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An Amphiphysin-Like Domain in Fus2p Is Required for Rvs161p Interaction and Cortical Localization

Author(s): Stein, Richard A; Smith, Jean A; Rose, Mark D

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Abstract: Cell–cell fusion fulfils essential roles in fertilization, development and tissue repair. In the budding yeast, Saccharomyces cerevisiae, fusion between two haploid cells of opposite mating type generates the diploid zygote. Fus2p is a pheromone-induced protein that regulates cell wall removal during mating. Fus2p shuttles from the nucleus to localize at the shmoo tip, bound to Rvs161p, an amphiphysin. However, Rvs161p independently binds a second amphiphysin, Rvs167p, playing an essential role in endocytosis. To understand the basis of the Fus2p–Rvs161p interaction, we analyzed Fus2p structural domains. A previously described N-terminal domain (NTD) is necessary and sufficient to regulate nuclear/cytoplasmic trafficking of Fus2p. The Dbl homology domain (DBH) binds GTP-bound Cdc42p; binding is required for cell fusion, but not localization. We identified an approximately 200 amino acid region of Fus2p that is both necessary and sufficient for Rvs161p binding. The Rvs161p binding domain (RBD) contains three predicted alpha-helices; structural modeling suggests that the RBD adopts an amphiphysin-like structure. The RBD contains a 13-amino-acid region, conserved with Rvs161p and other amphiphysins, which is essential for binding. Mutations in the RBD, predicted to affect membrane binding, abolish cell fusion without affecting Rvs161p binding. We propose that Fus2p/Rvs161p form a novel heterodimeric amphiphysin required for cell fusion. Rvs161p binding is required but not sufficient for Fus2p localization. Mutations in the C-terminal domain (CTD) of Fus2p block localization, but not Rvs161p binding, causing a significant defect in cell fusion. We conclude that the Fus2p CTD mediates an additional, Rvs161p-independent interaction at the shmoo tip.
Publication Date: Feb-2016
Electronic Publication Date: 17-Dec-2015
Citation: Stein, Richard A, Smith, Jean A, Rose, Mark D. (2016). An Amphiphysin-Like Domain in Fus2p Is Required for Rvs161p Interaction and Cortical Localization. G3: Genes|Genomes|Genetics, 6 (2), 337 - 349. doi:10.1534/g3.115.023960
DOI: doi:10.1534/g3.115.023960
EISSN: 2160-1836
Pages: 337 - 349
Type of Material: Journal Article
Journal/Proceeding Title: G3; Genes|Genomes|Genetics
Version: Final published version. This is an open access article.

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