The Mechanism of Membrane Fission at the Recycling Endosome of C. Elegans
Abstract
Membrane fission, or the controlled pinching off of vesicles and tubules from intracellular organelles and the plasma membrane, is the first step in vesicle trafficking in eukaryotic cells. The mechanoenzyme dynamin-1 was the first protein identified to catalyze membrane fission and remains one of the best understood fission reactions. However the diversity of membrane fission mechanisms has increased over the past two decades. The focus of this research is identifying the mediator of membrane fission at the basolateral recycling endosome, a pathway well characterized in the model organism C. elegans. Fission at the basolateral recycling endosome returns proteins back to the plasma membrane and requires the activities of at least two interacting proteins, amphiphysin 1 (AMPH-1) and receptor mediated endocytosis-1 (RME1). In order to address the roles of these two proteins, fission reactions were measured by Burst Analysis Spectroscopy (BAS). This single particle fluorescence-based method was developed as a novel approach to quantitatively study membrane fission reactions and validated using the ENTH domain of epsin, a well-characterized membrane fission protein. Using this technique we identified AMPH-1 as the mediator of membrane fission. This fission activity was stimulated in the presence of GTP, an unexpected result because AMPH-1 has no known nucleotide binding domain. Additionally the GTP-stimulated fission activity of AMPH-1 is dependent on the amphipathic helices, and is regulated by RME-1. Therefore we propose that mechanism of membrane fission at the basolateral recycling endosome is through the insertion of the amphipathic helices of AMPH-1.
Citation
Kustigian, Lauren (2019). The Mechanism of Membrane Fission at the Recycling Endosome of C. Elegans. Doctoral dissertation, Texas A&M University. Available electronically from https : / /hdl .handle .net /1969 .1 /187531.