Enigma prevents Cbl-c-mediated ubiquitination and degradation of RETMEN2A.
The Cbl proteins (Cbl, Cbl-b, and Cbl-c) are a highly conserved family of RING finger ubiquitin ligases (E3s) that function as negative regulators of tyrosine kinases in a wide variety of signal transduction pathways. In this study, we identify a new Cbl-c interacting protein, Enigma (PDLIM7). This interaction is specific to Cbl-c as Enigma fails to bind either of its closely related homologues, Cbl and Cbl-b. The binding between Enigma and Cbl-c is mediated through the LIM domains of Enigma as removal of all three LIM domains abrogates this interaction, while only LIM1 is sufficient for binding. Here we show that Cbl-c binds wild-type and MEN2A isoforms of the receptor tyrosine kinase, RET, and that Cbl-c enhances ubiquitination and degradation of activated RET. Enigma blocks Cbl-c-mediated RETMEN2A ubiquitination and degradation. Cbl-c decreased downstream ERK activation by RETMEN2A and co-expression of Enigma blocked the Cbl-c-mediated decrease in ERK activation. Enigma showed no detectable effect on Cbl-c-mediated ubiquitination of activated EGFR suggesting that this effect is specific to RET. Through mapping studies, we show that Cbl-c and Enigma bind RETMEN2A at different residues. However, binding of Enigma to RETMENA prevents Cbl-c recruitment to RETMEN2A. Consistent with these biochemical data, exploratory analyses of breast cancer patients with high expression of RET suggest that high expression of Cbl-c correlates with a good outcome, and high expression of Enigma correlates with a poor outcome. Together, these data demonstrate that Cbl-c can ubiquitinate and downregulate RETMEN2A and implicate Enigma as a positive regulator of RETMEN2A through blocking of Cbl-mediated ubiquitination and degradation.
Women's Malignancies Branch, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, Maryland, United States of America.
Stephen C
SC
Kales
Women's Malignancies Branch, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, Maryland, United States of America.
Marion M
MM
Nau
Center for Cancer Research Bioinformatics Core, Advanced Biomedical Computing Center, SAIC-Frederick, Frederick, Maryland, United States of America.
Anand S
AS
Merchant
Women's Malignancies Branch, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, Maryland, United States of America.
Stanley
S
Lipkowitz
D048868:Adaptor Proteins, Signal Transducing / Q000378:metabolism*; D015153:Blotting, Western; D001943:Breast Neoplasms / Q000378:metabolism* / Q000401:mortality / Q000473:pathology; D002478:Cells, Cultured; D003598:Cytoskeletal Proteins / Q000378:metabolism*; D005260:Female; D057809:HEK293 Cells; D006801:Humans; D047468:Immunoprecipitation; D060588:LIM Domain Proteins / Q000378:metabolism*; D046228:Microarray Analysis; D018813:Multiple Endocrine Neoplasia Type 2a / Q000378:metabolism*; D010190:Pancreatic Neoplasms / Q000378:metabolism* / Q000473:pathology; D059748:Proteolysis; D050721:Proto-Oncogene Proteins c-cbl / Q000378:metabolism*; D051096:Proto-Oncogene Proteins c-ret / Q000378:metabolism*; D020798:Two-Hybrid System Techniques; D054875:Ubiquitination
D048868:Adaptor Proteins, Signal Transducing; D003598:Cytoskeletal Proteins; D060588:LIM Domain Proteins; C484380:PDLIM7 protein, human; D050721:Proto-Oncogene Proteins c-cbl; D051096:Proto-Oncogene Proteins c-ret; C099282:RET protein, human