RESUMEN
During endosomal recycling, Sorting Nexin 17 (SNX17) facilitates the transport of numerous membrane cargo proteins by tethering them to the Retriever complex. Despite its importance, the mechanisms underlying this interaction have remained elusive. Here, we report the structure of the Retriever-SNX17 complex determined using cryogenic electron microscopy (cryo-EM). Our structure reveals that the C-terminal tail of SNX17 engages with a highly conserved interface between the VPS35L and VPS26C subunits of Retriever. Through comprehensive biochemical, cellular, and proteomic analyses, we demonstrate that disrupting this interface impairs the Retriever-SNX17 interaction, subsequently affecting the recycling of SNX17-dependent cargos and altering the composition of the plasma membrane proteome. Intriguingly, we find that the SNX17-binding pocket on Retriever can be utilized by other ligands that share a consensus acidic C-terminal tail motif. By showing how SNX17 is linked to Retriever, our findings uncover a fundamental mechanism underlying endosomal trafficking of critical cargo proteins and reveal a mechanism by which Retriever can engage with other regulatory factors.
RESUMEN
The normally nuclear HNRNPH2 is mutated in HNRNPH2 -related X-linked neurodevelopmental disorder causing the protein to accumulate in the cytoplasm. Interactions of HNRNPH2 with its importin Karyopherin-ß2 (Transportin-1) had not been studied. We present a structure that shows Karyopherin-ß2 binding HNRNPH2 residues 204-215, a proline-tyrosine nuclear localization signal or PY-NLS that contains a typical R-X 2-4 -P-Y motif, 206 RPGPY 210 , followed a new Karyopherin-ß2 binding epitope at 211 DRP 213 that make many interactions with Karyopherin-ß2 W373. Mutations at each of these sites decrease Karyopherin-ß2 binding affinities by 70-100 fold, explaining aberrant accumulation in cells and emphasizing the role of nuclear import defects in the disease. Sequence/structure analysis suggests that the new epitope C-terminal of the PY-motif, which binds Karyopherin-ß2 W373, is rare and thus far limited to close paralogs HNRNPH2, HNRNPH1 and HNRNPF. Karyopherin-ß2 W373, a HNRNPH2-binding hotspot, corresponds to W370 of close paralog Transportin-2, a site of pathological variants in patients with neurodevelopmental abnormalities, suggesting that Transportin-2-HNRNPH2/H1/F interactions may be compromised in the abnormalities. Summary: HNRNPH2 variants in HNRNPH2 -related X-linked neurodevelopmental disorder aberrantly accumulate in the cytoplasm. A structure of Karyopherin-ß2â¢HNRNPH2 explains nuclear import defects of the variants, reveals a new NLS epitope that suggests mechanistic changes in pathological variants of Karyopherin-ß2 paralog Transportin-2.
