How Nuclear Pore Complexes Form in the First Stages of Life
A reservoir of maternally provided NPCs in the ER enables nuclear divisions during early embryogenesis
To the point
- In oocytes cytoplasmic NPCs form from precursor condensates
- These cytoplasmic NPCs are inherited by the embryo and insert into nuclei
- The cellular environment defines if NPCs are symmetric or asymmetric
The Nuclear Pore Complex (NPC) is the largest non-polymeric protein complex in a eukaryotic cell. It is built from a set of ~30 nucleoporins (Nups) that assemble in a stereotyped, repetitive fashion into a mature complex that in humans comprises up to ~1000 proteins.
Evidently assembling such giant architectures is an intricate task. While significant progress has been made to understand NPC assembly in cultured cell lines and in yeast, information on how nuclear pores form in a developing animal is very limited. Does the mode of NPC assembly obey to different developmental requirements? Do pore number or composition adjust at developmental transitions?
We leverage Drosophila as a genetically tractable model system and apply a combination of live-imaging, biochemistry and in situ structural biology to address the cell biology of NPC assembly during the earliest steps of development.
In embryos and oocytes across animal species NPCs do not only populate the nuclear envelope (NE), but also stacked ER sheets termed annulate lamellae (AL, Figure 1). This compartment was already identified in the 1950s by classical transmission Electron Microscopy, but its function has remained elusive. Over the last years our work could contribute novel insights into the biogenesis, fate and architecture of these cytoplasmic pores.
NPCs in the ooplasm assemble from nucleoporin condensates
In all animals earliest embryogenesis is ensured primarily by maternally provided components that are stored in the mother’s oocyte. A reservoir of pre-assembled cytoplasmic NPCs forms already in the ooplasm of the mother. Both known NPC assembly pathways involve a nuclear compartment, so how can these NPCs form in the cytoplasm, away from the oocyte nucleus?
Our findings suggest that in flies, Nups condense into phase separated granules that serve as spatial cue to form NPCs on ER membranes, alleviating the need of a nucleus [Hampoelz et al., Cell 2019 and comment]. Condensates of the cytoplasmic NPC component Nup358 have a key role, as they harbor several Nups en route to the oocyte. How nucleoporin condensates interact with each other and with membranes to eventually form cytoplasmic NPCs is not known.
Preassembled cytoplasmic NPCs insert into the nuclear envelope
Upon fertilization this pool of ooplasmic cNPCs is inherited by the embryo and undergoes cycles of disassembly and reformation at each mitosis (Figure 2). By imaging nucleoporins in living embryos, we could capture that cytoplasmic NPCs insert in batch into the nuclear envelope when the nuclear surface expands during the very fast interphases of the syncytial blastoderm. We deciphered a topological model how his might work without compromising the impermeability of the nuclear envelope [Hampoelz et al., Cell 2016 and comment]. This insertion process is prominent before the induction of the zygotic genome but ceases later in development. Thus a developmental regulation of NPC assembly is likely, yet we lack molecular understanding.
Regulation of NPC (a)symmetry
Transport dependent and independent functions of NPCs rely on the asymmetric distribution of compartment specific Nups to the symmetric core scaffold of the complex. By focusing on NPCs that are located outside of the NE, we shed light on how this asymmetry is established [Sachweh et al., Cell 2025].
Vitrification and cryo-Electron Tomography of isolated cells from Drosophila ovaries allowed us to decipher that in contrast to the asymmetric NPCs at the NE, pores entirely surrounded by cytoplasm are symmetric. This indicates that the peripheral Nup configuration of the NPC is determined by the surrounding cellular milieu. We also found that the nucleotide state of the small GTPase Ran, a distinguishing factor between the nuclear and the cytoplasmic compartment, molecularly regulates which set of Nups binds to the symmetric core scaffold. Thus the same molecular mechanism controls nuclear transport and the composition of the transport channel itself.