Aug 2026· Molecules and Cells· 0 citations· 83 references
Medicine
TL;DR
It is demonstrated here that the interaction between H-NS and StpA fine-tunes the physico-chemical properties of nucleoid-associated compartments, thereby preserving gene repression and optimizing bacterial growth under stress.
Abstract
Intragenomic homologs are widespread, but their physiological roles are often masked by redundancy. Histone-like nucleoid structuring protein (H-NS), a nucleoid-associated protein in Gram-negative bacteria, typically coexists with homologs like StpA, whose functions are obscured by a lack of strong phenotypes. We demonstrate here that the interaction between H-NS and StpA fine-tunes the physico-chemical properties of nucleoid-associated compartments. Although H-NS forms dynamic condensates in vitro, StpA assembles into stable insoluble fibrils. However, together the two proteins form liquid-like droplets, whose fluidity and stability are tunable by their relative stoichiometry. By increasing the levels of StpA over H-NS, bacteria stabilize heterochromatin-associated compartments, thereby preserving gene repression and optimizing bacterial growth under stress. Structural differences at these proteins' dimerization sites help explain their distinct phase behaviors. Our findings reveal a paradigm in which intragenomic homologs that are positioned at the opposite ends of the phase spectrum can fine-tune subcellular organization to promote survival in fluctuating environments.
Intracellular biomolecular condensation forms multicomponent signaling hubs that regulate development, stress responses, and environmental adaptation. While the molecular grammar encoded within scaffold proteins defines the basal associative features driving condensation, heterotypic condensates are intrinsically dynamic, multicomponent, and far-from-equilibrium systems. Consequently, how condensates organize component composition, stoichiometry, and functional specificity in space and time under physiological conditions remains poorly understood. Addressing this challenge requires integrative frameworks that combine predictive biophysical features with experimental information on protein abundance, interaction networks, subcellular localization, and evolutionary conservation. Here, we first analyzed phase separation (PS) proteins across the Tree of Life in 1,106 species, revealing a stark contrast in computationally predicted phase-separation propensity between eukaryotes and prokaryotes, with genome size as a key determinant. Through a broad analysis of amino acid homorepeat-containing proteins (HRPs) across all species, we uncovered how phase separation evolves via a balance between functional condensation and avoidance of harmful, aggregation-prone sequences. We further identified potential signaling hubs and components across kingdoms by integrating PS-positive proteins with experimentally derived abundance and interactome data from four model eukaryotic species. Using Arabidopsis as a model, we dissect the relationships among PS propensity, condensation hub prediction, HRPs, subcellular localization, and structural conservation. Consequently, we developed PhaseHub (https://phasehub.sbs.ntu.edu.sg/), a user-friendly interface for exploring scaffold-client dynamics, PS components, sequence signatures within each PS protein, and hubs. Our work provides an evolutionary framework for understanding multicomponent PS hubs by integrating molecular grammar with physiological context, thereby facilitating hypothesis generation and rational design.
An evolved hierarchy of paralog-specific JDP couplings that dynamically rewires the Hsp70 network from active repair to protection during stress is revealed, revealing an evolved hierarchy of paralog-specific JDP couplings that dynamically rewires the Hsp70 network from active repair to protection during stress.
Roni Suhler, Lars J. W. van Beurden, Merav D. Shmueli et al.· Proceedings of the National...· 0 citations
Among non‐classical nucleic acid secondary structures, G‐quadruplexes (G4s) play diverse roles in cellular functions and disease pathogenesis. However, the molecular mechanisms underlying the assembly of endogenous G4s into punctate condensates in cells remain unclear. Biomolecular condensates can arise from weak multivalent intermolecular interactions involving proteins and/or nucleic acids; this phenomenon is frequently linked to liquid–liquid phase separation. Recent research has provided compelling evidence for G4s driving biomolecular condensation. In this review, we first summarize the latest breakthroughs in the structural classification of G4s. In addition to frequently reported intramolecular G4s, intermolecular G4s have also been observed in cellular environments. Next, we discuss the regulatory role of G4s in condensation. Although G4s can independently form condensates, they primarily serve as structural platforms that facilitate condensate formation and regulate their phase transitions. Ultimately, this review reveals the multifaceted physiological and pathological functions of G4‐driven condensates, including chromatin organization, assembly of stress granules and paraspeckles, abnormal transcriptional activation, telomere maintenance, neurodegenerative disease‐associated protein aggregation, and viral inclusion body formation.
