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Related Concept Videos

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Base complementarity between the three base pairs of mRNA codon and the tRNA anticodon is not a failsafe mechanism. Inaccuracies can range from a single mismatch to no correct base pairing at all. The free energy difference between the correct and nearly correct base pairs can be as small as 3 kcal/ mol. With complementarity being the only proofreading step, the estimated error frequency would be one wrong amino acid in every 100 amino acids incorporated. However, error frequencies observed in...
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During most eukaryotic translation processes, the small 40S ribosome subunit scans an mRNA from its 5' end until it encounters the first start AUG codon. The large 60S ribosomal subunit then joins the smaller one to initiate protein synthesis. The location of the translation initiation is largely determined by the nucleotides near the start codon as there may be multiple translation initiation sites present on the mRNA.  Marilyn Kozak discovered that the sequence RCCAUGG (where R...
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The large ribosomal subunit has several important structures essential to translation. These include the peptidyl transferase center (PTC) - which is the site where the peptide bond is formed - and a large, internal, water-filled tube through which the nascent polypeptide moves. This latter structure is called the Peptide Exit Tunnel, and it begins at the PTC and spans the body of the large ribosomal subunit. During translation, as the nascent polypeptide chain is synthesized, it passes through...
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Initiating translation is complex because it involves multiple molecules. Initiator tRNA, ribosomal subunits, and eukaryotic initiation factors (eIFs) are all required to assemble on the initiation codon of mRNA. This process consists of several steps that are mediated by different eIFs.
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Translational regulation in prokaryotes ensures efficient protein synthesis by controlling ribosome access to mRNA. This regulation is mediated by secondary RNA structures, including translational riboswitches, RNA thermometers, and small RNAs (sRNAs), which respond to intracellular and environmental signals to modulate gene expression.Translational RiboswitchesRiboswitches in the leader region of mRNAs can regulate translation by altering the accessibility of the Shine-Dalgarno (SD) sequence,...
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Translational buffering by ribosome stalling in upstream open reading frames.

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Upstream open reading frames (uORFs) can buffer protein expression by causing ribosome stalls. These stalls prevent decreases in main protein output, acting as a protective mechanism during cellular stress.

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Area of Science:

  • Molecular Biology
  • Gene Regulation
  • Translational Control

Background:

  • Upstream open reading frames (uORFs) are prevalent in human mRNAs, influencing downstream translation.
  • Mechanisms of uORF-mediated translational regulation include ribosome siphoning, re-initiation modulation, and ribosome interactions.
  • The precise impact of these mechanisms on protein expression regulation is not fully understood.

Purpose of the Study:

  • To investigate uORF-mediated translational regulation using the cytomegaloviral UL4 mRNA as a model.
  • To test alternative models of how uORFs control downstream open reading frame (ORF) translation in human cells.

Main Methods:

  • Systematic measurements of the UL4 mRNA 5' untranslated region (UTR) in human cells.
  • Analysis of ribosome stalling and its effect on protein expression.
  • Computational kinetic modeling to understand ribosome-stalled interactions.

Main Results:

  • A terminal diproline-dependent ribosome stall in the UL4 uORF buffers main ORF protein expression against reduced ribosome loading.
  • This buffering effect is independent of the ribosome stall's position within the uORF.
  • Computational modeling indicates scanning ribosomes dissociate upon encountering stalled ribosomes, rather than queuing.

Conclusions:

  • Ribosome stalls within uORFs, particularly those involving diproline motifs, serve as a general mechanism to buffer main ORF translation.
  • This buffering is crucial for maintaining protein expression during cellular stress and developmental changes.
  • Identified human uORFs with similar diproline motifs that repress main ORF expression, suggesting a conserved regulatory strategy.