In Vitro Homotypic ER Membrane Fusion Assay Using Isolated Yeast Microsomes

Eunhong Jang; Youngsoo Jun

doi:10.1007/978-1-0716-4314-3_11

In Vitro Homotypic ER Membrane Fusion Assay Using Isolated Yeast Microsomes

Methods Mol Biol. 2025:2887:167-173. doi: 10.1007/978-1-0716-4314-3_11.

Authors

Eunhong Jang¹, Youngsoo Jun²

Affiliations

¹ School of Life Sciences, Gwangju Institute of Science and Technology, Gwangju, South Korea.
² School of Life Sciences, Gwangju Institute of Science and Technology, Gwangju, South Korea. [email protected].

PMID: 39806153
DOI: 10.1007/978-1-0716-4314-3_11

Abstract

Cell-free in vitro assays offer several advantages for elucidating molecular mechanisms underlying various biological processes. Here, we describe a simple and quantitative in vitro assay using isolated yeast microsomes to measure homotypic ER membrane fusion. In this assay, membrane fusion between ER microsomes is monitored by reconstitution of luciferase activity from split luciferase fragments. Our findings reveal that homotypic ER membrane fusion requires not only Sey1p, the yeast atlstin, but also ER-resident SNAREs, such as Sec22p and Sec20p, in Saccharomyces cerevisiae.

Keywords: Atlastin; Endoplasmic reticulum; Membrane fusion; Microsome; SNARE.

MeSH terms

Biological Assay / methods
Endoplasmic Reticulum* / metabolism
Membrane Fusion*
Microsomes* / metabolism
SNARE Proteins / metabolism
Saccharomyces cerevisiae Proteins / metabolism
Saccharomyces cerevisiae* / metabolism

Substances

Saccharomyces cerevisiae Proteins
SNARE Proteins