Document Type

Honors Thesis

Major

Biochemistry

Advisor(s)

Joshua Kelley

Committee Members

Julie Gosse, Samantha Jones, Jennifer Newell-Caito, Robert Wheeler

Graduation Year

May 2021

Publication Date

2025

Abstract

Saccharomyces cerevisiae utilize a G-protein coupled receptor (GPCR) signaling pathway for mating, that recognizes mating factor and leads to cell polarization and elongation. For directed growth to occur, endocytosis is required to internalize regions of the membrane and septins must form structures along this area for polarity to form and for the cytoskeleton to rearrange. Epsins are proteins that support membrane curvature and assist in selective endocytosis. They also contain a domain that is known to interact with septin regulators. Therefore, we suspect that epsins are involved in coupling endocytosis to septin deposition. To test the role of epsins in septin distribution, we engineered yeast deletion strains that lacked epsins (ENT1 and ENT2), exposed them to pheromone using microfluidics devices and monitored them over 12 hours to track polarization and elongation. Additionally, we deleted epsins in a hyperactive strain (Gpa1G302S) that manifests improper septin deposition and an inability to track a gradient, to further investigate our hypothesis. We also created hyperactive deletion strains lacking gics (proteins involved in recruiting septins to the membrane). By deleting gics we were able to slow the rate of septin deposition and rescue the hyperactive morphology, suggesting that proper septin localization is dependent on the rate of septin deposition. This was further supported by the observation that exposing these cells to pheromone at sub-optimal temperatures also partially rescued the distribution of septins. Our data suggests that septin localization is rate dependent and that epsins are involved in temporally regulating septin deposition through their interacts with other downstream proteins and receptor endocytosis.

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