Endosomal Organizers of post-Golgi trafficking in polarized epithelial cells
Epithelial cells are characterized by a polarized organization of their plasma membrane which is divided into apical and basolateral domains. This architecture is maintained by highly specific cargo sorting machinery that efficiently delivers newly synthesized polypeptides to their correct target me...
Klinische Zytobiologie und Zytopathologie
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|Zusammenfassung:||Epithelial cells are characterized by a polarized organization of their plasma membrane which is divided into apical and basolateral domains. This architecture is maintained by highly specific cargo sorting machinery that efficiently delivers newly synthesized polypeptides to their correct target membrane. After TGN exit apical cargo is segregated by at least two distinct sorting mechanisms into lipid-raft-dependent or lipid-raft-independent apical pathways in MDCK cells. The aim of this study was the identification of proteins which are essential for the transport of apically sorted proteins. In the first part of the current study, a member of kinesin-1 group, KIF5C, was identified as a kinesin motor for apical trafficking of sucrase-isomaltase, the marker for the raft-associated pathway, and of non-raft-associated p75. KIF5C was found by mass spectrometry in vesicle enriched fractions and on immunoisolated post-Golgi vesicles carrying apical cargo. KIF5C associates with vesicles of both raft-dependent and raft-independent pathways directly after TGN exit. The specific knockdown of KIF5C interfered the apical trafficking of both raft-associated and non-raft associated marker proteins significantly (Astanina and Jacob, 2010). In the second part, annexin XIIIb was identified in raft-independent apical trafficking by mass spectrometry, immunoblotting and confocal microscopy. Annexin XIIIb accumulated in endosomal compartments that are traversed by raft-dependent and raft-independent apical cargo after TGN release. Finally, a specific reduction of annexin XIIIb expression by RNA interference resulted in a significant decrease in the apical delivery of the raft- as well as non-raft apical markers (Astanina et al., 2010). Taken together, both proteins – KIF5C and annexin XIIIb – act as endosomal organizers of apical protein trafficking in polarized epithelial cells. Based on the confocal microscopy studies and TGN release experiments we came to the conclusion that both proteins function on the first transport steps after TGN exit and accomplish trafficking of raft-associated, as well as non-raft-associated apical cargo.|