Insights on transcription factors and chromatin dynamics through single-molecule tracking in live cells

DIEGO M PRESMAN

Punta del Este

Simposio; The 11th International Weber Symposium on Innovative Fluorescence Methodologies in Biochemistry and Medicine; 2023

Laboratory of Fluorescence Dynamics

Transcription factors (TFs) scan the nucleus in search of their consensus binding motifs located within enhancers or promoter-proximal regions. The mechanism by which TFs navigate the complex nuclear environment to assemble the transcriptional machinery at specific genomic loci remains elusive. Using single-molecule tracking (SMT), coupled with machine learning, we examined the mobility of multiple transcription factors and coregulators both in space and time. From a temporal perspective, our results indicate that steroid hormone receptors, as well as other transcriptional coregulators, architectural proteins, and remodelers, follow a power-law distribution of dwell-times, blurring the temporal line between non-specific and specific binding, suggesting that productive binding may involve longer binding events than previously believed. From a spatial perspective, we find that all studied proteins display two distinct low-mobility states. Ligand activation results in a dramatic increase in the proportion of TFs in both low-mobility states. Strikingly, histone H2B also exhibits the same two low-mobility states and examination of several different cell lines demonstrates the universality of these states for both TFs and H2B. Taken together, our data suggest that both low-mobility states are intimately coupled with mobile chromatin. Importantly, these states are not spatially separated as individual H2B and TF molecules can dynamically switch between the two low-mobility states. Overall, we propose a continuum of affinities model to explain TF dynamics and found two unique and distinct low-mobility states of transcriptional regulators that appear to represent common pathways for transcription activation in eucaryotic cells.