Gene-expression events are intricately linked — and there is now a way to view the complete set of events together in a single living cell. Reporting in Cell, David Spector and colleagues have developed a system to simultaneously visualize gene expression at the DNA, RNA and protein levels.

Spector and co-workers constructed an inducible plasmid with a transcriptional unit that encodes cyan fluorescent protein (CFP) — allowing visualization of the protein — and so-called translational operator RNA (a 19-nt RNA stem loop). Upstream of the transcriptional unit are repeats of the bacterial lac operator sequence, so that, in the presence of a lac-repressor-binding protein that is expressed as a fluorescent fusion protein, the transgene locus can be visualized. And, transcribed RNA can be visualized by expression of yellow fluorescent protein (YFP) fused to MS2, a protein that binds the translational operator RNA. The authors selected a stable transfectant that expressed ∼200 copies of the transgene that was integrated in a euchromatic region in human chromosome 1.

The authors simultaneously analysed the chromatin and RNA dynamics at the transcription site, and showed that, in the absence of transcriptional activation, the CFP–lac repressor marks the chromatin of the transgene locus as a highly condensed dot and MS2–YFP is diffusely distributed throughout the nucleus. On transcriptional induction, the locus decondenses, proteins of the gene-expression machinery colocalize with the decondensed locus, and MS2–YFP associates with the nascent transcripts at the locus and with messenger ribonucleoprotein (mRNP) particles in the nucleoplasm.

Next, Spector and colleagues measured the kinetics of RNA synthesis at the site of transcription. They detected an early increase in RNA levels (5 min post-induction), as shown by a small spot of MS2–YFP at the locus, while the chromatin was still highly condensed. The first chromatin changes were detected at ∼17.5 min, by which time transcripts were seen across the entire locus. RNA levels peaked by ∼130 min and subsequently decreased: the induction of transcription seems to slow over time.

Heterochromatin protein 1 (HP1), as well as histone H3 that is tri-methylated at lysine 9 (this modification serves as the HP1-binding site), colocalize with the condensed locus, which indicates that it is heterochromatic in nature. After transcriptional activation, HP1 and tri-methylated histone H3 are depleted while the locus becomes decondensed, and the histone variant H3.3 becomes deposited. So, histone exchange might be a possible mechanism for removing methylated histones from chromatin during transcriptional activation.

Spector and colleagues have developed a powerful tool for studying gene expression and chromatin, in time and space, in living cells. They suggest that this system of transgene arrays might also prove useful for studying the mechanisms of RNA-interference-mediated gene silencing in vivo.