Whereas different importin α paralogs share the ability to bind and import classical NLS substrates only certain animal isoforms can recognize specific non-classical cargos.
In this respect it is noteworthy that there are five importin-α isoforms in mammalian cells; while all isoforms are capable of recognizing cNLSs, some isoforms show striking substrate specificity for non-classical cargoes.
This showed that the stapling procedure did not interfere dramatically with the transport of classical anterograde cargo.
Interestingly, in the crystal structure of PTHrP-ncNLS bound to importin β, the site of phosphorylation at Thr85 is located in close proximity to two well conserved tryptophan residues of importin β, which are known to be important for nuclear import of classical NLS cargos [ 125].
We used the classical caveolae-dependent cargo protein BSA-FITC [ 43, 44] to perform endocytosis assays in T-FGM AhR+/+and AhR−/− cells.
Although Sec16 is thought to be required for ER export, the effect of Sec16 depletion on the transport of the classical transmembrane secretory cargo tsO45-G from the ER is fairly minimal; tsO45-G can still exit the ER but at a slower rate.
In the presence of hematoxylin, the interaction between importin α and HIV1 Vpr NLS is inhibited in a dose-dependent manner, whereas nuclear import of classical NLS-containing cargos is not affected by hematoxylin (Suzuki et al. 2009).
By enlargement, classical early endosomes mix cargo materials and form several membrane domains of maturation.
Many of the endogenous cargos for classical nuclear import interact with other cellular components, either in the nucleus, the cytoplasm or both.
In the more well-defined classical transport pathway, NLS-cargo proteins interact with an adapter protein belonging to the importin α family, which in turn forms a heterocomplex with importin β1 for nuclear transport [16], [20].
Unlike a unified route for classical protein secretion, leaderless cargoes undergoing unconventional secretion employ multiple means of protein delivery, the details of which are largely unknown (Ding et al., 2012; Nickel, 2010; Rabouille et al., 2012; Zhang and Schekman, 2013).
