Protein–protein interactions (PPIs) represent the main mode of the proteome organization in the cell. In the last decade, several large-scale representations of PPI networks have captured generic aspects of the functional organization of network components but mostly lack the context of cellular states. However, the generation of context-dependent PPI networks is essential for structural and systems-level modeling of biological processes—a goal that remains an unsolved challenge. Here we describe an experimental/computational strategy to achieve a modeling of PPIs that considers contextual information. This strategy defines the composition, stoichiometry, temporal organization, and cellular requirements for the formation of target assemblies. We used this approach to generate an integrated model of the formation principles and architecture of a large signalosome, the TNF–receptor signaling complex (TNF-RSC). Overall, we show that the integration of systems- and structure-level information provides a generic, largely unexplored link between the modular proteome and cellular function.

Unidirectional transport of mRNA from the nucleus to the cytoplasm via nuclear pore complexes is an essential step in the gene expression of all eukaryotes. Although factors involved in mRNA transport have been characterized, a comprehensive mechanistic understanding of this critical process and its regulation is lacking. Here, we use real-time single RNA imaging to demonstrate that acute depletion of the budding yeast DEAD-box ATPase Dbp5 causes rapid nuclear accumulation of mRNAs in vivo and dramatic changes in nuclear dynamics of RNA export factors. In particular, the essential export factor Nab2 ceases to shuttle between the nucleus and cytoplasm and forms an RNA-dependent condensate throughout the nucleus. Phase-separation can be recapitulated in vitro, with Nab2 forming RNA-dependent liquid droplets, which depend on the presence of Dbp5. Intriguingly, in glucose stress, condensation of Nab2 blocks bulk mRNA export while selectively allowing the passage of stress-induced mRNAs from the nucleus to the cytoplasm to elicit a timely cellular stress response. This is accompanied by a lowered abundance of the DEAD-box ATPase Dbp5 at the cytoplasmic sites of nuclear pore complexes, which leads to the formation of the Nab2 condensates. Our results suggest that cells use selective mRNA retention in nuclear Nab2 condensates to re-wire mRNA export and to regulate gene expression during stress.

Protein-protein interactions (PPI) represent the main mode of the proteome organization in the cell. In the last decade, several large-scale representations of PPI networks have captured generic aspects of the functional organization of network components, but mostly lack the context of cellular states. However, the generation of contextual representations of PPI networks is essential for structural and systems-level modeling of biological processes and remains an unsolved challenge. In this study we describe an integrated experimental/computational strategy to achieve a contextualized modeling of PPI. This strategy defines the composition, stoichiometry, spatio-temporal organization and cellular requirements for the formation of target assemblies. We used this approach to generate an integrated model of the formation principles and architecture of a large signalosome, the TNF-receptor signaling complex (TNF-RSC). Overall, we show that the integration of systems- and structure-level information provides a generic, largely unexplored link between the modular proteome and cellular function.

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