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Updated: May 28, 2026
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Single-molecule Imaging of Gene Regulation In vivo Using Cotranslational Activation by Cleavage (CoTrAC)
Published on: March 15, 2013
Cell cycle dependent TN-C promoter activity determined by live cell imaging.
Michael Halter1, Daniel R Sisan, Joe Chalfoun
1Cell Systems Science Group/Biochemical Science Division, Chemical Science and Technology Laboratory, National Institute of Standards and Technology, Gaithersburg, MD 20899, USA. michael.halter@nist.gov
Summary
Tenascin-C (TN-C) promoter activity rises in the latter 40% of the cell cycle, varying between individual cells. This study uses live cell microscopy and automated analysis to reveal subtle gene regulation mechanisms.
Area of Science:
- Cell biology
- Molecular biology
- Biochemistry
Background:
- Tenascin-C (TN-C) is an extracellular matrix protein vital in development, wound healing, and cancer.
- The precise control mechanisms and physiological functions of TN-C are not fully understood.
Purpose of the Study:
- To investigate the dynamic regulation of the TN-C promoter activity.
- To understand how TN-C gene expression is controlled throughout the cell cycle.
Main Methods:
- Utilized live cell microscopy with phase contrast and fluorescence imaging.
- Employed an NIH 3T3 cell line with a TN-C promoter driving destabilized green fluorescent protein (GFP).
- Applied automated image analysis for quantifying GFP in single cells over 62-hour live imaging experiments.
Main Results:
- Observed significant cell-to-cell variation in TN-C promoter activity patterns across the cell cycle.
- Demonstrated an average increase in TN-C promoter activity during the final 40% of the cell cycle.
- Found that the magnitude of promoter activity increase correlated with earlier activity levels.
Conclusions:
- Live cell microscopy combined with automated analysis provides a powerful tool for dissecting gene regulatory mechanisms.
- TN-C promoter activity exhibits dynamic regulation tied to the cell cycle.
- This approach overcomes limitations of traditional population-averaged measurements for revealing subtle gene expression dynamics.
