Chapter 7 Messenger RNA

Figure 7.01: Protein synthesis uses three types of RNA. 7.1 Introduction All three types of RNA are central players during the process of gene expression. Figure 7.01: Protein synthesis uses three types of RNA.

7.2 mRNA Is Produced by Transcription and Is Translated Within a gene, only one of the two strands of DNA is transcribed into RNA. Figure 7.02: Gene expression = transcription + translation.

7.3 The Secondary Structure of Transfer RNA Is a Cloverleaf A tRNA has a sequence of 74 to 95 bases that folds into a cloverleaf secondary structure with four constant arms (and an additional arm in the longer tRNAs). tRNA is charged to form aminoacyl-tRNA by forming an ester link from the 2' or 3' OH group of the adenylic acid at the end of the acceptor arm to the COOH group of the amino acid.

7.3 The Secondary Structure of Transfer RNA Is a Cloverleaf Figure 7.03: tRNA is an adaptor.

7.3 The Secondary Structure of Transfer RNA Is a Cloverleaf The sequence of the anticodon is solely responsible for the specificity of the aminoacyl-tRNA during translation.

7.3 The Secondary Structure of Transfer RNA Is a Cloverleaf Figure 7.05: The anticodon determines tRNA specificity.

7.4 The Acceptor Stem and Anticodon Are at Opposite Ends of the tRNA Tertiary Structure The cloverleaf forms an L-shaped tertiary structure with the acceptor arm at one end and the anticodon arm at the other end.

Figure 7.06: All tRNAs share a tertiary structure. 7.4 The Acceptor Stem and Anticodon Are at Opposite Ends of the tRNA Tertiary Structure Figure 7.06: All tRNAs share a tertiary structure.

7.5 Messenger RNA Is Translated by Ribosomes Ribosomes are characterized by their rate of sedimentation. 70S for bacterial ribosomes and 80S for eukaryotic ribosomes. A ribosome consists of a: large subunit (50S or 60S for bacteria and eukaryotes) small subunit (30S or 40S) The ribosome provides the environment in which aminoacyl-tRNAs add amino acids to the growing polypeptide chain in response to the corresponding triplet codons. A ribosome moves along an mRNA from 5' to 3'.

7.5 Messenger RNA Is Translated by Ribosomes Figure 7.08: Ribosomes dissociate into subunits.

7.6 Many Ribosomes Can Bind to One mRNA An mRNA is simultaneously translated by several ribosomes. Each ribosome is at a different stage of progression along the mRNA. Figure 7.09: Each ribosome has a polypeptidyl-tRNA and an aminoacyl-tRNA.

Figure 7.10: Hemoglobin is synthesized on pentasomes. Photo courtesy of Alexander Rich, Massachusetts Institute of Technology

Figure 7.11: Ribosomes recycle for translation.

Figure 7.12: ~30% of bacterial dry mass is concerned with gene expression.

7.7 The Cycle of Bacterial Messenger RNA Transcription and translation occur simultaneously in bacteria. Ribosomes begin translating an mRNA before its synthesis has been completed. Bacterial mRNA is unstable and has a half-life of only a few minutes.

7.7 The Cycle of Bacterial Messenger RNA Figure 7.13: Trancription - translation - degradation.

7.7 The Cycle of Bacterial Messenger RNA A bacterial mRNA may be polycistronic in having several coding regions that represent different genes. Figure 7.15: Bacterial mRNA is polycistronic.

7.8 Eukaryotic mRNA Is Modified During or after Its Transcription A eukaryotic mRNA transcript is modified in the nucleus during or shortly after transcription. The modifications include the addition of a methylated cap at the 5' end and a sequence of poly(A) at the 3' end.

7.8 Eukaryotic mRNA Is Modified During or after Its Transcription Figure 7.16: Eukaryotic mRNA is modified at both ends.

7.8 Eukaryotic mRNA Is Modified During or after Its Transcription The mRNA is exported from the nucleus to the cytoplasm only after all modifications have been completed.

7.8 Eukaryotic mRNA Is Modified During or after Its Transcription Figure 7.17: Eukaryotic mRNA is modified and exported.

7.9 The 5’ End of Eukaryotic mRNA Is Capped A 5' cap is formed by adding a G to the terminal base of the transcript via a 5'–5' link. One to three methyl groups are added to the base or ribose of the new terminal guanosine.

7.9 The 5’ End of Eukaryotic mRNA Is Capped Figure 7.18: Eukaryotic mRNA has a methylated 5' cap.

7.10 The 3’ Terminus of Eukaryotic mRNA Is Polyadenylated A length of poly(A) ~200 nucleotides long is added to a nuclear transcript after transcription. The poly(A) is bound by a specific protein (PABP). The poly(A) stabilizes the mRNA against degradation.

7.11 Bacterial mRNA Degradation Involves Multiple Enzymes The overall direction of degradation of bacterial mRNA is 5'–3'. Degradation results from the combination of endonucleolytic cleavages followed by exonucleolytic degradation of the fragment from 3'→5'.

7.11 Bacterial mRNA Degradation Involves Multiple Enzymes Figure 7.19: mRNA is degraded by exo- and endo- nucleases.

7.12 Two Pathways Degrade Eukaryotic mRNA The modifications at both ends of mRNA protect it against degradation by exonucleases. Specific sequences within an mRNA may have stabilizing or destabilizing effects. Destabilization may be triggered by loss of poly(A).

7.12 Two Pathways Degrade Eukaryotic mRNA Figure 7.20: The structure and sequence of eukaryotic mRNA determine stability.

Figure 7.21: An ARE in a 3’ nontranslated region initiates degradation of mRNA.

7.12 Two Pathways Degrade Eukaryotic mRNA Degradation of yeast mRNA requires removal of the 5' cap and the 3' poly(A). Figure 7.22: Deadenylation allows decappaing to occur, which leads endonucleolytic cleavage from the 5’ end.

7.12 Two Pathways Degrade Eukaryotic mRNA One yeast pathway involves exonucleolytic degradation from 5'→3'. Another yeast pathway uses a complex of several exonucleases that work in the 3'→5' direction. The deadenylase of animal cells may bind directly to the 5' cap. Either mutation causes slower degradation of mRNA, but loss of both pathways is lethal in yeast.

Figure 7.23: The 3'-5' pathway has three stages.

7.13 Nonsense Mutations Trigger a Surveillance System Nonsense mutations cause mRNA to be degraded. Genes coding for the degradation system have been found in yeast and worms.

7.13 Nonsense Mutations Trigger a Surveillance System Figure 7.24: A surveillance system degrades mutant mRNA.

Figure 7.25: Splicing junctions are marked by proteins.

7.14 Eukaryotic RNAs Are Transported RNA is transported through a membrane as a ribonucleoprotein particle. All eukaryotic RNAs that function in the cytoplasm must be exported from the nucleus. tRNAs and the RNA component of a ribonuclease are imported into mitochondria. mRNAs can travel long distances between plant cells.

7.14 Eukaryotic RNAs Are Transported Figure 7.26: Eukaryotic RNA can be transported between cell compartments.

7.15 mRNA Can Be Localized Within A Cell Yeast ASH1 mRNA forms a ribonucleoprotein that binds to a myosin motor. A motor transports it along actin filaments into the daughter bud. It is anchored and translated in the bud, so that the protein is found only in the bud.

7.15 mRNA Can Be Localized Within A Cell Figure 7.27: ASH1 mRNA is connected to a motor.