BRC-BIO: investigating the molecular mechanisms of fungal cell fusion

This National Science Foundation (NSF) Project Grant, under the Biological Sciences program (CFDA 47.074), was awarded to Stetson University, Inc. in the amount of $502,998 with a period of performance from August 1, 2023 to July 31, 2026. The project aims to investigate the molecular mechanisms of fungal cell fusion, using the yeast Saccharomyces cerevisiae as a model system. Specifically, the research will focus on identifying novel regulators of cell wall degradation and plasma membrane fusion during the cell fusion process. This work will be performed by undergraduate students through course-based research experiences and independent research opportunities, providing them with hands-on training in genetic manipulation techniques and advanced microscopy. The findings from this project have the potential to provide insights into the fundamental process of cell fusion in higher eukaryotes, including humans.

Abstract

The process of two cells combining into one, known as cell fusion, happens frequently throughout the development of mammals, including humans. During reproduction, sperm and EGG cells must fuse for fertilization to occur, and later in fetal development muscle cells fuse into long skeletal muscle fibers that allow for movement. These are just two examples of the many fusion events that occur and highlight the importance of this general process. Cell fusion does not happen spontaneously and is highly regulated by proteins that ensure the process occurs at the correct time and place. Identifying these regulators and determining how they function is crucial for understanding cell fusion but has proven difficult to study in mammals. The yeast, saccharomyces cerevisiae, undergoes a fusion event similar to fertilization during their life cycle making them an excellent model system to study fusion. Yeast are easy to grow and manipulate and contain many of the same genes and proteins as human cells. This project aims to study the mechanisms of cell fusion in s. Cerevisiae by identifying novel regulators and analyzing known proteins required for the process. This work will be performed by undergraduates through course-based research experiences and year-long independent research opportunities. Students will gain experience genetically manipulating yeast and performing cutting edge microscopy techniques. This project will encourage the next generation of scientists by allowing them access to research experiences where they will identify mechanisms of yeast fusion that can aid in the understanding of cell fusion in mammals. Yeast cells have a cell wall surrounding their plasma membrane, meaning that once two cells come into contact, they must first degrade their cell walls specifically where they will fuse, similar to the degradation of extracellular matrix required in mammalian systems. Once this occurs, proteins that promote plasma membrane fusion can act, leading to cytoplasmic mixing. While some regulators and morphological events of yeast fusion have been identified, there are still unanswered questions about the process. Overall, this proposal aims to uncover how cell wall degradation and plasma membrane fusion are regulated through mutational analysis of an understudied, key regulator of cell wall degradation, visualization of enzyme secretion during fusion, and identification of novel plasma membrane fusion regulators. Fluorogen-activating tags will be used to develop a microscopic assay for fusion-specific secretion, addressing a long-standing hypothesis in the field. Additionally, suppressor analysis of a membrane fusion mutation recently identified by a student researcher will allow for identification of novel plasma membrane fusion regulators, which have long-eluded scientists. Along with significant advancements to the understanding of yeast fusion, this work involves proteins that are conserved from yeast to humans allowing the potential to provide insight into this fundamental process in higher eukaryotes. 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.