Functional annotation of genes is a challenge in the post-genomic era. Even in the best-studied model Arabidopsis, a significant proportion of genes and their encoded proteins remain functionally uncharacterized. Determining the protein-protein interaction (PPI) network is the basis for annotating protein function, considering that proteins interact for structural, regulatory, and catalytic purposes, and that interacting proteins are typically involved in the same biological processes (guilt-by-association). Cross-linking mass spectrometry is a powerful tool for the determination of PPIs by providing direct evidence of interactions between the interacting complex, and precise localization of the contact sites at the amino acid level. Here, we applied cross-linking mass spectrometry (XL-MS) to study the Arabidopsis protein network. Our analysis revealed about 27,000 intra-crosslinks and 3000 inter-crosslinks, providing insights into protein structure, protein complex topologies, and novel interactions. Among the inter-crosslinks, a significant proportion (>70%) is supported by STRING analysis, while 30% represent novel interactions. These interactions have led to new biological hypotheses and improved our understanding of cellular processes in plants.
This work was funded by the NIH grant R01GM135706 and S10OD030441 to S-LX and by Carnegie endowment fund to Carnegie mass spectrometry facility.