Multi-layered proteomic analyses decode compositional and functional effects of cancer mutations on kinase complexes

Abstract

Rapidly increasing availability of genomic data and ensuing identification of disease associated mutations allows for an unbiased insight into genetic drivers of disease development. However, determination of molecular mechanisms by which individual genomic changes affect biochemical processes remains a major challenge. Here, we develop a multilayered proteomic workflow to explore how genetic lesions modulate the proteome and are translated into molecular phenotypes. Using this workflow we determine how expression of a panel of disease-associated mutations in the Dyrk2 protein kinase alter the composition, topology and activity of this kinase complex as well as the phosphoproteomic state of the cell. The data show that altered protein-protein interactions caused by the mutations are associated with topological changes and affected phosphorylation of known cancer driver proteins, thus linking Dyrk2 mutations with cancer-related biochemical processes. Overall, we discover multiple mutation-specific functionally relevant changes, thus highlighting the extensive plasticity of molecular responses to genetic lesions.

The DYRK Family of Kinases in Cancer: Molecular Functions and Therapeutic Opportunities

“…The activity of DYRK2 has often been linked to its ability to negatively regulate the stability of its target proteins—in particular, through its interaction with the UBR5/EDD-DNA damage-binding protein 1 (DDB1)-DDB1- and cullin 4-associated factor homolog 1 (DCAF1/VPRBP) (EDVP) E3 ubiquitin ligase complex [ 38 ] ( Figure 5 A). The relevance of this interaction is highlighted by the alterations in the assembly of the EDVP complex detected in the analysis of certain DYRK2 mutants found in cancer samples [ 153 ]. In addition, it is worth noting that many of the proteins that are degraded following DYRK2 phosphorylation are targets of the tumor suppressor F-box/WD repeat-containing protein 7 (FBXW7) ( Figure 5 A), suggesting possible cross-talk with E3 ubiquitin ligase protein complexes made up of this factor.…”

Data-independent acquisition mass spectrometry (DIA-MS) for proteomic applications in oncology

“…For instance, Mehnert et al developed a multi-layered proteomic approach to study effects of different mutations of Dual specificity tyrosine-phosphorylationregulated kinase 2 (Dyrk2) on protein topology, protein-protein interactions (PPI) and global proteomic and phosphoproteomic profiles. 70 Through interactions with the EDVP E3 ubiquitin ligase complex, Dyrk2 plays a key role in cell cycle and apoptosis and has been identified as both a putative tumour suppressor and oncogene. 71,72 Based on published data, the authors generated a series of cancer-associated Dyrk2 mutants which were expressed in HEK293 cells.…”

Mass spectrometry‐based protein–protein interaction networks for the study of human diseases

“…Mehnert et al combined AP-MS, BioID, and XL-MS to establish the molecular response to five cancer-associated DYRK2 mutations. These results were compared with WT DYRK2 and a catalytically inactive, non-cancerous DYRK2 mutation to determine the effect of each mutation on cancer-relevant interactions and biochemical pathways (Mehnert et al, 2020). The different mutations resulted in varied perturbations of the interactome, with a C-terminal truncated mutant and the catalytically inactive mutant causing the strongest interactome changes, including a loss of interactions with the DYRK2 kinase core complex.…”