Conditional Switch between Frameshifting Regimes upon Translation of dnaX mRNA

Area of Science:

• Molecular Biology
• Bacterial Genetics
• Protein Synthesis

Background:

• Ribosome frameshifting is a key mechanism in bacterial gene expression, allowing for the synthesis of multiple protein isoforms from a single mRNA.
• The bacterial dnaX gene is known to undergo programmed -1 ribosomal frameshifting, producing both full-length and truncated products.

Purpose of the Study:

• To elucidate the distinct molecular mechanisms and conditions governing different ribosome frameshifting routes during bacterial dnaX translation.
• To investigate the role of aminoacyl-tRNA availability and mRNA structure in directing frameshifting pathways.

Main Methods:

• Kinetic experiments were employed to analyze the dynamics of tRNA translocation and frameshifting events.
• Quantitative mass spectrometry was used to identify and quantify the polypeptides produced in vivo under different conditions.

Main Results:

• -1 frameshifting predominantly occurs via a stem-loop dependent pathway when aminoacyl-tRNAs are abundant, involving simultaneous translocation of two tRNAs.
• Under aminoacyl-tRNA limitation, ribosomes switch to a stem-loop independent pathway initiated by stalling at a vacant codon, leading to peptidyl-tRNA slippage.
• A -2 frameshifted product, identified via mass spectrometry, is synthesized in vivo, particularly when the -1 frameshift aminoacyl-tRNA is absent.

Conclusions:

• Ribosome frameshifting in bacterial dnaX is a regulated process that dynamically switches between distinct pathways based on cellular conditions, specifically aminoacyl-tRNA availability.
• This pathway switching mechanism allows for the adaptation of gene expression levels in response to nutrient or tRNA limitations.
• The findings provide new insights into the intricate regulation of bacterial protein synthesis and gene expression plasticity.