Translation at higher than an optimal level interferes with coupling at an intercistronic junction

Area of Science:

• Bacteriophage genetics
• Molecular biology
• Gene regulation

Background:

• Bacterial polycistronic mRNAs typically couple translation of upstream genes to downstream ones.
• Efficient coupling ensures similar protein levels, but variations exist.
• The filamentous phage IKe gene V and gene VII pair shows unusually low translational coupling (1%).

Purpose of the Study:

• To characterize the inefficient translational coupling between phage IKe gene V and gene VII.
• To identify the genetic region responsible for this inefficient coupling.
• To elucidate the mechanism by which upstream translation affects downstream gene expression.

Main Methods:

• Utilized chimeric sequences between phage IKe and f1 to map the coupling region.
• Manipulated gene V translation rates (elongation, amber codon suppression).
• Altered the Shine-Dalgarno interaction strength for gene VII initiation.

Main Results:

• The region responsible for inefficient coupling mapped upstream of the intercistronic region, associated with gene V.
• Decreasing gene V translation (elongation rate, suppression efficiency) increased coupling efficiency.
• Enhancing the Shine-Dalgarno interaction at gene VII significantly boosted coupling.

Conclusions:

• Upstream translation acts as a negative factor, limiting downstream gene expression in the IKe phage system.
• Excessive upstream translation interferes with translational coupling at the intercistronic junction.
• This finding offers a novel mechanism for regulating gene expression in polycistronic operons.