EAGER: Tools4Cells: Detecting biomolecular condensation via genetically encoded biosensors

Ohio State University, The, Columbus OH

Investigators

Abstract

Biomolecular condensates, formed through liquid-liquid phase separation (LLPS) and related phase transitions, represent a novel mechanism to organize diverse cellular processes in biological systems. These dynamic structures play pivotal roles in fundamental processes such as growth, development, and stress responses. Currently, the standard approach for investigating biomolecular condensation involves microscopy-based techniques extensively applied in individual cells and in vitro settings. However, tools are lacking to explore condensates within the complex environments of tissues, organs, and whole organisms. The goal of this EAGER project is to develop innovative biosensors to probe condensate dynamics with expanded spatiotemporal resolution in multicellular plants at the organismal level. The success of this innovative tool will greatly extend the timescale of detecting condensates from minutes to several days, providing a more comprehensive understanding of condensate dynamics over longer periods. The broader impacts of this interdisciplinary research extend to education, diversity and science communication. The project will also contribute to the training of scientists at the undergraduate, doctoral, and postdoctoral levels. Cellular compartmentalization is a fundamental process in all living systems to control biochemical reactions in space and time. In addition to the canonical membrane-enclosed organelles, biomolecular condensates assembled through phase transitions create another layer of cellular compartments without biological membranes. To date, cell-based assays have been extensively employed to understand the function of condensates. Significant knowledge gaps exist in delineating the contribution of condensates to physiologically relevant processes in vivo, particularly in multicellular organisms such as plants and mammals. This gap is in part due to a dearth of tools to detect biomolecular condensation at the organismal level. Microscopy-based methods are powerful to monitor condensates’ behavior in individual cells but do not support the throughput and sensitivity in organs, tissues, and organisms. This work is guided by a recent discovery by the PI of a new LLPS-mediated host defense mechanism in Arabidopsis thaliana, wherein plants assemble biomolecular condensates mobilized by a group of immune GTPases, Guanylate-Binding Protein-Like (GBPLs) proteins, to combat infection. This project aims to develop novel genetically encoded biosensors as a proxy for GBPL condensate formation at the organismal level over a long period of time. The successful development of the tool will benefit both plant and animal research communities and holds significant promise in uncovering more fascinating insights into how these membraneless compartments contribute to cellular dynamics. This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.