Publications
RNA Polymerase II cluster dynamics predict mRNA output in living cells
WK Cho, N Jayanth, BP English, T Inoue, JO Andrews, W Conway, JB Grimm, JH Spille, LD Lavis, T Lionnet, II Cisse. Elife 5, e13617 (2016).
Abstract:
Protein clustering is a hallmark of genome regulation in mammalian cells. However, thedynamic molecular processes involved make it difficult to correlate clustering with functionalconsequences in vivo. We developed a live-cell super-resolution approach to uncover thecorrelation between mRNA synthesis and the dynamics of RNA Polymerase II (Pol II) clusters at agene locus. For endogenousb-actin genes in mouse embryonic fibroblasts, we observe that short-lived (~8 s) Pol II clusters correlate with basal mRNA output. During serum stimulation, astereotyped increase in Pol II cluster lifetime correlates with a proportionate increase in thenumber of mRNAs synthesized. Our findings suggest that transient clustering of Pol II mayconstitute a pre-transcriptional regulatory event that predictably modulates nascent mRNA output.
We show that transient RNA Pol II clustering with a lifetime of a few seconds is a mechanism for controlling gene expression. Pol II was tagged with the photo-convertable fluorescent protein Dendra2 and cluster properties evaluated by time-correlated analysis of PALM data (tcPALM). The lifetime of Pol II clusters (but not their size, strength, or frequency) at the beta actin gene locus is increased 12.5 minutes after serum stimulation. The mRNA output as quantified by MS2 fluorescence is proportional to the cluster lifetime. We used simulations of Dendra2 photophysics to estimate the number of Pol II molecules in a cluster. It's up to a hundred, but only 4-20 get loaded onto the gene!
