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Identification of Growth Inhibition Phenotypes Induced by Expression of Bacterial Type III Effectors in Yeast
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Medium dependent factors govern the functionality of engineered type III secretion systems.

SangKu Yi1, Beom Seok Kim1, Eunna Choi2

  • 1School of Life Sciences, BK21 FOUR KNU Creative BioResearch Group, Kyungpook National University, Daegu, 41566, Republic of Korea.

Journal of Biological Engineering
|December 16, 2025
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Summary

Bacterial culture conditions significantly impact engineered type III secretion systems (T3SS). Growth medium affects T3SS performance, influencing protein secretion efficiency and assembly, crucial for synthetic biology applications.

Keywords:
Resource allocationTranslational burdenType III secretion system

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Area of Science:

  • Synthetic Biology
  • Microbiology
  • Molecular Biology

Background:

  • The type III secretion system (T3SS) is a bacterial machine for injecting effector proteins.
  • T3SS engineering for biotechnological applications is promising.
  • The effect of bacterial host culture conditions on T3SS performance is largely unstudied.

Purpose of the Study:

  • To investigate how bacterial host culture conditions influence engineered T3SS performance.
  • To characterize an IPTG-inducible, refactored T3SS (iT3SS) circuit in Salmonella enterica.
  • To determine the impact of different growth media on iT3SS functionality.

Main Methods:

  • Constructed an IPTG-inducible T3SS circuit (iT3SS) using a PrgH-GFP fusion in Salmonella.
  • Assessed PrgH-GFP expression dynamics and SptP effector secretion in LB and glucose minimal media.
  • Utilized RNA-sequencing (RNA-seq) to analyze gene expression under varying IPTG concentrations and media conditions.

Main Results:

  • Secretion efficiency was maintained in glucose minimal medium but not in rich LB medium across IPTG concentrations.
  • In LB medium, secretion and invasion efficiencies did not scale proportionally with IPTG, with reduced efficiency at high inducer levels.
  • RNA-seq indicated that transcription was not limiting in LB medium; translational/post-translational burdens and stress responses likely limited iT3SS function.

Conclusions:

  • Bacterial growth medium significantly impacts engineered T3SS performance, potentially due to host resource allocation affecting translational efficiency.
  • Impaired translation and assembly of iT3SS components compromise their membrane insertion.
  • A medium-aware framework is essential for optimizing engineered secretion systems in synthetic biology.