Septin Organization in Multinucleated Cells

Dartmouth College, Hanover NH

Investigators

Abstract

Proteins at the cell cortex link the cell interior and exterior by transmitting and responding to a variety of signals. Thus, the cell cortex is an information processing center where cells can receive and react to diverse stimuli in their environment or within the cell. A conserved family of proteins called septins organizes and regulates structures and processes at the cortex of eukaryotic cells. In addition to performing essential functions specific to certain cell types, septins are also important for normal cell division; they can act as "molecular fences" in membranes to keep proteins in specific locations and they are thought to act as protein "scaffolds" to assemble specific groups of proteins at the cell cortex. In most cells, septin proteins self-assemble into filaments that further assemble into complex higher-order structures such as rings and fibers. Despite the widespread distribution and varied functions of septins, little is known about the molecular mechanisms that direct septin assembly into filaments and the higher order organization of these filaments. A major goal of this project is to determine how septins assemble into an array of morphologically different forms within a single cell. The PI and her colleagues will investigate how septin structures are assembled, reorganized and regulated in the multinucleated, filamentous fungus, Ashbya gossypii. In this filamentous fungus, septins assemble into many morphologically distinct and dynamic structures, making it an excellent model for understanding the molecular controls of complex septin organization. The simple lifestyle and small genome of the fungus facilitates generation and analysis of mutants in regulatory proteins. Additionally, septin proteins can be visualized in living cells by linking the septin proteins to fluorescent proteins that are visible under the microscope. The specific goals of this project are to: 1. Identify the composition and dynamics of different septin complexes that coexist in one A. gossypii cell. 2. Establish the regulatory pathways that control the assembly of different septin rings. 3. Determine if the nuclear division cycle directs changes in the septin cortex in a multinucleated cell. These goals will be achieved using a combination of time-lapse fluorescence microscopy, electron microscopy and biochemistry for mutant analysis. This work is significant because the septins are a nearly ubiquitous, highly conserved protein family, yet major gaps exist in our knowledge of how they assemble and function in complexes in living cells. Furthermore, little is known about how internal or external signals lead to the assembly of elaborate types of septin structures. Knowledge gained from this work will provide a mechanistic foundation for understanding septin organization in other eukaryotic cells. The project will have substantial broader impact on education and training. The PI will mentor graduate and undergraduate students from multiple programs in research projects; undergraduate students will perform experiments, attend weekly lab meetings and present their data at Dartmouth's undergraduate research symposia.