Receptor-mediated endocytosis involves tyrosine phosphorylation of cortactin.
Efficient internalization of cell surface receptors requires actin polymerization mediated by Arp2/3 complex and cortactin, a prominent substrate of the protein-tyrosine kinase Src. However, the significance of cortactin tyrosine phosphorylation in endocytosis is unknown. We found that overexpression of a cortactin mutant deficient in tyrosine phosphorylation decreased transferrin uptake. Suppression of cortactin expression by RNA interference also reduced transferrin internalization. Such inhibition was effectively rescued by overexpressing wild-type cortactin but not a cortactin mutant deficient in tyrosine phosphorylation or a mutant with deletion of the Src homology 3 domain. Likewise, purified phosphorylation-null cortactin failed to restore the formation of clathrin-coated vesicles in a cortactin-depleted cell extract. In vitro analysis revealed that Src-mediated phosphorylation enhanced the association of cortactin with dynamin-2 in a tyrosine phosphorylation-dependent manner. Quantitative analysis demonstrated that Src enhances the affinity of cortactin for dynamin-2 by more than 3-fold. On the other hand, Src-treated dynamin-2 had no effect on its interaction with cortactin. These data indicate that Src kinase is implicated in clathrin-mediated endocytosis by phosphorylation of cortactin.
Zhu
Jianwei
J
Affiliated Hospital of Nantong University, 226001 Nantong, China.
Yu
Dan
D
Zeng
Xian-Chun
XC
Zhou
Kang
K
Zhan
Xi
X
D051376:Actin-Related Protein 2-3 Complex / Q000378:metabolism; D000595:Amino Acid Sequence / Q000235:genetics; D002966:Clathrin / Q000378:metabolism; D051356:Cortactin / Q000235:genetics / Q000378:metabolism*; D034285:Dynamin II / Q000378:metabolism; D004705:Endocytosis / Q000502:physiology*; D006367:HeLa Cells; D006801:Humans; D010766:Phosphorylation; D011499:Protein Processing, Post-Translational / Q000502:physiology*; D017434:Protein Structure, Tertiary / Q000235:genetics; D034622:RNA Interference; D011940:Receptor Aggregation / Q000502:physiology; D011990:Receptors, Transferrin / Q000378:metabolism; D017384:Sequence Deletion; D014168:Transferrin / Q000378:metabolism; D014443:Tyrosine / Q000378:metabolism; D019061:src-Family Kinases / Q000378:metabolism
D051376:Actin-Related Protein 2-3 Complex; D002966:Clathrin; D051356:Cortactin; D011990:Receptors, Transferrin; D014168:Transferrin; D014443:Tyrosine; D019061:src-Family Kinases; D034285:Dynamin II