BIOCHEMISTRY, MOLECULAR & CELL BIOLOGY FIELD SEMINAR. Anna Allen – Fromme Lab. Wednesday, October 9^th, 2024. 12:15 PM, Caldwell Hall 100.

Toward a structural and functional understanding of Arf1-GEFs Gea1 and Gea2.

In eukaryotic organisms, the Golgi complex serves as the central hub of the secretory pathway. Traffic within the Golgi is tightly controlled by resident GTPases such as Arf1, which recruit the necessary effector proteins required for cisternal maturation, including cargo adaptors and vesicle coats. As a GTPase, the activity of Arf1 is dependent on whether it is GTP-bound (active) or GDP-bound (inactive). Resident guanine nucleotide exchange factors (GEFs) are distributed throughout the Golgi to catalyze the exchange of GDP for a new molecule of GTP following GTP hydrolysis. In Saccharomyces cerevisiae (budding yeast), there are three Golgi resident Arf1-GEFs: Gea1, Gea2, and Sec7, which distinctly occupy the early (cis-), medial, and late (trans-Golgi network (TGN)) Golgi compartments, respectively. While all three GEFs are similar, particularly in their catalytic domains, Gea1 and Gea2 are genetically paralogous. In my project, I aim to uncover the discrete, localized roles of Gea1 and Gea2 in Arf1 regulation by investigating their mechanisms of localization. To approach this question, I am testing the localization and function of chimeric Gea1/2 constructs that target the N-terminal DCB-HUS domain, which has been shown in previous studies to target each Gea to its unique Golgi compartment. Additionally, with cryoEM, I aim to uncover the full structural picture of the Gea2 activity cycle. My current focus is to characterize the transient Arf1-Gea2-GDP complex by locking it with a GEF-inhibitory drug. Ultimately, defining the functional and structural mechanisms that dictate the behavior and catalytic activity of Gea1 and Gea2 will be crucial to our understanding of precise Arf1 regulation, and therefore global regulation of trafficking within the Golgi complex.

Committee: Chris Fromme, Richa Sardana, Toshi Kawate.