Journal article
Interacting partners of Golgi-localized small G protein Arl5b identified by a combination of in vivo proximity labelling and GFP-Trap pull down
FJ Houghton, C Makhoul, EHJ Cho, NA Williamson, PA Gleeson
FEBS Letters | Published : 2022
Abstract
The small G protein Arl5b is localised on the trans-Golgi network (TGN) and regulates endosomes-to-TGN transport. Here, we combined in vivo and in vitro techniques to map the interactive partners and near neighbours of Arl5b at the TGN, using constitutively active, membrane-bound Arl5b(Q70L)-GFP in stably expressing HeLa cells, and the proximity labelling techniques BioID and APEX2 in parallel with GFP-Trap pull down. From MS analysis, 22 Golgi proteins were identified; 50% were TGN-localised Rabs, Arfs and Arls. The scaffold/tethering factors ACBD3 (GCP60) and PIST (GOPC) were also identified, and we show that Arl5b is required for TGN recruitment of ACBD3. Overall, the combination of in vi..
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Awarded by Australian Research Council
Funding Acknowledgements
Confocal Microscopy was performed at the Biological Optical Microscopy Platform (BOMP), University of Melbourne and mass spectrometry in the Bio21 Mass Spectrometry and Proteomics Facility (MSPF) at the Bio21 Molecular Science and Biotechnology Institute, University of Melbourne. This work was supported by funding from the Australian Research Council (DP160102394). We thank Dr Ismail Mahmond for the construction of Arl1-myc. CM was supported by a University of Melbourne Graduate Scholarship. Open access publishing facilitated by The University of Melbourne, as part of the Wiley -The University of Melbourne agreement via the Council of Australian University Librarians.