PROGRAM ABSTRACT: Integrated mass spectrometry-based chemoproteomic and genomic technolo-
gies for studying dynamic kinase interactomes
Dynamic changes in protein-protein and protein-DNA interactions (PPIs and PDIs) control most cellular pro-
cesses, including cell signaling and transcription; devastating diseases rewire PPI and PDI networks to drive
disease progression, therapy resistance, and immune escape. Novel methods for mapping dynamic interaction
networks are, therefore, urgently required to identify disease mechanisms and drug targets. Protein kinases are
critical regulatory nodes in most cellular PPI and PDI networks, are often dysregulated in disease, and are highly
druggable with synthetic, ATP-competitive inhibitors. Accordingly. insights into how diseases utilize kinases to
rewire PPI and PDI networks are extremely relevant for combating many diseases. Advances in quantitative
mass spectrometry (MS) have revolutionized proteomics, yet, facile methods for the systematic, sensitive, and
high-throughput profiling of kinase PPIs, locus-specific PDIs, and their dynamics are lacking. We will develop
transformative methods that combine cell-permeable affinity probes with chemical crosslinking and proximity
labeling to globally encode kinase interactomes in situ, followed by integrated LC-MS and sequencing analyses.
Cellular plasticity drives physiological and pathological de- and transdifferentiation, and lineage switching,
critically contributing to development, tissue repair, cancer metastasis, organ fibrosis, and therapy and immune
escape in numerous diseases. To identify drug targets for combating these disease phenotypes, we pressingly
need to understand the signaling and transcriptional network that underly cellular plasticity. Our studies of patho-
logical kinome rewiring linked ~20% of human kinases to cellular plasticity, among them numerous understudied
kinases. We found that 70% these kinases localize to the nucleus and interact with transcription factors and
chromatin remodelers. We also found that cellular plasticity dynamically alters the post-translational modifica-
tions (PTMs) and PPIs of these kinases. How plasticity pathways coordinate dynamic changes in PTM, PPI and
PDI networks to systematically alter chromatin states and transcription, however, remains largely unknown, leav-
ing critical molecular mechanisms and drug targets unexplored. We will develop streamlined workflows for stud-
ying nuclear kinase dynamics, combining kinobead/LC-MS kinome profiling with global proteomics, epigenomics,
and transcriptomics analyses, and our novel interactomic platforms, and apply these workflows to unravel how
plasticity pathways spatiotemporally control kinases during cellular de- and transdifferentiation.
To summarize, our program seeks to develop novel bioanalytical methods and workflows to systematically
study dynamic kinase interactomes, and to illuminate the mechanisms of pathological cellular plasticity. Pursuing
our goals, we created an ambitious, rigorous, and productive research program that fosters inclusiveness and
creativity, training the next generation of scientific leaders in proteomics, cell signaling, and chemical biology.
