A structurally defined nucleoplasmic-facing surface of Nup170 controls subtelomeric gene silencing.

The nuclear pore complex (NPC) mediates nucleocytoplasmic transport while contributing to nuclear organization and genome regulation. In Saccharomyces cerevisiae, the nucleoporin Nup170, positioned in the NPC's inner ring, plays a critical role in subtelomeric gene silencing, telomere tethering to the nuclear envelope, and association of silencing factors with subtelomeric chromatin. However, the molecular basis of these functions and their relationship to NPC architecture remain unclear. Here, we define a nucleoplasm-exposed region of Nup170 required for subtelomeric gene silencing. Systematic truncation analysis reveals that Nup170 residues between ∼990 and 1200 are necessary to restore silencing, telomere positioning, and subtelomeric gene expression in nup170Δ cells. Mapping these truncations onto high-resolution structural models of the NPC places this region on a nucleoplasmic-facing surface of the inner ring, accessible through openings in the outer ring scaffold. Targeted single amino acid substitutions within this region, including nup170D1099G, phenocopy the subtelomeric silencing defect, telomere tethering defect, and reduced SIR complex binding of nup170Δ. These results demonstrate that a short, structurally defined surface of Nup170 within the NPC scaffold directly supports epigenetic regulation at the nuclear periphery. Our findings reveal how architectural nucleoporins perform specific genome regulatory functions through discrete interfaces, highlighting functional specialization among individual NPCs.