Intrinsically Disordered Regions as Facilitators of the Transcription Factor Target Search
Transcription factors (TFs) play a critical role in organismal development and function by regulating gene expression. Despite decades of intense research, the factors that determine the specificity and speed at which eukaryotic TFs locate their target binding sites remain a subject of intrigue. Intrinsically disordered regions (IDRs) within TFs have recently emerged as significant players in the TF target search process. However, the inherent challenges in studying IDRs stem from their ability to confer specificity despite low sequence complexity and maintain functional conservation even amidst rapid sequence divergence.
Recent advances in computational and experimental methodologies are beginning to unfold the sequence-function relationship within TF IDRs. These insights are shedding light on the potential mechanisms by which IDRs direct the search for DNA targets, including their role in the formation of biomolecular condensates, TF co-localization, and the hypothesis of IDRs directly interacting with specific genomic regions.
The principles of gene regulation, birthed through studies on the Escherichia coli Lac operon nearly a century ago, laid the foundation for understanding TFs. Since then, research has established that cells across life’s kingdoms employ hundreds to thousands of TFs to bind specific genomic sites and regulate gene expression. Eukaryotic TFs consist of structured regions—often located in the DNA-binding domains (DBDs)—and intrinsically disordered regions (IDRs). While DBDs have been extensively studied for their critical role in binding specificity, the nuanced contribution of IDRs has garnered increasing attention.
IDRs tend to exhibit low sequence complexity and contain fewer hydrophobic residues than folded domains. They lack a stable 3D structure and instead transition through a flexible ensemble of conformations or remain permanently disordered. Despite their lack of stable folding, IDRs can interact with other biomolecules, forming complexes with varying degrees of structural rigidity and conformational freedom. TFs’ IDRs are generally elongated, often spanning hundreds of amino acids, and feature distinct compositions compared to IDRs of other proteins. These regions can serve as linkers between functional domains or perform autonomous functions such as acting as TF effector domains, nuclear localization signals, and stability-regulating degrons. A notable role of IDRs involves their incorporation of TFs into biomolecular condensates, a phenomenon evident in super-enhancers.
IDRs have presented significant challenges for investigation, primarily due to the ineffectiveness of alignment-based sequence analysis, a traditional method used to study protein function. Comparative analysis typically suggests that functional protein regions exhibit a degree of sequence conservation—useful for folded domains that require accurate residue ordering to function. However, functionally conserved IDRs display minimal sequence conservation using these methods, pointing to unique sequence-function relationships independent of linear residue ordering. This prompted the development of novel computational and experimental methods designed to analyze IDRs, which accommodate their characteristics by detecting alignment-free features and considering the length and robustness of their functional regions.
This article summarizes evidence supporting the involvement of IDRs in guiding the TF target search across eukaryotic genomes. Studies mapping in vivo TF binding locations and tracking TF dynamics through single-molecule live microscopy lend critical insights into this process. We will delve into the emerging computational and experimental approaches for analyzing TF IDR sequences, highlighting the challenges and insights associated with the IDR sequence determinants that promote TF targeting. These studies frequently draw from substantial preliminary understanding derived from Msn2, a well-characterized IDR-directed TF, inspiring discussions about a potential new sequence ‘grammar’ that could guide IDR-chromatin interactions.
Though we do not cover in vitro IDR analysis methods here, given their prior comprehensive reviews, their potential and significance in understanding IDR-based TF specificity cannot be overstated.
