Insights into the ER-mitochondria contact sites using protein interaction assay BioID

Identification: Aaltonen, Mari


Description

Insights into the ER-mitochondria contact sites using protein interaction assay BioID
 
Mari Aaltonen1, Hana Antonicka1, Alexandre Janer1, Zhen-Yuan Lin2, Anne-Claude Gingras2, Eric A. Shoubridge1*
1Montreal Neurological Institute, McGill University, Montreal, Canada; 2 Lunenfeld-Tanenbaum Research Institute & University of Toronto; Toronto, Ontario, Canada
*Corresponding Author (optional)
 
The membrane contact sites between endoplasmic reticulum (ER) and mitochondria are essential for several processes requiring organellar communication, such as mitochondrial division, autophagy and exchange of calcium and lipids between ER and mitochondria. However, it is not fully understood which proteins are present at these contact sites in mammals. To identify contact site proteins in human cells, we are using a novel protein interaction assay BioID. In BioID, the protein of interest is labelled with a modified bacterial biotin ligase BirA*. Inside the living cell, the ligase biotinylates its proximal proteins, which can then be affinity purified and detected by mass spectrometry. BioID is proving a powerful tool to reveal novel protein interactions and environments as it allows the detection of dynamic and long-range interactions. We set to test whether BioID can be used to detect interactions across membrane contact sites. First, we have looked at the interaction profile of mitochondrial proteins that have been previously characterized to play a role in ER-mitochondria tethering. PTPIP51 has been shown to interact with the ER proteins VAPB and ORP8 in classical co-immunoprecipitation experiments. Our BioID analysis of PTPIP51 protein confirmed these interactions, validating the use of BioID as a tool to study membrane contact sites. In addition, we found several novel interacting partners of PTPIP51, including subunits of the ER membrane protein complex EMC which has been linked to lipid transfer between ER and mitochondria in yeast. With the BioID as a tool, we are continuing to explore the ER-mitochondria interface to reveal the proteome of these contact sites.
 

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