Cbl interacts with multiple E2s in vitro and in cells

Many receptor tyrosine kinases (RTKs, such as EGFR, MET) are negatively regulated by ubiquitination and degradation mediated by Cbl proteins, a family of RING finger (RF) ubiquitin ligases (E3s). Loss of Cbl protein function is associated with malignant transformation driven by increased RTK activit...

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Bibliographic Details
Published in:PloS one 2019-05, Vol.14 (5), p.e0216967-e0216967
Main Authors: Liyasova, Mariya S, Ma, Ke, Voeller, Donna, Ryan, Philip E, Chen, Jinqiu, Klevit, Rachel E, Lipkowitz, Stanley
Format: Article
Language:English
Subjects:
Biochemistry
Biodegradation
Biology and Life Sciences
Cancer
Cbl protein
Chains
EDTA
Enzymes
Epidermal growth factor
Epidermal growth factor receptors
ErbB Receptors - metabolism
Gene Expression Regulation
Gene mutation
Gene Silencing
Genetic transformation
HEK293 Cells
HeLa Cells
Humans
Immunoglobulins
Kinases
Ligands
Ligases
Medical research
Medical screening
Mutation
Phenols (Class of compounds)
Phosphorylation
Protein Binding
Proteins
Proto-Oncogene Proteins c-cbl - genetics
Proto-Oncogene Proteins c-cbl - metabolism
Research and Analysis Methods
Screens
Substrates
Syk protein
Transformation
Two-Hybrid System Techniques
Tyrosine
Ubiquitin
Ubiquitin - metabolism
Ubiquitin-conjugating enzyme
Ubiquitin-Conjugating Enzymes - metabolism
Ubiquitin-protein ligase
Ubiquitination
Yeast
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Summary:Many receptor tyrosine kinases (RTKs, such as EGFR, MET) are negatively regulated by ubiquitination and degradation mediated by Cbl proteins, a family of RING finger (RF) ubiquitin ligases (E3s). Loss of Cbl protein function is associated with malignant transformation driven by increased RTK activity. RF E3s, such as the Cbl proteins, interact with a ubiquitin-conjugating enzyme (E2) to confer specificity to the ubiquitination process and direct the transfer of ubiquitin from the E2 to one or more lysines on the target proteins. Using in vitro E3 assays and yeast two-hybrid screens, we found that Ube2d, Ube2e families, Ube2n/2v1, and Ube2w catalyze autoubiquitination of the Cbl protein and Ube2d2, Ube2e1, and Ube 2n/2v1 catalyze Cbl-mediated substrate ubiquitination of the EGFR and SYK. Phosphorylation of the Cbl protein by by Src resulted in increased E3 activity compared to unphosphorylated cbl or Cbl containing a phosphomimetic Y371E mutation. Ubiquitin chain formation depended on the E2 tested with Cbl with Ube2d2 forming both K48 and K63 linked chains, Ube2n/2v1 forming only K63 linked chains, and Ube2w inducing monoubiquitination. In cells, the Ube2d family, Ube2e family, and Ube2n/2v1 contributed to EGFR ubiquitination. Our data suggest that multiple E2s can interact with Cbl and modulate its E3 activity in vitro and in cells.
ISSN:1932-6203
1932-6203
DOI:10.1371/journal.pone.0216967