The lac repressor displays facilitated diffusion in living cells

Area of Science:

• Molecular biology
• Genetics
• Biophysics

Background:

• Transcription factors (TFs) regulate gene expression by binding specific DNA sites.
• Facilitated diffusion (1D DNA sliding + 3D cytoplasmic diffusion) is a proposed mechanism for rapid TF target location.
• In vitro evidence exists, but in vivo experimental proof in living cells is lacking.

Purpose of the Study:

• To experimentally investigate the DNA sliding mechanism of transcription factors in living bacterial cells.
• To quantify the sliding distance of the lac repressor on chromosomal DNA.
• To determine if DNA-bound proteins impede TF sliding.

Main Methods:

• Development of a novel single-molecule assay for observing TF behavior in living bacteria.
• In vivo observation of the lac repressor's interaction with chromosomal DNA.
• Analysis of sliding distances and frequency of operator interaction.

Main Results:

• The lac repressor slides an average of 45 ± 10 base pairs on chromosomal DNA.
• Sliding can be hindered by other proteins bound to nearby DNA sequences.
• The lac repressor repeatedly slides over its target operator (>90% of cases) before binding.

Conclusions:

• The study provides the first experimental evidence of DNA sliding by a transcription factor in living cells.
• DNA sliding is a crucial component of the facilitated diffusion mechanism for gene regulation in vivo.
• The findings suggest a balance between efficient searching of DNA and rapid binding to specific operator sites.