Research Overview

Research

Figure 1: A general overview of our research program. The scheme highlights various research areas involving the studies of phase transitions in soft living matter and their functions.

A primary focus of our research program is on understanding the physical principles of (non) folding and phase behavior of Intrinsically Disordered Proteins (IDPs) and their complexes with RNA and DNA that govern their biological functions. Our studies utilize an innovative integration of sensitive, high-resolution fluorescence microscopy, optical tweezer technology, microfluidics, cell biology, polymer theories, and computational methods. IDPs account for a significant portion of the eukaryotic proteome (30 ‒ 40%). Although they challenge the classical protein structure-function paradigm, it is well established now that the disordered proteome performs important organizational, regulatory, and signaling functions in living systems. Furthermore, IDPs are commonly associated with a broad repertoire of human diseases including neurodegeneration and cancer. Three broad research areas in my group are (Fig. 1)

Dissecting Phase Behavior of Protein-RNA Condensates

Stimuli-responsive Multilayered Protein-RNA Condensates

In biological cells, multivalent disordered RNA-binding proteins form dynamic condensates through phase transitions that play essential roles as intracellular storage compartments and signaling hubs. Examples include the nucleolus, stress granules (SG), processing bodies (P-bodies), transcription factories, PML bodies, heterochromatin domains, and para-speckles. In our laboratory, we investigate the most fundamental properties of these bio-condensates utilizing a combination of in vitro reconstitutions and cell culture models (Fig. 1). Specifically, we are studying how protein primary sequence encodes features that drive the formation of liquid-like protein droplets through biomolecular phase transitions, and how RNA binding regulates their phase behavior, compositional specificity, and transport properties. Our studies are expected to shed light on how protein condensate liquid-phase physical properties are encoded by the constituent biopolymers, and how human disease-associated mutations lead to abnormal intracellular condensation of several RNA-binding proteins.

Quantifying Dynamical Properties and Aging Behavior of Biomolecular Condensates

Quantitative understanding of the molecular driving forces underlying the complex material properties of RNA/DNA-binding protein condensates necessitates experimental tools and a conceptual/theoretical framework to systematically probe how their dynamical properties across different lengths- and timescales are encoded within the constituent protein molecules. A major effort in my laboratory is dedicated to developing and employing multi-parametric methodologies featuring correlative single-molecule fluorescence microscopy, optical tweezers, and microfluidics (Fig. 1), a combination that can robustly quantify the structure and dynamical properties of nascent and matured biomolecular condensates on nano-to-microscale. We are employing this toolbox to dissect and target sequence-encoded molecular interactions that govern the network structure, viscoelasticity, and aging of biomolecular condensates.

Single-Molecule Biophysics of Phase Transitions and Genome Packaging

Nuclear transcriptional condensates are formed by transcription factors, coactivators and RNA polymerase II at specific DNA sites to activate or repress genes.

A major long-term research focus of my research group is chromatin organization via biomolecular phase transitions. We seek to understand how living cells utilize the phase separation of multivalent chromatin-associated factors to organize the genome into membrane-less compartments and regulate gene expression (Fig. 1). In this regard, a key question that we seek to address is how transcriptionally active vs. repressive condensates mechanically alter chromatin/DNA and influence local chromatin accessibility to transcriptional machinery. To achieve these goals, my group is pushing the development of novel technologies primarily combining optical tweezers and fluorescence microscopy (Fig. 1).