Defense Date

2021

Document Type

Thesis

Degree Name

Master of Science

Department

Biology

First Advisor

Derek Prosser

Second Advisor

Rima Franklin

Third Advisor

Matthew Hartman

Fourth Advisor

Jason Newton

Abstract

Endocytosis, or internalization of material from the plasma membrane, is an important process for many aspects of eukaryotic cell function. Although clathrin-mediated endocytosis (CME), which requires the coat protein clathrin to facilitate endocytic vesicle formation, is the predominant mechanism for internalization, numerous clathrin-independent (CIE) pathways exist that remain poorly characterized. While the budding yeast Saccharomyces cerevisiae has been a powerful model organism for studying CME, it was long thought to lack any CIE pathways. Recently, a mutant yeast strain defective in CME was used to discover the first known yeast CIE pathway, which requires the small GTPase Rho1. Rho1 regulates numerous aspects of cell function, including organization of the actin cytoskeleton which is critical for endocytosis. The Rho1 GTPase itself is regulated by cycles of activation and inactivation, where guanine nucleotide exchange factors (GEFs) facilitate activation, and GTPase-activating proteins (GAPs) promote inactivation. Evidence suggests that the Rho1 GEFs Rom1 and Rom2 stimulate CIE; however, the role of GAPs in this process have not yet been assessed. Here, I demonstrate that the Rho1 GAP Sac7 acts as a negative regulator of CIE, most likely by spatially restricting Rho1 activity at the plasma membrane. By deleting each of the four known Rho1 GAPs, I show that loss of SAC7, but not any of the other GAPs, restores endocytosis in CME-deficient cells. Moreover, Rho1 becomes more broadly activated at the plasma membrane in sac7∆ cells, suggesting that Sac7 locally regulates Rho1 activation. Thus, Sac7 appears to be a novel regulator of CIE in yeast.

Rights

© The Author

Is Part Of

VCU University Archives

Is Part Of

VCU Theses and Dissertations

Date of Submission

12-11-2021

Available for download on Saturday, October 24, 2026

Included in

Cell Biology Commons

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