Lecture Connections 26 | RNA Metabolism © 2009 W H Freeman and Company CHAPTER 26 RNA Metabolism Key topics: – Transcription: DNA-dependent synthesis of RNA – Capping and splicing: RNA processing Overview of RNA Function • Ribonucleic acids play three well-understood roles in living cells – Messenger RNAs encode the amino acid sequences of all the polypeptides found in the cell – Transfer RNAs match specific amino acids to triplet codons in mRNA during protein synthesis – Ribosomal RNAs are the constituents and catalytic appropriate amino acid • Ribonucleic acids play several less-understood functions in eukaryotic cells – Micro RNA appears to regulate the expression of genes, possibly via binding to specific nucleotide sequences • Ribonucleic acids act as genomic material in viruses Overview of RNA Metabolism • Ribonucleic acids are synthesized in cells using DNA as a template in a process called the transcription – Transcription is tightly regulated in order to control the concentration of each protein in the cell at optimal level • Being mainly single stranded, many RNA molecules can fold into compact structures with specific functions – Some RNA molecules can act as catalysts (ribozymes), often using metal ions as cofactors • Most eukaryotic ribonucleic acids are processed after synthesis – Elimination of introns; joining of exons – Poly-adenylation of the 3’ end – Capping the 5’ end Transcription in E coli • The nucleoside triphosphates add to the the 3’ end of the growing RNA strand • The growing chain is complementary to the template strand in DNA • The synthesis is catalyzed by enzyme (RNA polymerase) • RNA polymerase covers about 35 bp-long segment of DNA Replication vs Transcription • Both add nucleotides via an attack of the 3’ hydroxyl of the growing chain to -phosphorus of nucleoside triphosphates – RNA synthesis requires ribonucleoside triphosphates – RNA synthesis pairs A with U instead of dA with dT • Both require catalysis by a Mg++-dependent enzyme – RNA synthesis has lower fidelity – RNA synthesis does not require a primer for initiation • Both require a single strand of DNA as molecular template for building the new strand Both DNA Strands may Encode for Proteins • Adenovirus is one of the causative agents of common cold • Adenovirus has a linear genome • Each strand encodes for a number of proteins Capping the 5’ of mRNA • Capping protects mRNA from 5’exonuclease degradation Capping Enzymes Are Tethered to the C-terminal Domain of Polymerase II Four Major Groups of Introns • Spliceosomal introns are spliced by splicesomes – These are most common introns – Frequent in protein-coding regions of eukaryotic genomes • Group I and Group II introns are self splicing – Interrupt mRNA, tRNA and rRNA genes – Found within nuclear, mitochondrial, and chloroplast genomes – Common in fungi, algae, and plants, also found in bacteria – Group I and Group II differ mainly by the splicing mechanism • tRNA introns are spliced by protein-based enzymes – Found in certain tRNAs in eukaryotes and archae – Primary transcript cleaved by endonuclease – Exons are joined by ATP-dependent ligase Splicing of Group I Introns Transesterification in Splicing of Group I and Group II Introns • In case of group I intron splicing, the nucleophile is 3’ hydroxyl of free guanosine, GMP, GDP, or GTP • In case of group II intron splicing, the nucleophile is an hydroxyl in RNA Splicing of Group II Introns Overview of RNA Processing Chapter 26: Summary In this chapter, we learned that: • RNA polymerase synthesizes RNA using a strand of DNA as a template and nucleoside triphosphates as substrates • The primary RNA transcript in eukaryotes requres processing before it becomes messanger RNA • The processing involves capping 5’end with methylguanosine to stabilize the RNA molecule • The processing involves splicing out introns • Some introns have an amazing ability to carry out their own splicing ...CHAPTER 26 RNA Metabolism Key topics: – Transcription: DNA-dependent synthesis of RNA – Capping and splicing: RNA processing Overview of RNA Function • Ribonucleic acids... rRNAs) • RNA polymerase II is responsible for synthesis of mRNA – Very fast (500 – 1000 nucleotides / sec) – Specifically inhibited by mushroom toxin -amanitin • RNA polymerase III makes tRNAs... some small RNA products • Plants appear to have RNA polymerase IV that is responsible for the synthesis of small interfering RNAs • Mitochondria have their own RNA polymerase Eukaryotic mRNA Transcription