A dominant-negative form of POM121 binds chromatin and disrupts the two separate modes of nuclear pore assembly

Abstract

Nuclear pore complexes (NPCs) are formed during two separate stages of the metazoan cell cycle. They are assembled into the re-forming nuclear envelope (NE) at the exit from mitosis and into an intact, expanding NE during interphase. Here, we show that a soluble internal fragment of the membrane nucleoporin POM121 has a dominant-negative effect on both modes of assembly in a cell-free reconstitution system. The soluble POM121 fragment binds chromatin at sites that are distinct from ELYS–Nup107–160 ‘seeding’ sites and prevents membrane enclosure and NPC formation. Importin-β negatively regulates chromatin binding by the POM121 fragment through a conserved NLS motif and is also shown to affect the recruitment of the endogenous membrane protein to chromatin in the full assembly system. When an intact NE is present before the addition of the dominant-negative fragment, NPCs are inserted into the NE but membrane expansion is inhibited. This results in densely packed NPCs with no intervening membrane patches, as visualized by scanning electron microscopy. We conclude that POM121 plays an important role in both modes of assembly and links nuclear membrane formation and expansion to nuclear pore biogenesis.