TY  - GEN
UR  - https://archiv.ub.uni-heidelberg.de/volltextserver/10678/
AV  - public
N2  - Ran GTPase has a well characterized role in interphase and metaphase.  Due to high levels of RanGTP in proximity to chromatin, import receptors are  dissociated from nuclear localization signal (NLS) bearing proteins in interphase  and mitosis, thus facilitating both nuclear import and mitotic spindle assembly.  In this thesis we investigated if Ran GTPase regulates the nucleoporin Pom121  through a similar mechanism during post-mitotic nuclear reassembly.  We found that importin !/" interacts with both of the two predicted NLS  sites of Xenopus Pom121 and is released by RanGTP. Importin !/" binding is  completely abolished upon mutation of NLS sites, which also affected binding of  a group of nucleoporins to Pom121. The NLS sites and transmembrane domain  of Xenopus Pom121 are required for correct nuclear envelope (NE) localization  of Pom121 in human U2OS cell lines. In these cells, RNAi depletion of human  Pom121 greatly reduced cell viability and diminished the levels of a subset of  nucleoporins detected by the monoclonal antibody 414 (mAb414). The signal  was present, however, if the human Pom121 was knocked down in a cell line  stably expressing wild-type Xenopus Pom121, while cell viability remained low.  Furthermore, in the absence of human Pom121, stable expression of an NLS  mutant form of Xenopus Pom121 decreased cell viability even more than U2OS  cells depleted of endogenous Pom121. In cells depleted of human Pom121,  expression of NLS mutant Xenopus Pom121 restored mAb414 levels as  wildtype Xenopus Pom121 expressing cells. However, these NLS mutant  Xenopus Pom121 expressing cells displayed decreased nuclear import kinetics  compared to the cells expressing wildtype Xenopus Pom121. Electron  microscopy analysis showed that wild-type Xenopus Pom121 localized at the  NPCs in U2OS cells. In contrast, the NLS mutant Xenopus Pom121 localized in  cytoplasmic membrane stacks interestingly together with other NPC  components. Despite the presence of NPC components, these membrane  stacks lacked NPC-like structures.   Taken together, this data shows that the importin !/" binding sites on  Pom121 are important for the NE localization of the protein and its interaction  with other nucleoporins. A Pom121 mutant defective in importin binding results  in nuclear transport defective pores and induces the formation of cytoplasmic  membrane stacks which lack NPC-like structures. We propose that the importin  !/"-Pom121 interaction is important for the formation of proper NPC function  and structure at the NE and in cytoplasmic membrane stacks. 
A1  - Yavuz, Emine Sevil
ID  - heidok10678
Y1  - 2009///
TI  - Interplay between the transmembrane nucleoporin Pom121 and the Ran GTPase system
KW  - Pom121 
KW  -  nucleoporin 
KW  -  importins 
KW  -  nuclear envelope 
KW  -  NPC
ER  -