Sorting nexin 9 participates in clathrin-mediated endocytosis through interactions with the core components.
Sorting nexin 9 (SNX9) belongs to a family of proteins, the sorting nexins, that are characterized by the presence of a subclass of the phosphoinositide-binding phox domain. SNX9 has in its amino terminus a Src homology 3 domain and a region with predicted low complexity followed by a carboxyl-terminal part containing the phox domain. We previously found that SNX9 is one of the major proteins in hematopoietic cells that binds to the alpha and beta2-appendages of adaptor protein complex 2 (AP-2), a protein with a critical role in the formation of clathrin-coated vesicles at the plasma membrane. In the present study we show that clathrin and dynamin-2, two other essential molecules in the endocytic process, also interact with SNX9. We found that both AP-2 and clathrin bind to the low complexity region in SNX9 in a cooperative manner, whereas dynamin-2 binds to the Src homology 3 domain. In the cytosol, SNX9 is present in a 14.5 S complex containing dynamin-2 and an unidentified 41-kDa protein. In HeLa cells, SNX9 co-localized with both AP-2 and dynamin-2 at the plasma membrane or on vesicular structures derived from it but not with the early endosomal marker EEA1 or with AP-1. The results suggest that SNX9 may be recruited together with dynamin-2 and become co-assembled with AP-2 and clathrin at the plasma membrane. Overexpression in both K562 and HeLa cells of truncated forms of SNX9 interfered with the uptake of transferrin, consistent with a role of SNX9 in endocytosis.
Lundmark
Richard
R
Department of Medical Biochemistry and Biophysics, Umeå University, S-901 87 Umeå, Sweden.
Carlsson
Sven R
SR
D033962:Adaptor Protein Complex 2 / Q000378:metabolism; D001665:Binding Sites; D002352:Carrier Proteins / Q000235:genetics / Q000378:metabolism* / Q000502:physiology; D002966:Clathrin / Q000378:metabolism*; D022163:Clathrin-Coated Vesicles / Q000378:metabolism; D034285:Dynamin II / Q000378:metabolism; D004705:Endocytosis; D006367:HeLa Cells; D006801:Humans; D020014:K562 Cells; D011485:Protein Binding; D017384:Sequence Deletion; D058305:Sorting Nexins; D014162:Transfection; D033921:Vesicular Transport Proteins; D018909:src Homology Domains
D033962:Adaptor Protein Complex 2; D002352:Carrier Proteins; D002966:Clathrin; C400533:SNX9 protein, human; D058305:Sorting Nexins; D033921:Vesicular Transport Proteins; D034285:Dynamin II