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-β2 (Transportin-1) had not been studied. We present a structure that shows Karyopherin-β2 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-β2 binding epitope at 211 DRP 213 that make many interactions with Karyopherin-β2 W373. Mutations at each of these sites decrease Karyopherin-β2 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-β2 W373, is rare and thus far limited to close paralogs HNRNPH2, HNRNPH1 and HNRNPF. Karyopherin-β2 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-β2•HNRNPH2 explains nuclear import defects of the variants, reveals a new NLS epitope that suggests mechanistic changes in pathological variants of Karyopherin-β2 paralog Transportin-2.