Spatial proteomics defines the content of trafficking vesicles captured by golgin tethers.

Shin JJH, Crook OM, Borgeaud AC, Cattin-Ortolá J, Peak-Chew SY, Breckels LM, Gillingham AK, Chadwick J, Lilley KS, Munro S

Intracellular traffic between compartments of the secretory and endocytic pathways is mediated by vesicle-based carriers. The proteomes of carriers destined for many organelles are ill-defined because the vesicular intermediates are transient, low-abundance and difficult to purify. Here, we combine vesicle relocalisation with organelle proteomics and Bayesian analysis to define the content of different endosome-derived vesicles destined for the trans-Golgi network (TGN). The golgin coiled-coil proteins golgin-97 and GCC88, shown previously to capture endosome-derived vesicles at the TGN, were individually relocalised to mitochondria and the content of the subsequently re-routed vesicles was determined by organelle proteomics. Our findings reveal 45 integral and 51 peripheral membrane proteins re-routed by golgin-97, evidence for a distinct class of vesicles shared by golgin-97 and GCC88, and various cargoes specific to individual golgins. These results illustrate a general strategy for analysing intracellular sub-proteomes by combining acute cellular re-wiring with high-resolution spatial proteomics.

Keywords:

Hela Cells

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Endosomes

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trans-Golgi Network

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Mitochondria

,

Humans

,

Membrane Proteins

,

Autoantigens

,

Proteomics

,

Gene Knockdown Techniques

,

HEK293 Cells

,

Spatial Analysis

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Golgi Matrix Proteins