Multiplexed measurements of protein-protein interactions and protein abundance across cellular conditions using Prod&PQ-seq

ABSTRACT Methods to profile protein-protein interactions (PPIs) have limited scalability and can only study a handful of conditions and/or targets. Here, we introduce Prod&PQ-seq, a framework for multiplexed detection and quantification of PPIs and proteins. Our framework uses cross-linked cells, antibody-oligonucleotide conjugates (ab-oligos), and captures PPIs by the DNA-caliper, a specialized oligonucleotide for bidirectional priming of proximal ab-oligos. We benchmarked Prod&PQ-seq using recombinant complexes, titrations and cell mixture experiments and show that our framework is quantitative, reproducible, sensitive and specific. Applying Prod&PQ-seq to study Polycomb Repressive Complex 2 (PRC2) shows that EZH2 inhibition and expression of the oncohistone H3.3K27M weakens both PRC2-H3K27me3 interactions and PPIs within PRC2. Further, H3.1K27M and H3.3K27M variants lead to distinct PPI profiles such as the intensity of H3K27ac-K27M or H3K27ac-EED. Together, Prod&PQ-seq enables detection of changes in PPI composition and intensity and protein quantification across biological conditions, small molecule inhibition and genetic perturbations. Competing Interest Statement T.X., S.H, C.B and A.G. are inventors on a related patent application; J.F., M.N, and C.J.F. are employees of Cell Signaling Technology (CST). Footnotes Fixed an error in middle name of an author and added a missing funding info