Comprehensive Profiling of Mammalian Tribbles Interactomes Implicates TRIB3 in Gene Repression

SIMPLE SUMMARY: Tribbles proteins play various roles in cancer initiation and progression. However, still little is known about their molecular actions. Here we developed a mass spectrometry-based approach to study the Tribbles interactomes, allowing us to discover new interactors and functions that might help to understand their behavior better. Our proteomics data highlight the ability of TRIB3 to interact with transcription regulatory proteins and point to a new role in gene repression. Systematic analyses like these will help to evaluate the potential of Tribbles proteins as biomarkers for disease diagnosis and prognosis. ABSTRACT: The three human Tribbles (TRIB) pseudokinases have been implicated in a plethora of signaling and metabolic processes linked to cancer initiation and progression and can potentially be used as biomarkers of disease and prognosis. While their modes of action reported so far center around protein–protein interactions, the comprehensive profiling of TRIB interactomes has not been reported yet. Here, we have developed a robust mass spectrometry (MS)-based proteomics approach to characterize Tribbles’ interactomes and report a comprehensive assessment and comparison of the TRIB1, -2 and -3 interactomes, as well as domain-specific interactions for TRIB3. Interestingly, TRIB3, which is predominantly localized in the nucleus, interacts with multiple transcriptional regulators, including proteins involved in gene repression. Indeed, we found that TRIB3 repressed gene transcription when tethered to DNA in breast cancer cells. Taken together, our comprehensive proteomic assessment reveals previously unknown interacting partners and functions of Tribbles proteins that expand our understanding of this family of proteins. In addition, our findings show that MS-based proteomics provides a powerful tool to unravel novel pseudokinase biology.