Wenmeng Wang, Qingqing Xu, Yuxin Zhang et al.· Advancement of science· 0 citations
The ParABS system orchestrates chromosome segregation in many bacterial species. The centromere-like parS sites serve as nucleation points for the initial binding of the ParB protein. Subsequent diffusion on adjacent, non-specific DNA regions (spreading) in the presence of CTP and binding of more ParB molecules along with DNA looping via ParB-ParB interactions bring distal parts of the chromosome into proximity. ParB interaction with the ParA-ATPase motor protein, then, drives genomic segregation. It has been shown that in some bacterial species, the ParB-parS complex undergoes phase separation into a condensate. However, the physico-chemical properties of such condensates and their response to forces, such as those they may face in the cell, have not yet been characterized. Performing turbidity measurements in the presence of CTP and various concentrations of DNA and physiologically relevant mono and divalent salt It was shown that Mg2+ facilitates, while K+ concentrations higher than ∼20 mM disfavors, condensate formation. Microrheology measurements showed that condensates of ParB and DNA including parS sites (ParB-parS DNA) in the presence of CTP, are viscoelastic with a viscosity at Troom of ∼5 Pa·s and able to quickly respond to deformations with a network relaxation time of 0.1 s. Additionally, fluorescence combined with force spectroscopy showed that mechanical disruption of ParB-DNA condensates in the presence of CTP requires ∼ 5-7.5 pN of tension in the DNA, which is lower than the force required to stall a molecular motor such as RNA polymerase, but higher than the force required for the relocation of chromosomes and plasmids during segregation. These results support the idea that ParB-parS condensates dynamically rearrange at the molecular level while maintaining the cohesion necessary to sustain the drag force of segregation without interfering with genomic transactions. This physical mechanism could be the basis for the critical role of ParB-parS condensates in organizing and partitioning bacterial chromosomes. Statement of Significance Condensates formed by liquid-liquid phase separation enable cellular compartmentalization without the energy-costly production of a membrane enclosure and have been implicated in a wide array of cellular functions. Here, we analyze their chemo-physical properties in relation to their activity in bacterial chromosome segregation using the C. crescentus ParB protein and DNA containing parS specific binding sites. We find that the cohesion of condensates of ParB parS-containing DNA confers the mechanical stability necessary to pull genetic material towards the cell pole without interfering with genomic functions. We propose a mechanism to explain the function of ParB-parS DNA in chromosomal segregation.
Ritika Gupta, Suleyman Ucuncuoglu, W. Childers et al.· bioRxiv· 0 citations
Histone proteins are critical for the compaction and organization of DNA. In eukaryotes, these proteins are typically encoded by a multigene family. Replication-dependent histones are highly conserved, both in their amino acid sequence and in their S-phase linked expression. Despite striking protein and regulatory similarities, histone genes often reside in wildly different organizations within genomes, even those of closely related species. At the same time, distantly-related species sometimes share organizational themes. How each species similarly regulates histone genes despite organizational differences remains unclear, and a multi-species approach is necessary to address this gap in knowledge. Here, we distill, categorize, and describe the histone gene organizations of over 250 species spanning 11 phyla. Our comprehensive curation of histone gene organizations provides new insights into the divergence and evolutionary changes of this ancient, conserved gene family. Additionally, we suggest that histone gene organization may contribute to subsequent regulation, as well as misregulation in disease.
Sisi Falcone, P. Raman, William F. Marzluff et al.· Molecular Biology of the Cel...· 0 citations
