INTERACTIONS OF DROSOPHILA CBL WITH EPIDERMAL GROWTH-FACTOR RECEPTORSAND ROLE OF CBL IN R7 PHOTORECEPTOR CELL-DEVELOPMENT

The human proto-oncogene product c-Cbl and a similar protein in Caenor habditis elegans (Sli-1) contain a proline-rich COOH-terminal region t hat binds Src homology 3 (SH3) domains of proteins such as the adapter Grb2. Cbl-Grb2 complexes can be recruited to tyrosine-phosphorylated epidermal growth factor (EGF) receptors through the SH2 domain of Grb2 . Here we identify by molecular cloning a Drosophila cDNA encoding a p rotein (Drosophila Cbl [D-Cbl]) that shows high sequence similarity to the N-terminal region of human c-Cbl but lacks proline-rich sequences and fails to bind Grb2. Nonetheless, in COS-1 cells, expression of he magglutinin epitope-tagged D-Cbl results in its coimmunoprecipitation with EGF receptors in response to EGF. EGF also caused tyrosine phosph orylation of D-Cbl in such cells, but no association of phosphatidylin ositol 3-kinase was detected in assays using anti-p85 antibody. A poin t mutation in D-Cbl (G305E) that suppresses the negative regulation of LET-23 by the Cbl homolog Sli-1 in C. elegans prevented tyrosine phos phorylation of D-Cbl as well as binding to the liganded EGF receptor i n COS-1 cells. Colocalization of EGF receptors with both endogenous c- Cbl or expressed D-Cbl in endosomes of EGF-treated COS-1 cells is also demonstrated by immunofluorescence microscopy. In lysates of adult tr ansgenic Drosophila melanogaster, GST-DCbl binds to the tyrosine-phosp horylated 150-kDa torso-DER chimeric receptor. Expression of D-Cbl dir ected by the sevenless enhancer in intact Drosophila compromises sever ely the development of the R7 photoreceptor neuron. These data suggest that despite the lack of Grb2 binding sites, D-Cbl functions as a neg ative regulator of receptor tyrosine kinase signaling in the Drosophil a eye by a mechanism that involves its association with EGF receptors or other tyrosine kinases.