التفاصيل البيبلوغرافية
| العنوان: |
A checkpoint function for Nup98 in nuclear pore formation suggested by novel inhibitory nanobodies. |
| المؤلفون: |
Solà Colom, Mireia, Fu, Zhenglin, Gunkel, Philip, Güttler, Thomas, Trakhanov, Sergei, Srinivasan, Vasundara, Gregor, Kathrin, Pleiner, Tino, Görlich, Dirk |
| المصدر: |
EMBO Journal; Jun2024, Vol. 43 Issue 11, p2198-2232, 35p |
| مصطلحات موضوعية: |
NUCLEAR pore complex, IMMUNOGLOBULINS, NUCLEOCYTOPLASMIC interactions, NUCLEAR transport, NUCLEAR membranes, EUKARYOTIC cells, ARTIFICIAL satellite tracking |
| مستخلص: |
Nuclear pore complex (NPC) biogenesis is a still enigmatic example of protein self-assembly. We now introduce several cross-reacting anti-Nup nanobodies for imaging intact nuclear pore complexes from frog to human. We also report a simplified assay that directly tracks postmitotic NPC assembly with added fluorophore-labeled anti-Nup nanobodies. During interphase, NPCs are inserted into a pre-existing nuclear envelope. Monitoring this process is challenging because newly assembled NPCs are indistinguishable from pre-existing ones. We overcame this problem by inserting Xenopus-derived NPCs into human nuclear envelopes and using frog-specific anti-Nup nanobodies for detection. We further asked whether anti-Nup nanobodies could serve as NPC assembly inhibitors. Using a selection strategy against conserved epitopes, we obtained anti-Nup93, Nup98, and Nup155 nanobodies that block Nup–Nup interfaces and arrest NPC assembly. We solved structures of nanobody-target complexes and identified roles for the Nup93 α-solenoid domain in recruiting Nup358 and the Nup214·88·62 complex, as well as for Nup155 and the Nup98 autoproteolytic domain in NPC scaffold assembly. The latter suggests a checkpoint linking pore formation to the assembly of the Nup98-dominated permeability barrier. Synopsis: Nuclear pores complexes (NPCs) conduct nucleocytoplasmic transport in eukaryotic cells. This work introduces a nanobody toolkit for studying NPC biogenesis and describes a checkpoint that links assembly of the NPC scaffold to that of the NPC permeability barrier. Direct tracking of postmitotic NPC assembly with fluorescent nanobodies that recognize well-accessible Nup epitopes. Novel interphase NPC assembly assay using frog-specific anti-Nup nanobodies to track the insertion of new frog NPCs into a human template nucleus. Nanobodies that block Nup-Nup interactions and inhibit NPC assembly. Structures of Nup complexes with inhibitory nanobodies. Inhibitory anti-Nup98 nanobodies identify a checkpoint that avoids non-selective NPCs without a permeability barrier. A nanobody toolkit for tracking or blocking nuclear pore complex (NPC) biogenesis reveals that assembly of the NPC scaffold is linked to formation of a functional permeability barrier. [ABSTRACT FROM AUTHOR] |
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| قاعدة البيانات: |
Complementary Index |
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