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Updated: Apr 17, 2026

Purification and Quality Control of Recombinant Septin Complexes for Cell-Free Reconstitution
Published on: June 23, 2022
Septins associate with AP-3 to support trafficking to the vacuole/lysosome in yeast
Mitchell Leih1, Michaela McCright1, Cortney Angers2
1Department of Molecular Cellular and Developmental Biology, University of Colorado, Boulder, CO 80309.
Abstract:
Adaptor protein complex 3 (AP-3) mediates clathrin-independent transport to lysosomes, yet accessory factors supporting this pathway remain incompletely defined. In Saccharomyces cerevisiae, the C-terminal intrinsically disordered regions (IDRs) of both AP-3 large subunits (δ and β3) serve as platforms for association with accessory factors. Through proteomic analysis of proteins associated with these IDRs, we identify the septin cytoskeleton as a candidate AP-3-associated factor. Bimolecular fluorescence complementation (BiFC) reveals a hierarchical pattern of association: AP-3 shows preferential proximity to core septin subunits (Cdc10, Cdc3, and Cdc12) over terminal subunits (Cdc11 and Shs1). These terminal subunits serve as alternative caps of septin octamers, generating structurally distinct assemblies. Significantly, dysfunction of Cdc11 but not Shs1 selectively impairs AP-3-dependent cargo sorting without affecting the parallel vacuolar protein sorting (VPS) pathway to the vacuole (lysosome in yeast), providing genetic evidence for a specific functional connection between Cdc11-containing septin assemblies and AP-3-mediated transport.
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