c-Src trafficking and co-localization with the EGF receptor promotes EGF ligand-independent EGF receptor activation and signaling.
c-Src is a non-receptor tyrosine kinase that associates with both the plasma membrane and endosomal compartments. In many human cancers, especially breast cancer, c-Src and the EGF receptor (EGFR) are overexpressed. Dual overexpression of c-Src and EGFR correlates with a Src-dependent increase in activation of EGFR, and synergism between these two tyrosine kinases increases the mitogenic activity of EGFR. Despite extensive studies of the functional interaction between c-Src and EGFR, little is known about the interactions in the trafficking pathways for the two proteins and how that influences signaling. Given the synergism between c-Src and EGFR, and the finding that EGFR is internalized and can signal from endosomes, we hypothesized that c-Src and EGFR traffic together through the endocytic pathway. Here we use a regulatable c-SrcGFP fusion protein that is a bona fide marker for c-Src to show that c-Src undergoes constitutive macropinocytosis from the plasma membrane into endocytic compartments. The movement of c-Src was dependent on its tyrosine kinase activity. Stimulation of cells with EGF revealed that c-Src traffics into the cell with activated EGFR and that c-Src expression and kinase activity prolongs EGFR activation. Surprisingly, even in the absence of EGF addition, c-Src expression induced activation of EGFR and of EGFR-mediated downstream signaling targets ERK and Shc. These data suggest that the synergy between c-Src and EGFR also occurs as these two kinases traffic together, and that their co-localization promotes EGFR-mediated signaling.
Donepudi
Mrudula
M
Cell Biology Program, Memorial Sloan-Kettering Cancer Center, New York, NY 10065, United States.
Resh
Marilyn D
MD
D000818:Animals; D019556:COS Cells; D002462:Cell Membrane / Q000187:drug effects / Q000201:enzymology; D002645:Chickens; D002522:Chlorocebus aethiops; D011992:Endosomes / Q000187:drug effects / Q000201:enzymology; D004789:Enzyme Activation / Q000187:drug effects; D004815:Epidermal Growth Factor / Q000494:pharmacology*; D066246:ErbB Receptors / Q000378:metabolism*; D017930:Genes, Reporter; D049452:Green Fluorescent Proteins / Q000378:metabolism; D042541:Intracellular Space / Q000187:drug effects / Q000201:enzymology; D008024:Ligands; D051379:Mice; D041681:NIH 3T3 Cells; D010873:Pinocytosis / Q000187:drug effects; D021381:Protein Transport / Q000187:drug effects; D016392:Proto-Oncogene Proteins pp60(c-src) / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D015398:Signal Transduction / Q000187:drug effects*; D013347:Subcellular Fractions / Q000187:drug effects; D033921:Vesicular Transport Proteins / Q000378:metabolism
D008024:Ligands; D011993:Recombinant Fusion Proteins; D033921:Vesicular Transport Proteins; C093846:early endosome antigen 1; D049452:Green Fluorescent Proteins; D004815:Epidermal Growth Factor; D066246:ErbB Receptors; D016392:Proto-Oncogene Proteins pp60(c-src)