Transport and InsP₈ gating mechanisms of the human inorganic phosphate exporter XPR1

Inorganic phosphate (Pi) has essential metabolic and structural roles in living organisms. The Pi exporter, XPR1/SLC53A1, is critical for cellular Pi homeostasis. When intercellular Pi is high, cells accumulate inositol pyrophosphate (1,5-InsP₈), a signaling molecule required for XPR1 function. Inactivating XPR1 mutations lead to brain calcifications, causing neurological symptoms including movement disorders, psychosis, and dementia. Here, cryo-electron microscopy structures of dimeric XPR1 and functional characterization delineate the substrate translocation pathway and how InsP₈ initiates Pi transport. Binding of InsP₈ to XPR1, but not the related inositol polyphosphate InsP₆, rigidifies the intracellular SPX domains, with InsP₈ bridging the dimers and SPX and transmembrane domains. Locked in this state, the C-terminal tail is sequestered, revealing the entrance to the transport pathway, thus explaining the obligate roles of the SPX domain and InsP₈. Together, these findings advance our understanding of XPR1 transport activity and expand opportunities for rationalizing disease mechanisms and therapeutic intervention.