Dynamic switching of transcriptional regulators between two distinct low-mobility chromatin states

KAUSTUBH WAGH; STAVREVA, DIANA A.; JENSEN, RIKKE A. M.; VILLE PAAKINAHO; GREGORY FETTWEIS; R. LOUIS SCHILTZ; SUSANNE MANDRUP; DIEGO M PRESMAN; UPADHYAYA, ARPITA; GORDON L. HAGER

Congreso; CHSL meeting Mechanisms of Eucaryotic Transcription 2023; 2023

CSHL

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. Technological advances over the past 20 years have enabled us to follow single TF molecules within live cells as they interact with chromatin. Using single-molecule tracking, coupled with machine learning, we developed a framework to study the mobility of chromatin and transcriptional regulators within the nucleus. We show that histone H2B and multiple chromatin-bound transcriptional regulators display two distinct low-mobility states. Ligand activation results in a dramatic increase in the propensity of steroid receptors to bind in the lowest mobility state. Mutational analysis revealed that interactions with chromatin in the lowest mobility state require an intact DNA-binding domain as well as oligomerization domains. Importantly, these states are not spatially separated as previously believed but in fact, individual chromatin-bound TF molecules can switch between two dynamic chromatin polymer states (marked by H2B) on timescales of seconds. Single bound-TF molecules with different mobilities exhibit different dwell time distributions, suggesting that the mobility of TFs is intimately coupled with their binding dynamics. Together, our results identify two unique and distinct low-mobility states that appear to represent common pathways for transcription activation in mammalian cells.