RNA length has a non-trivial effect in the stability of biomolecular condensates formed by RNA-binding proteins

Biomolecular condensates formed via liquid-liquid phase separation (LLPS) play a crucial role in the spatiotemporal organization of the cell material. Nucleic acids can act as critical modulators in the stability of these protein condensates. Here, we present a multiscale computational strategy, exploiting the advantages of both a sequence-dependent coarse-grained representation of proteins and a minimal coarse-grained model that represent proteins as patchy colloids, to unveil the role of RNA length in regulating the stability of RNA-binding protein (RBP) condensates. We find that for a constant RNA/protein ratio in which phase separation is enhanced, the protein fused in sarcoma (FUS), which can phase separate on its own-i.e., via homotypic interactions-only exhibits a mild dependency on the RNA strand length, whereas, the 25-repeat proline-arginine peptide (PR25), which does not undergo LLPS on its own at physiological conditions but instead exhibits complex coavervation with RNA-i.e., via heterotypic interactions-shows a strong dependence on the length of the added RNA chains. Our minimal patchy particle simulations, where we recapitulate the modulation of homotypic protein LLPS and complex coacervation by RNA length, suggest that the strikingly different effect of RNA length on homotypic LLPS versus complex coacervation is general. Phase separation is RNA-length dependent as long as the relative contribution of heterotypic interactions sustaining LLPS is comparable or higher than that committed by protein homotypic interactions. Taken together, our results contribute to illuminate the intricate physicochemical mechanisms that influence the stability of RBP condensates through RNA inclusion.

Bacterial Death and TRADD-N domains help define novel apoptosis and immunity mechanisms shared by prokaryotes and metazoans

Several homologous domains are shared by eukaryotic immunity and programmed cell-death systems and poorly understood bacterial proteins. Recent studies show these to be components of a network of highly regulated systems connecting apoptotic processes to counter-invader immunity, in prokaryotes with a multicellular habit. However, the provenance of key adaptor domains, namely those of the Death-like and TRADD-N superfamilies, a quintessential feature of metazoan apoptotic systems, remained murky. Here, we use sensitive sequence analysis and comparative genomics methods to identify unambiguous bacterial homologs of the Death-like and TRADD-N superfamilies. We show the former to have arisen as part of a radiation of effector-associated α-helical adaptor domains that likely mediate homotypic interactions bringing together diverse effector and signaling domains in predicted bacterial apoptosis- and counter-invader systems. Similarly, we show that the TRADD-N domain defines a key, widespread signaling bridge that links effector deployment to invader-sensing in multicellular bacterial and metazoan counter-invader systems. TRADD-N domains are expanded in aggregating marine invertebrates and point to distinctive diversifying immune strategies probably directed both at RNA and retro- viruses and cellular pathogens that might infect such communities. These TRADD-N and Death-like domains helped identify several new bacterial and metazoan counter-invader systems featuring under-appreciated, common functional principles: the use of intracellular invader-sensing lectin-like (NPCBM and FGS), transcription elongation GreA/B-C, glycosyltransferase-4 family, inactive NTPase (serving as nucleic-acid-receptors) and invader-sensing GTPase switch domains. Finally, these findings point to the possibility of multicellular bacteria-stem metazoan symbiosis in the emergence of the immune/apoptotic systems of the latter.

Coarse Grained Molecular Dynamic Simulations for the Study of TNF Receptor Family Members' Transmembrane Organization

The Tumor Necrosis Factor (TNF) and the TNF receptor (TNFR) superfamilies are composed of 19 ligands and 30 receptors, respectively. The oligomeric properties of ligands, both membrane bound and soluble, has been studied most. However, less is known about the oligomeric properties of TNFRs. Earlier reports identified the extracellular, membrane-distal, cysteine-rich domain as a pre-ligand assembly domain which stabilizes receptor dimers and/or trimers in the absence of ligand. Nevertheless, recent reports based on structural nuclear magnetic resonance (NMR) highlight the intrinsic role of the transmembrane domains to form dimers (p75NTR), trimers (Fas), or dimers of trimers (DR5). Thus, understanding the structural basis of transmembrane oligomerization may shed light on the mechanism for signal transduction and the impact of disease-associated mutations in this region. To this end, here we used an in silico coarse grained molecular dynamics approach with Martini force field to study TNFR transmembrane homotypic interactions. We have first validated this approach studying the three TNFR described by NMR (p75NTR, Fas, and DR5). We have simulated membrane patches composed of 36 helices of the same receptor equidistantly distributed in order to get unbiassed information on spontaneous proteins assemblies. Good agreement was found in the specific residues involved in homotypic interactions and we were able to observe dimers, trimers, and higher-order oligomers corresponding to those reported in NMR experiments. We have, applied this approach to study the assembly of disease-related mutations being able to assess their impact on oligomerization stability. In conclusion, our results showed the usefulness of coarse grained simulations with Martini force field to study in an unbiased manner higher order transmembrane oligomerization.

Dichotomy between heterotypic and homotypic interactions by a common chemical law

This study introduces a general correction to the classical chemical rate law to avoid overestimating the frequency of homotypic interactions.