Biology subjects

Stavreva, D. A.

Publications and source records attributed to Stavreva, D. A..

4 recordsLinked to original sources

bioRxiv2026

QuantiTrack: A unified software to study protein dynamics in living cells

Linking the spatiotemporal dynamics of proteins in live cells to biological function is a fundamental challenge in biology. Single molecule tracking (SMT) has emerged as a powerful technique to investigate protein dynamics at the single molecule level. However, SMT analysis often requires expertise in biophysical modeling and programming, and integrating results from different analyses can be challenging. To address these barriers, we developed QuantiTrack: a MATLAB-based SMT analysis software with a simple graphical user interface. This provides a much-needed end-to-end solution where a user can load a movie, detect and track single molecules, and perform complementary downstream analyses within a standardized workflow. QuantiTrack includes quantitative metrics for selecting detection and tracking parameters and troubleshooting experimental design, and includes a detailed step-by-step User Guide. We used simulations to demonstrate how signal intensity, labeling density, and motion blur affect detection and tracking fidelity. Using multi-state simulations, we further benchmarked complementary methods to identify distinct mobility states from heterogeneous trajectory populations. Finally, we applied QuantiTrack to real experimental data where we address how the glucocorticoid receptor (GR), a hormone-regulated transcription factor, responds to treatment and washout of its cognate hormone. Hormone washout results in rapid (in minutes) downregulation of GR target genes to basal levels. By integrating complementary analyses within QuantiTrack, we showed that hormone washout substantially reduced the bound fraction of GR, its occupancy in the mobility state associated with GR activation, and dwell times. Together, these analyses showcase QuantiTrack as an integrated platform for extracting biologically meaningful measurements from single molecule trajectories.

biophysics↗

bioRxiv2025

Bile acids target an exposed cavity in the glucocorticoid receptor modulating receptor self-assembly, chromatin binding and transcriptional activity

The glucocorticoid receptor (GR) is an essential transcription factor that controls metabolism and homeostasis. Glucocorticoids (GCs) activate the GR upon occupying the internal ligand-binding pocket (LBP) of its ligand-binding domain (GR-LBD), which has been the focus of most previous structure-function studies. Synthetic GCs such as dexamethasone are widely used to treat inflammatory diseases, but their chronic use results in major side effects, whose molecular underpinnings remain unresolved. Here we present a thorough analysis of the topography of GR-LBD and its ability to bind small-molecule compounds, especially cholesterol derivatives. We show that one important class of steroids, bile acids, bind to previously unidentified and highly conserved, surface-exposed cavities on GR-LBD. We show that bile acids affect GR turnover and self-assembly in living cells, modulating receptor transcriptional activity. These findings reveal a previously unrecognized mechanism of GR regulation, with implications for the design of GCs with novel mechanisms of action. TeaserBile acids modulate the activity of the glucocorticoid receptor upon binding to an exposed allosteric pocket thereby influencing transcriptional regulation and receptor self-assembly in living cells.

molecular biology↗

bioRxiv2024

Transcription factors form a ternary complex with NIPBL/MAU2 to localize cohesin at enhancers

While the cohesin complex is a key player in genome architecture, how it localizes to specific chromatin sites is not understood. Recently, we and others have proposed that direct interactions with transcription factors lead to the localization of the cohesin-loader complex (NIPBL/MAU2) within enhancers. Here, we identify two clusters of LxxLL motifs within the NIPBL sequence that regulate NIPBL dynamics, interactome, and NIPBL-dependent transcriptional programs. One of these clusters interacts with MAU2 and is necessary for the maintenance of the NIPBL-MAU2 heterodimer. The second cluster binds specifically to the ligand-binding domains of steroid receptors. For the glucocorticoid receptor (GR), we examine in detail its interaction surfaces with NIPBL and MAU2. Using AlphaFold2 and molecular docking algorithms, we uncover a GR-NIPBL-MAU2 ternary complex and describe its importance in GR-dependent gene regulation. Finally, we show that multiple transcription factors interact with NIPBL-MAU2, likely using interfaces other than those characterized for GR.

molecular biology↗

bioRxiv2022

Single-molecule tracking reveals two low-mobility states for chromatin and transcriptional regulators within the nucleus

How transcription factors (TFs) navigate the complex nuclear environment to assemble the transcriptional machinery at specific genomic loci remains elusive. Using single-molecule tracking, coupled with machine learning, we examined the mobility of multiple transcriptional regulators. We show that H2B and ten different transcriptional regulators display two distinct low-mobility states. Our results indicate that both states represent dynamic interactions with chromatin. Ligand activation results in a dramatic increase in the proportion of steroid receptors in the lowest mobility state. Mutational analysis revealed that only chromatin interactions 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 H2B and TF molecules can dynamically switch between them. Together, our results identify two unique and distinct low-mobility states of transcriptional regulators that appear to represent common pathways for transcription activation in mammalian cells.

biophysics↗