Discovery of an N-terminal Amphipathic Phase Code for Exosome Targeting Using Dynamical Geometric Embeddings

Protein targeting to extracellular vesicles and exosomes remains poorly understood, with few generalizable sequence determinants beyond classical signal peptides and transmembrane domains. This work reports the discovery of a previously uncharacterized short N-terminal targeting signal associated with exosome localization, identified using a deterministic dynamical geometric embedding framework (PhaseOS). A single latent embedding coordinate reproducibly discriminates exosome-annotated proteins in the CAFA6 dataset. Through systematic perturbation, phase rotation, masking, positional jitter, reduced-alphabet remapping, and targeted motif knockout and insertion experiments, the signal is localized to an 8–12 residue N-terminal window exhibiting strong phase sensitivity and short-period amphipathic structure. Reduced-alphabet log-odds analysis reveals a small family of degenerate residue grammars, exemplified by patterns including HGCP, HGPC, HCGP, and HCPG, which are enriched in independently annotated UniProt exosome proteins. These findings support the existence of a phase-encoded amphipathic targeting mechanism distinct from canonical signal peptides. The identified signal, termed the Amphipathic Phase EXosome (APEX) phase code, suggests that vesicle targeting can be mediated by short, phase-encoded amphipathic helices and demonstrates the utility of dynamical geometric embeddings for uncovering latent biophysical targeting rules not captured by conventional sequence models.