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Related Concept Videos

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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 flow of genetic information in cells from DNA to mRNA to protein is described by the central dogma, which states that genes specify the sequence of mRNAs, which in turn specify the sequence of amino acids making up all proteins. The decoding of one molecule to another is performed by specific proteins and RNAs. Because the information stored in DNA is so central to cellular function, it makes intuitive sense that the cell would make mRNA copies of this information for protein synthesis...
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Ribosome profiling or ribo-sequencing is a deep sequencing technique that produces a snapshot of active translation in a cell. It selectively sequences the mRNAs protected by ribosomes to get an insight into a cell’s translation landscape at any given point in time.
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Probing RNA Structure with Dimethyl Sulfate Mutational Profiling with Sequencing In Vitro and in Cells
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RNA Structural Determinants of Optimal Codons Revealed by MAGE-Seq.

Eric D Kelsic1, Hattie Chung2, Niv Cohen3

  • 1Department of Systems Biology, Harvard Medical School, Boston, MA 02115, USA; Wyss Institute for Biologically Inspired Engineering, Harvard Medical School, Boston, MA 02115, USA.

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Summary

Codon choices throughout genes impact RNA structure and function. Even distant codons can disrupt essential RNA conformations, influencing gene expression and fitness.

Keywords:
RNA structurecodoncodon optimizationcodon usagecomputational biologymolecular biologysynthetic biologysystems biology

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Area of Science:

  • Molecular Biology
  • Genetics
  • Bioinformatics

Background:

  • Synonymous codon usage influences gene expression, particularly at the start of genes by optimizing 5' RNA structures for translation initiation.
  • Mechanisms governing codon optimization downstream within a gene remain less understood.

Purpose of the Study:

  • To investigate the determinants of codon choices across an entire gene.
  • To understand how codon mutations affect gene fitness through RNA structural changes.

Main Methods:

  • Generated 12,726 in situ codon mutants in the Escherichia coli essential gene infA.
  • Utilized multiplex automated genome engineering mutagenesis with amplicon deep sequencing (MAGE-seq) to measure fitness.
  • Correlated predicted 5' RNA structures with measured fitness data.

Main Results:

  • Codons distant from the start codon can be deleterious if they disrupt native 5' RNA conformation.
  • Long-range RNA structural interactions impose context-dependent rules on codon selection, extending beyond intrinsic codon preferences.
  • Natural codon distributions are optimized to prevent disruption of structures near the 5' untranslated region (UTR).

Conclusions:

  • Codon optimization is influenced by long-range RNA structural interactions, not just local effects or intrinsic codon biases.
  • Understanding these context-dependent rules is crucial for predicting gene expression and for synthetic biology applications.
  • Findings provide insights into natural codon distributions and offer a basis for improving engineered gene expression systems.