Biochemical basis for the requirement of kinase activity for Cbl-dependent ubiquitinylation and degradation of a target tyrosine kinase

Amiya K. Ghosh, Alagarsamy L. Reddi, Navin L. Rao, Lei Duan, Vimla Band, Hamid Band

Research output: Contribution to journalArticle

14 Citations (Scopus)

Abstract

Members of the Cbl family of ubiquitin ligases have emerged as crucial negative regulators of tyrosine kinase signaling. These proteins preferentially interact with and target activated tyrosine kinases for ubiquitinylation, thereby facilitating the lysosomal sorting of receptor tyrosine kinases or proteasomal degradation of nonreceptor tyrosine kinases. Recent work has indicated a crucial role of the target kinase activity in Cbl-dependent ubiquitinylation and degradation, but the biochemical basis for this requirement is not understood. Here, we have used the Src-family kinase Fyn, a well characterized Cbl target, to address this issue. Using defined Fyn mutants, we demonstrate that the kinase activity of Fyn is crucial for its Cbl-dependent ubiquitinylation and degradation, but a low level of ubiquitinylation and degradation of kinase-inactive Fyn mutants was consistently observed. Mutational induction of an open conformation enhanced the susceptibility of kinase-active Fyn to Cbl but was insufficient to promote the ubiquitinylation and degradation of kinase-inactive Fyn. Notably, the Cbl-dependent degradation of Fyn did not require the Fyn-mediated phosphorylation of Cbl. Finally, we show that the major determinant of the susceptibility of Fyn protein to Cbl-dependent ubiquitinylation and degradation is the extent to which it physically associates with Cbl; kinase activity of Fyn serves as a critical determinant to promote its association with Cbl, which we demonstrate is mediated by multiple protein-protein interactions. Our results strongly suggest that promotion of association with Cbl is the primary mechanism by which the kinase activity of the targets of Cbl contributes to their susceptibility to Cbl.

Original languageEnglish (US)
Pages (from-to)36132-36141
Number of pages10
JournalJournal of Biological Chemistry
Volume279
Issue number34
DOIs
StatePublished - Aug 20 2004

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Protein-Tyrosine Kinases
Phosphotransferases
Degradation
Proteins
Association reactions
Phosphorylation
src-Family Kinases
Receptor Protein-Tyrosine Kinases
Ligases
Ubiquitin
Sorting
Conformations

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

Cite this

Biochemical basis for the requirement of kinase activity for Cbl-dependent ubiquitinylation and degradation of a target tyrosine kinase. / Ghosh, Amiya K.; Reddi, Alagarsamy L.; Rao, Navin L.; Duan, Lei; Band, Vimla; Band, Hamid.

In: Journal of Biological Chemistry, Vol. 279, No. 34, 20.08.2004, p. 36132-36141.

Research output: Contribution to journalArticle

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