Stable and EGF-Induced Temporal Interactome Profiling of CBL and CBLB Highlights Their Signaling Complex Diversity

The
epidermal growth factor receptor (EGFR) signal modulates cell
proliferation, migration, and survival. Aberrant activation of EGFR
constitutes the major cause of various cancers. Receptor ubiquitination
and degradation mediated by CBL proteins play negative regulatory
roles and control the intensity and duration of the signaling. With
the construction of stable cell lines inducibly expressing FLAG-tagged
CBL or CBLB, we identified 102 and 82 stable interacting proteins
of CBL and CBLB, respectively, through the affinity purification followed
by mass spectrometry (AP-MS) approach. Time-resolved profiling at
six different time points combined with functional annotations of
the temporal interactomes provides insights into the dynamic assembly
of signal proteins upon EGFR signaling activation. Comparison between
the interactomes of CBL and CBLB indicates their redundant but also
complementary functions. Importantly, we validated the stable association
of EPS15L1 and ITSN2 and temporal association of TNK2 to both CBL
and CBLB through biochemical assays. Collectively, these results offer
a useful resource for CBL and CBLB interactomes and highlight their
prominent and diverse roles in the EGFR signaling network.