Jul 2026· Current Opinion in Structural Biology· Vol 100, pp.
103334
· 0 citations· 57 references
Medicine
TL;DR
This work examines the physicochemical principles underlying RNA-mediated crowding, focusing on how sequence-driven multivalency, structural topology, and network connectivity govern molecular organization.
Abstract
Molecular crowding in cells is not solely a consequence of excluded volume but emerges from interaction networks shaped by RNA and proteins. Here we examine the physicochemical principles underlying RNA-mediated crowding, focusing on how sequence-driven multivalency, structural topology, and network connectivity govern molecular organization. Repeats, secondary structures, and higher-order motifs such as G-quadruplexes act as interaction modules that promote percolation and phase separation, while RNA length and concentration tune phase boundaries. These RNA-encoded features promote multivalent RNA-RNA and RNA-protein interactions that shape condensate assembly, dynamics, and organization across scales. In this framework, crowding emerges as an RNA-centered, interaction-driven property linking molecular features to mesoscale organization in both physiological and pathological contexts.
This minireview examines how RNA shapes the formation, composition, and material properties of biomolecular condensates, highlighting the molecular grammar encoded in RNA sequence, length and valency, structure, and chemical modifications.
Shibam Dey, O. Amster-Choder· Current Opinion in Structura...· 0 citations
Crowding-aware structural biology, including in-cell nuclear magnetic resonance, cryo-electron microscopy of condensates, single-molecule methods, and thermodynamic-kinetic modelling, will be important for resolving physiologically relevant intermediates.
Tejas Nikam, Shashi Prakash Patel, S. Saraf et al.· Current Opinion in Structura...· 1 citation
This review discusses how features of protein sequence, such as amino acid composition, interaction motifs, and post-translational modifications, influence condensate aging and highlights how interactions with RNA and lipid membranes modulate condensate behavior by altering interaction networks and interfacial properties.
Rebecca J. Thrush, Francesco A. Aprile· Current Opinion in Structura...· 0 citations
Reversible associations of RNAs among themselves, or with RNA binding proteins through the process of phase separation, are found to be important in many different cellular contexts, including cellular responses to stress, gene regulation, development, and disease. Short RNA repeat sequences, which are mostly linked with many repeat expansion neurodegenerative diseases, are found to undergo phase separation and yield protein-free biomolecular condensates in vitro and in cells. However, the physicochemical principles governing phase separation of RNAs considering both sequence and structural aspects, especially for short RNAs, remain elusive. It is intriguing, as well as challenging, to characterize the RNA phase behavior at a submolecular resolution. Based on atomistic enhanced sampling simulations, here we report potential dynamic structural effects in the mutual association properties of a tetra loop containing 14-mer hairpin RNA. We show that the folded and unfolded conformations of the hairpin fragment lead to different energetic barriers for the formation of an associated pair. Unfolded conformation leads to the formation of gel like energetically stable associated phases spontaneously; the folded hairpin motif is found to yield droplet like associated phases accompanying by cations in solution. Overall, our findings illustrate that dynamic association/dissociation is energetically more favorable for folded RNA hairpins, and the presence of additional salt in solution assists the formation of dense droplet like associated phases, while ionic concentration plays a critical role in the formation and stability of the droplet like associated phase. These observations open avenues for exploring structure-based phase separation mechanisms of RNAs in the context of RNA mediated functional biomolecular condensate formation within cells and designing RNA based novel biomaterials.
M. Mondal, Maodong Li, Y. Gao· Journal of Chemical Physics· 0 citations
RNA molecules populate complex structural landscapes that are continuously reshaped throughout the RNA lifecycle by equilibrium and non-equilibrium processes. Resolving these structural landscapes represents a central challenge in RNA biochemistry. We review recent advances in RNA chemical probing, sequencing, and computational deconvolution technologies that are revolutionizing our ability to measure the complexities of RNA folding in cells and the deep involvement of these complexities in RNA functional mechanisms. We highlight new methods for deconvolving structural ensembles, distinguishing isoform-specific architectures using long-read sequencing, capturing co-transcriptional folding intermediates in vivo, and measuring higher-order RNA structures while also underscoring remaining challenges. We conclude by outlining future directions in probe development, sequencing, and integrative modeling, and discuss how resolving RNA structural ensembles with increasingly high resolution will likely reveal new therapeutic opportunities to selectively target functional RNA heterogeneity.
Anthony M. Mustoe, D. Incarnato· Current Opinion in Structura...· 0 citations
It is demonstrated that single-molecule mass measurements with mass photometry can capture RNA-protein interactions in phase-separated protein systems and can distinguish between charge neutralization, which drives coacervation, and complex formation, which mediates phase re-entry, making it a highly complementary tool for the study of RNA-mediated phase separation.
Axel Leppert, Jesper Shiapan, Irena Papageorgiou et al.· bioRxiv· 0 citations
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