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Photocatalytic Proximity Labeling Coupled with Cross-linking for Deciphering of the Lysosome Proteome in Living Cells

ABSTRACT: Lysosomes are essential organelles that play vital roles in the degradation and recycling of a variety of biomolecules. To fully understand their molecular mechanisms, high spatiotemporal identification of the lysosome proteome in living cells is critical. Although photocatalytic proximity labeling is a powerful tool for profiling subcellular proteome, it often struggles with low labeling efficiency, particularly for low-abundance proteins. To overcome this, we introduced a novel strategy that integrates photocatalytic proximity labeling with cross-linking. This approach enabled difficult-to-label proteins to be linked to already labeled ones via cross-linking, thereby expanding the scope of pro-tein identification. Using this strategy, we successfully identified 238 lysosome-annotated proteins in living HeLa cells, a substantial in-crease compared to method without cross-linking (238 vs 197). The developed strategy enhanced lysosomal proteomic coverage via a high-precision integration platform, allowing systematic identification of both membrane-associated and luminal proteins within lysosome, thereby providing mechanistic insights into subcellular biology and unlocking new frontiers in cellular research.

ORGANISM(S): Homo Sapiens

SUBMITTER: Lihua Zhang

PROVIDER: PXD063423 | iProX | Tue Apr 29 00:00:00 BST 2025

REPOSITORIES: iProX

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