A proximity labeling strategy enables proteomic analysis of inter-organelle membrane contacts

Maoge Zhou; Bingjie Kong; Xiang Zhang; Ke Xiao; Jing Lu; Weixing Li; Min Li; Zonghong Li; Wei Ji; Junjie Hou; Tao Xu

doi:10.1016/j.isci.2023.107159

A proximity labeling strategy enables proteomic analysis of inter-organelle membrane contacts

iScience. 2023 Jun 17;26(7):107159. doi: 10.1016/j.isci.2023.107159. eCollection 2023 Jul 21.

Authors

Maoge Zhou^{1

2}, Bingjie Kong², Xiang Zhang², Ke Xiao², Jing Lu², Weixing Li², Min Li¹, Zonghong Li¹, Wei Ji^{2

3}, Junjie Hou², Tao Xu^{1

2

3

4}

Affiliations

¹ Guangzhou Laboratory, Guangzhou, Guangdong 510005, China.
² Institute of Biophysics, Chinese Academy of Sciences, Beijing 100101, China.
³ College of Life Science, University of Chinese Academy of Sciences, Beijing 100049, China.
⁴ School of Biomedical Engineering, Guangzhou Medical University, Guangzhou, Guangdong 511436, China.

Abstract

Inter-organelle membrane contacts are highly dynamic and act as central hubs for many biological processes, but the protein compositions remain largely unknown due to the lack of efficient tools. Here, we developed BiFCPL to analyze the contact proteome in living cells by a bimolecular fluorescence complementation (BiFC)-based proximity labeling (PL) strategy. BiFCPL was applied to study mitochondria-endoplasmic reticulum contacts (MERCs) and mitochondria-lipid droplet (LD) contacts. We identified 403 highly confident MERC proteins, including many transiently resident proteins and potential tethers. Moreover, we demonstrated that mitochondria-LD contacts are sensitive to nutrient status. A comparative proteomic analysis revealed that 60 proteins are up- or downregulated at contact sites under metabolic challenge. We verified that SQLE, an enzyme for cholesterol synthesis, accumulates at mitochondria-LD contact sites probably to utilize local ATP for cholesterol synthesis. This work provides an efficient method to identify key proteins at inter-organelle membrane contacts in living cells.

Keywords: Biophysics; Membranes; Proteomics.