RNA polymerase regulates efficiency of transcription © 2015 Pearson Education, Inc. Section 13.10: Consensus Sequences Consensus sequences – DNA sequences homologous in different genes of same organism E. coli promoters have two consensus sequences – TTGACA and TATAAT (Pribnow box) – Positioned at 35 and 10 with respect to the transcription initiation site © 2015 Pearson Education, Inc. Section 13.10: Chain Elongation Chain elongation – Ribosomes are added to RNA chain – subunit dissociates from holoenzyme – Elongation proceeds under direction of core enzyme (Figure 13-9c) © 2015 Pearson Education, Inc. © 2015 Pearson Education, Inc. Figure 13-9 Section 13.10: Termination Termination – Enzyme traverses entire gene until a termination nucleotide sequence is encountered – In bacteria: Termination transcribed into RNA causes newly formed transcript to fold back on itself (hairpin) – At times, termination depends on the rho () termination factor © 2015 Pearson Education, Inc. 13.11 Transcription in Eukaryotes Differs from Prokaryotic Transcription in Several Ways © 2015 Pearson Education, Inc. Section 13.11: Eukaryotic Transcription Transcription in eukaryotes – Occurs within nucleus (unlike prokaryotes) – mRNA must leave nucleus for translation – Chromatin remodeling: Chromatin must uncoil to make DNA accessible to RNA Pol – RNA polymerases rely on transcription factors (TFs) to scan/bind DNA – Enhancers and silencers control transcription regulation © 2015 Pearson Education, Inc. Section 13.11: RNA Polymerases Eukaryotes possess three forms of RNA polymerase – Each transcribes different types of genes (Table 13.7) – RNA Pol I – RNA Pol II – RNA Pol III © 2015 Pearson Education, Inc. © 2015 Pearson Education, Inc. Table 13.7 Section 13.11: RNA Pol II RNA polymerase II (RNAP II) – Responsible for transcription of wide range of genes in eukaryotes – Activity of RNAP II is dependent on cis-acting elements and trans-acting transcription factors – RNAP II core-promoter determines where RNAP II binds to DNA © 2015 Pearson Education, Inc. Section 13.11: TATA Box Regulatory sequences influence efficiency of transcription initiation by RNAP II – Proximal-promoter elements – Enhancers – Silencers TATA box – Core-promoter element – Binds TATA-binding protein (TBP) of transcription factor TFIID: determines start transcription start site © 2015 Pearson Education, Inc. Section 13.11: Enhancers and Silencers Enhancers and silencers – Found upstream, within, or downstream of gene – Enhancers increase transcription levels; silencers decrease them – Modulate transcription from a distance – Act to increase or decrease transcription in response to cell’s requirement for gene product © 2015 Pearson Education, Inc. Section 13.11: Transcription Factors Transcription factors facilitate RNAP II binding and initiation of transcription – General transcription factors: Required for all RNAP II-mediated transcription – Transcription activators and repressors: Influence efficiency or rate of RNAP II transcription initiation © 2015 Pearson Education, Inc. Section 13.11: Cap and Tail Eukaryotic mRNAs require processing to produce mature mRNAs Posttranscriptional modifications – Addition of 5 cap (7-mG cap) – Addition of 3 tail (poly-A tail) – Excision of introns (Figure 13-10) © 2015 Pearson Education, Inc. © 2015 Pearson Education, Inc. Figure 13-10 13.12 The Coding Regions of Eukaryotic Genes Are Interrupted by Intervening Sequences © 2015 Pearson Education, Inc. Section 13.12: Introns and Exons Introns (intervening sequences) – Regions of initial RNA transcript not expressed in amino acid sequence of protein – DNA sequences not represented in final mRNA product – Exons are sequence retained and expressed – Prokaryotes do not have introns – Heteroduplexes: Introns present in DNA but not mRNA loop out (Figure 13-11) © 2015 Pearson Education, Inc. © 2015 Pearson Education, Inc. Figure 13-11 Section 13.12: Splicing Posttranscriptional modification: Splicing – Introns are removed by splicing – Exons are then joined together in mature mRNA – Mature mRNA is smaller than initial RNA (Figure 13-12 and Table 13.8) © 2015 Pearson Education, Inc. © 2015 Pearson Education, Inc. Figure 13-12 Section 13.12: Self-Splicing RNAs and Spliceosome Self-Splicing RNAs – Self-excision group I introns occurs in bacteria, lower eukaryotes, and higher plants (Figure 13-13d) Spliceosome – Pre-mRNA introns spliced out by spliceosome – Reaction involves formation of lariat structure (Figure 13-14) © 2015 Pearson Education, Inc. © 2015 Pearson Education, Inc. Figure 13-13 © 2015 Pearson Education, Inc. Figure 13-14 © 2015 Pearson Education, Inc. Figure 13-15