Understanding how plants or animals function requires the ability to study molecular processes within specific cell types or even in single cells of the body. It is well known that cells within organisms respond to stimuli, such as light, gravity, hormones or any number of environmental triggers; however, when and how cells respond remains a mystery in part because it is technologically challenging to identify which cells are responding. Often responsive cells are not easily distinguished from non-responsive cells. In this high-risk, high reward project, the research team solves this problem by developing a molecular tool to identify and capture when and where cells respond to a stimulus, such as a hormone signal, in plant roots. The method will be broadly useful to detect other responses, and in many different plants and plant organs, due to its universal and modular design. The new technology will allow scientists to test new environments or treatments and identify how plant cells respond rapidly and precisely. The outcomes will impact researchers' ability to understand growth responses of diverse plants ranging from crops to model systems. Undergraduate students will have in-depth research training by contributing to the project during a summer research intensive.
Recent advances in molecular profiling of specific plant cell types have generated important breakthroughs in understanding cell differentiation and cell type-specific stress responses. The research team has played a key role in this through the development and application of our INTACT (Isolation of Nuclei TAgged specific Cell Types) and TRAP (Translating Ribosome Affinity Purification) and TRAP (Translating Ribosome Affinity Purification) methods in crop plants. Despite the utility of INTACT and TRAP, there remains a pressing need to be able to capture the subsets of cells of a given cell type as they respond to a stimulus. Cells undergoing a response to such a stimulus are a subset of a population and cannot be easily captured by available methods. This limited resolution hinders the ability to address the molecular details of plant-microbe interactions, responses to hormone signaling, and spatially non-uniform abiotic stresses. The research team will address this problem by developing a system for selective purification of nuclei and translating ribosomes from cell subpopulations that are actively undergoing a specific response, such as drought sensing or interaction with a microbe. This will be achieved by producing two-component INTACT and TRAP systems in which one component (the biotinylatable nuclear envelope- or ribosomal-tagging protein) is expressed from a cell type-specific promoter, while Biotin ligase is driven from a stimulus-inducible promoter. In this way, the entire population of a cell type can be analyzed by purification of organelles using anti-GFP beads, while the subset of responsive cells can be analyzed by Streptavidin bead purification.
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.
