Nuclear pore targeting of the yeast Pom33 nucleoporin depends on karyopherin and lipid binding

作者：Floch Aurelie G; Tareste David; Fuchs Patrick F J; Chadrin Anne; Naciri Ikrame; Leger Thibaut; Schlenstedt Gabriel; Palancade Benoit; Doye Valerie^*

来源：Journal of Cell Science, 2015, 128(2): 305-316.

DOI：10.1242/jcs.158915

• Nuclear pore complex
• NPC
• Integral membrane protein
• Karyopherin
• Amphipathic helix
• NPC targeting
• NPC distribution
• TMEM33 family

Pom33 is an integral membrane protein of the yeast nuclear pore complex (NPC), and it is required for proper NPC distribution and assembly. To characterize the Pom33 NPC-targeting determinants, we performed immunoprecipitation experiments followed by mass spectrometry analyses. This identified a new Pom33 partner, the nuclear import factor Kap123. In vitro experiments revealed a direct interaction between the Pom33 C-terminal domain (CTD) and Kap123. In silico analysis predicted the presence of two amphipathic alpha-helices within Pom33-CTD. Circular dichroism and liposome co-flotation assays showed that this domain is able to fold into alpha-helices in the presence of liposomes and preferentially binds to highly curved lipid membranes. When expressed in yeast, under conditions abolishing Pom33-CTD membrane association, this domain behaves as a Kap123-dependent nuclear localization signal (NLS). Although deletion of Pom33 C-terminal domain (Pom(Delta DCTD)-GFP) impaired Pom33 stability and NPC targeting, mutants affecting either Kap123 binding or the amphipathic properties of the alpha-helices did not display any detectable defect. However, combined impairment of lipid and Kap123 binding affects targeting of Pom33 to NPCs. These data highlight the requirement of multiple determinants and mechanisms for proper NPC localization of Pom33.

• 出版日期2015-1-15