Beyond the Nuclear Pore Complex, Nup358 Clusters at the Axon Initial Segment of Cultured Neurons

Nup358 is the largest nucleoporin of around 358 kDa molecular weight that localizes on the cytoplasmic face of the nuclear pore complex (NPC). It takes part in the overall activity of the NPC mediating the transport of nucleic acids and proteins between the nucleus and the cytoplasm. However, due to its multi-domain configuration, Nup358 has a more pleiotropic function in several cellular mechanisms such as mediating the stability of microtubules and axon specification. Since little is known about the non-conventional role of Nup358 in neuronal polarity, my PhD thesis was focused on characterizing the subcellular distribution and expression pattern of Nup358 protein in cultured neurons. My results show Nup358 present at the nuclear rim of neuronal cells, associated with NPCs, also in the cytoplasm having a spotted pattern along the neuronal processes. Interestingly, Nup358 was remarkably clustered at the axon initial segment (AIS) of mature neurons and dependent on a prior recruitment of the master AIS scaffold, Ankyrin-G (AnkG), to this specific region. Of the distinct domains present in Nup358 protein, the N-terminal region was found to be crucial for its localization at the AIS. Further, changes in Nup358 protein expression were monitored during neuronal development. Indeed, I detected the presence of a shorter isoform of Nup358 that was increasing as neurons develop whereas the full-length protein had an opposite decreasing trend. To gain knowledge about the functional role of Nup358 in neurons, I investigated its protein expression/distribution in response to an increasing or decreasing neuronal activity with specific drug treatments. Surprisingly, Nup358 protein expression was reduced following the stimulation or the depolarization of neurons, mediated by calcium influx and NMDA receptors. Overall, my results show that Nup358 has a unique subcellular distribution in neurons, being enriched at the AIS at advanced stages of development, therefore, suggest an involvement of Nup358 protein in maintaining and modulating neuronal polarity and activity.