Investigating transcription at the single-copy gene level

 

Signal intensity of a single copy gene transcription site (bright big spot) and cellular mRNAs (small scattered spots), versus a replicated transcription site.

Signal intensity of a single copy gene transcription site (bright big spot) and cellular mRNAs (small scattered spots), versus a replicated transcription site.

Gene expression is a complex and highly regulated process involving numerous steps, and is challenging to explore within living cells. One of the most important steps is called transcription. It results in an mRNA strand that exits the nucleus and then translates into a protein. Nevertheless, the way that transcription functions in time and space are not yet fully understood.

In collaboration with Dr. Yaron Shav-Tal’s lab, we developed a unique system to observe a single copy gene transcription integrated into the DNA of a human cell. With this system we can investigate transcription on a single-copy gene level in living cells. By probing fluorescence fluctuations at transcription sites we extract important parameters on the dynamics controlling transcription.

Using live cell imaging techniques we study the dynamics of transcription: transcription rates and variation in the number of RNA polymerases transcribing on the gene. Studying this process on an almost single molecule level enables us to quantify a single event statistics without the obscurants of an average on multiple genes.

Schematic presentation of transcription demonstrating the PolII  distribution. When the polymerase reaches the MS2 region, the mRNA MS2 stem-loops are coated by the MS2-GFP proteins.

Schematic presentation of transcription demonstrating the PolII distribution. When the polymerase reaches the MS2 region, the mRNA MS2 stem-loops are coated by the MS2-GFP proteins.


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