Cross-linking of the endolysosomal system reveals potential flotillin structures and cargo

Lysosomes are well-established as the main cellular organelles for the degradation of macromolecules and emerging as regulatory centers of metabolism. They are of crucial importance for cellular homeostasis, which is exemplified by a plethora of disorders related to alterations in lysosomal function. In this context, protein complexes play a decisive role, regulating not only metabolic lysosomal processes but also lysosome biogenesis, transport, and interaction with other organelles. Using cross-linking mass spectrometry, we analyze lysosomes and early endosomes. Based on the identification of 5376 cross-links, we investigate protein-protein interactions and structures of lysosome- and endosome-related proteins. In particular, we present evidence for a tetrameric assembly of the lysosomal hydrolase PPT1 and a heterodimeric structure of FLOT1/FLOT2 at lysosomes and early endosomes. For FLOT1-/FLOT2-positive early endosomes, we identify >300 putative cargo proteins and confirm eleven substrates for flotillin-dependent endocytosis, including the latrophilin family of adhesion G protein-coupled receptors. Protein complexes play a decisive role for lysosomal function. Here, the authors use cross-linking mass spectrometry and integrative modeling to investigate lysosomes and early endosomes; characterizing protein interactions, structures, and the cargo of flotillin-mediated endocytosis.

Automated summary

Lysosomes are crucial organelles for degradation and metabolism regulation. Protein complexes play a decisive role in lysosomal function. This study investigates lysosomes and early endosomes using cross-linking mass spectrometry, revealing protein interactions, structures, and cargo of flotillin-mediated endocytosis.