Transcription factors (TFs) are essential regulators of eukaryotic gene expression and orchestrate some of the most complex phenomena in cell biology, from cell division to death. Transcriptional activation is reliant on unstructured domains, which are hypothesized to facilitate the assembly of complex transcriptional machinery by forming highly concentrated and dynamic interaction hubs. However, current protein labeling methods have been shown to disrupt TF activity and traditional microscopy approaches are not sufficiently descriptive to understand the nuances associated with the activation process. My work focuses on developing chemical tools to uncover the molecular basis of TF activity, specifically addressing the major challenges associated with labeling TFs for functional investigation. My primarily focus is to develop a novel labeling strategy, based on a self-excising ligase, to afford perturbation free labeling through a fusion protein approach (Figure 1). Moreover, I am interested in using solvatochromic dyes to capture the molecular properties associated with the high concentrations of TFs required for transcriptional activity through using existing fusion proteins or novel strategies (Figure 2). Together, these tools will serve as useful complements to existing technologies used in current imaging and tracking experiments and provide new avenues for investigating the functional basis of TFs which eludes current methods.
Figure 1: Schematic representing the components and output of a fluorescence labeling approach based on a self-excising ligase.
Figure 2: Illustration of how the combination of solvatochromic dyes with covalent labeling strategies may allow for better resolution and description of TF hubs formation.