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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Main Authors: Mariya S Liyasova, Ke Ma, Donna Voeller, Philip E Ryan, Jinqiu Chen, Rachel E Klevit, Stanley Lipkowitz
Format: Article
Language:English
Published: Public Library of Science (PLoS) 2019-01-01
Series:PLoS ONE
Online Access:https://doi.org/10.1371/journal.pone.0216967
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author Mariya S Liyasova
Ke Ma
Donna Voeller
Philip E Ryan
Jinqiu Chen
Rachel E Klevit
Stanley Lipkowitz
author_facet Mariya S Liyasova
Ke Ma
Donna Voeller
Philip E Ryan
Jinqiu Chen
Rachel E Klevit
Stanley Lipkowitz
author_sort Mariya S Liyasova
collection DOAJ
description 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.
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spelling doaj.art-96dd9dd4f7ff4a2bbd1769476308f6722022-12-21T18:34:20ZengPublic Library of Science (PLoS)PLoS ONE1932-62032019-01-01145e021696710.1371/journal.pone.0216967Cbl interacts with multiple E2s in vitro and in cells.Mariya S LiyasovaKe MaDonna VoellerPhilip E RyanJinqiu ChenRachel E KlevitStanley LipkowitzMany 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.https://doi.org/10.1371/journal.pone.0216967
spellingShingle Mariya S Liyasova
Ke Ma
Donna Voeller
Philip E Ryan
Jinqiu Chen
Rachel E Klevit
Stanley Lipkowitz
Cbl interacts with multiple E2s in vitro and in cells.
PLoS ONE
title Cbl interacts with multiple E2s in vitro and in cells.
title_full Cbl interacts with multiple E2s in vitro and in cells.
title_fullStr Cbl interacts with multiple E2s in vitro and in cells.
title_full_unstemmed Cbl interacts with multiple E2s in vitro and in cells.
title_short Cbl interacts with multiple E2s in vitro and in cells.
title_sort cbl interacts with multiple e2s in vitro and in cells
url https://doi.org/10.1371/journal.pone.0216967
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