On the Spatial Organization of mRNA, Plasmids, and Ribosomes in a Bacterial Host Overexpressing Membrane Proteins

Area of Science:

• Microbiology
• Molecular Biology
• Biotechnology

Background:

• Heterologous membrane protein production in microbial hosts like Lactococcus lactis is crucial for biotechnology.
• Aberrant protein biogenesis can lead to cellular stress and reduced production yields.

Purpose of the Study:

• To investigate the real-time, single-cell dynamics of transcription, translation, and growth defects during heterologous membrane protein expression.
• To understand the mechanisms underlying failed membrane protein biogenesis and identify strategies for optimization.

Main Methods:

• Time-resolved single-cell fluorescence imaging in Lactococcus lactis.
• Monitoring mRNA localization, translation activity, and cell division.
• Analyzing the role of protein domains in membrane targeting.

Main Results:

• Accumulation of transcripts for poorly produced membrane proteins in polar mRNA-dense bodies.
• Polar mRNA foci correlate with cell division cessation and heat-shock response activation.
• Successful membrane protein localization depends on a native-like N-terminal domain for Signal Recognition Particle (SRP)-dependent targeting.

Conclusions:

• Aberrant membrane protein biogenesis disrupts cellular processes, including transcription, translation, and cell division.
• Proper folding and native-like domains are critical for SRP-dependent membrane targeting and efficient production.
• These findings provide insights for optimizing heterologous membrane protein production strategies.