The cell cortex distribution of SNX16 check FAQ is disrupted upon wortmannin treatment thus it is PI3 kinase dependent, which is consistent with the previous biochemical studies. SNX23KIF16B is another PX domain protein and it contains a kinesin domain which is usually involved in the microtubule filament dependent transport of cargos. Indeed, it has been demonstrated that SNX23 is able to regulate the microtubule dependent transport of FGFR containing vesicles or early endosomes. We found that a fraction of SNX23 co localizes with SNX16 at cell cortex and this observation suggests that SNX23 could be involved in the transport of SNX16 to cell cortex. We performed loss of function studies and revealed that SNX23 as well as the microtubule filaments are both required for the cell cortex transport of SNX16.

It is interesting to note that SNX16 does not localize to the LBPA containing multivesicular late endosomes in control Hela Inhibitors,Modulators,Libraries cells, how ever, it re distributes to this endosomes after the inhibition of microtubule. These observations suggest that a SNX23microtubule dependent transport route is critical for establishing proper subcellular distribution of SNX16. We tried but failed to detect a direct association between SNX16 and SNX23. Inhibitors,Modulators,Libraries It is possible that other adaptor pro teins are needed for the SNX23 mediated transport of SNX16. We report here that SNX16 plays a negative role during the migration or tumorigenesis of MCF 7 cells, but it is dispensable for the growth of these cells. Inhibitors,Modulators,Libraries SNX16 mediated vesicular trafficking is involved in signaling pathways such as EGF, BMP and Wnt pathways.

However, it is currently unknown whether Inhibitors,Modulators,Libraries or not these signaling pathways are in volved in cell migration or tumorigenesis in MCF 7 cells. Further studies are required to indentify the exact cargos associated with SNX16 during these processes. Conclusions SNX16 containing vesicles are identified near focal adhe sions at cell cortex in addition Inhibitors,Modulators,Libraries to their cytosolic distribu tion. The SNX23microtubule pathway and the PI3 kinase pathway are both required for the cell cortex distribution of SNX16. SNX16 negatively regulates cell migration in vitro and tumorigenesis in vivo. Methods Molecular cloning Molecular cloning was performed according to standard protocols. Human SNX16, SNX2 and Rab5 genes were amplified from cDNA and cloned into the eukaryotic expression vector pCR3.

1 uni tagged with FLAG, GFP FLAG or N GFP. SNX23 was purchased from FulenGen. SNX16 and SNX2 were subcloned into the lentivirus vec tor PlxnB for establishing stable cell lines. All constructs were confirmed by DNA sequencing. Detailed informa tion about these constructs is available upon request. Cell culture, transfection and small chemical treatment MCF 7, Hela, CT99021 NCI H460 and Bel7402 were cultured in RPMI 164010% FBS at 37 C with 5% CO2. HepG2 and 293T were cultured in DMEM10% FBS and GLC 82 was cultured in DMEM10% FBS plus 2 mM L glutamine.