Translational Regulation

• mRNAs lacking the 5' cap have an IRES upstream of the 5' start codon
• internal ribosome entry site = IRES
• The IRES recruits initiation factors and the small ribosomal subunit to begin translation
• Even IFs that have been cleaved by viruses can bind to the IRES site
• IRES enables co-expression of several genes under the same promoter
• Even though we can translate mRNA of interest and reporter mRNA together, the reporter protein may impact the native structure or function of the protein of interest
• Experimental application of IRES:

  • The mRNA of interest will be translated normally by initiation at the 5' cap, whereas translation of the reporter mRNA will begin at the IRES
• Nonsense-mediated mRNA decay: A process that detects and destroys transcripts with a nonsense mutation
• nonsense mutation = a premature stop codon
• During mRNA splicing, the site of each removed intron is marked by an EJC
• exon junction complex = EJC
• The ribosome removes EJCs as it moves along the mRNA
•  Nonsense-mediated mRNA decay

  1. The ribosome is released early and some EJCs remain on the mRNA
  2. EJCs recruit proteins that cleave the 5' cap, making the mRNA vulnerable to degradation by ribonucleases
• Non-stop mRNA decay: A process that detects and destroys transcripts that don't have a stop codon
• Without a stop codon, translation proceeds through the 3' poly(A) tail, resulting in a poly(lysine) tail
• Non-stop mRNA decay

  1. Translation proceeds through the 3' poly(A) tail encoding a poly(lysine) tail
  2. This causes the ribosome to stall and recruit a protein
  3. The protein triggers ribosome disassociation and mRNA degradation by a 3' to 5' ribonuclease
  4. The defective polypeptide is degraded by a protease that recognises the C-terminal poly(lysine) tag
• No-go mRNA decay: A process that destroys mRNAs containing ribosomes that have stalled before the stop codon is reached
• Possible causes of ribosome stalling:

  1. Rare conditions in which a rare codon is encountered and there is no tRNA to match
  2. Secondary structures are formed, either internal base pairing of one mRNA molecule or between two mRNA molecules
• No-go mRNA decay:

  The stalled ribosome recruits factors that promote ribosome disassociation, mRNA cleavage and mRNA degradation
• Small RNAs regulate mRNA stability
• RNA interference: Process of mRNA degradation induced by double-stranded RNA
• Exogenous dsDNA can be from viruses or experimentally introduced
• Regulation by exogenous dsDNA

  1. Exogenous dsRNA is cleaved by Dicer into siRNA
  2. siRNA binds to proteins to form RISC
  3. RISC binds to target mRNA to stop translation and/or cause mRNA degradation
• Dicer is a ribonuclease
• RNA induced silencing complex = RISC
• small interfering RNA = siRNA
• Endogenous regulation by miRNAs

  1. miRNAs are generated from larger transcripts made by RNA polymerase I or RNA polymerase III
  2. miRNAs are cleaved into dsDNA
  3. This dsDNA is further processed into single stranded RNA which binds to proteins to form RISC
  4. RISC binds to target mRNA to stop translation and/or cause mRNA degradation
• MicroRNAs = miRNAs
• Transferrin receptor: Binds the iron-transferrin complex to become endocytosed and enable entry into cells
• Transferrin: Carries iron in the blood
• Ferritin: Stores iron in the liver and kidney
• Ferritin and transferrin receptor levels are reciprocally related
• The rate of transcription of ferritin and transferrin receptors does not change, but is instead regulated during translation
• Ferritin mRNA has a stem-loop called the iron response element (IRE) in the 5' untranslated region which binds to the IRE binding protein
• When [iron] is low, the IRE binding protein binds to ferritin mRNA and blocks the initiation of translation
• When [iron] is high, the IRE binding protein binds to iron and cannot bind to ferritin mRNA, allowing ferritin to be translated and store excess iron
• Transferrin receptor mRNA has several IRE regions in the 3' untranslated region which does not interfere with translation if IRE binding protein binds
• When [iron] is low, IRE binding protein binds to transferrin receptor mRNA and translation proceeds as normal
• When [iron] is high, IRE binding protein binds to iron and detaches from the transferrin receptor mRNA, leaving the mRNA vulnerable to degradation and unable to be translated