Dailey, G. | Search bioRxiv

bioRxiv2017

Dynamic and Selective Low-Complexity Domain Interactions Revealed by Live-Cell Single-Molecule Imaging

Many eukaryotic transcription factors (TFs) contain intrinsically disordered low-complexity domains (LCDs) but how they perform transactivation functions remains unclear. Recent studies report that TF-LCDs can undergo hydrogel formation or liquid-liquid phase separation in vitro. Here, live-cell single-molecule imaging reveals that TF-LCDs form local high concentration interaction hubs at synthetic and endogenous genomic loci. TF-LCD hubs stabilize DNA binding, recruit RNA polymerase II (Pol II) and activate transcription. LCD-LCD interactions within hubs are highly dynamic, display selectivity with binding partners, and are differentially sensitive to disruption by hexanediols. These findings suggest that under physiological conditions, rapid reversible and multivalent LCD-LCD interactions occur between TFs and the Pol II machinery, which underpins a central mechanism for transactivation and plays a key role in gene expression and disease.

biophysics↗

bioRxiv2017

Identifying a TFIID interactome via integrated biochemical and high-throughput proteomic studies

The core promoter recognition TFIID complex acts as a central regulator for eukaryotic gene expression. To direct transcription initiation, TFIID binds the core promoter DNA and aids recruitment of the transcription machinery (e.g., RNA polymerase II) to the transcription start site. Many transcription factors target TFIID to control vital cellular processes. Current studies on finding TFIID interactors have predominantly focused on transcription factors. Yet, a comprehensive interactome of mammalian TFIID has not been established. Therefore, this study sought to reveal potential TFIID-nucleated networks by identifying likely co-regulatory factors that bind TFIID. By using intact native human TFIID complexes, we have exploited three independent approaches including a high-throughput Next Generation DNA sequencing coupled with proteomic analysis. Among these methods, we found some overlapping and new candidates in which we further assessed three putative interactors (i.e., Sox2, H2A and EMSY) by co-immunoprecipitation assays. Notably, in addition to known TFIID interactors, we identified a number of novel factors that participate either in co-regulatory pathways or non-transcription related functions of TFIID. Overal, these results indicate that, in addition to transcription initiation, mammalian TFIID may be involved in broader regulatory pathways than previous studies suggested.

biochemistry↗