Transcription templated assembly of the nucleolus in the embryo.

The nucleolus is a multicomponent structure made of RNA and proteins that serves as the site of ribosome biogenesis within the nucleus. It has been extensively studied as a prototype of a biomolecular condensate whose assembly is driven by phase separation. While the steady-state size of the nucleolus is quantitatively accounted for by the thermodynamics of phase separation, we show that experimental measurements of the assembly dynamics are inconsistent with a simple model of a phase-separating system relaxing to its equilibrium state.

Phase Separation in Biology and Disease; Current Perspectives and Open Questions.

In the past almost 15 years, we witnessed the birth of a new scientific field focused on the existence, formation, biological functions, and disease associations of membraneless bodies in cells, now referred to as biomolecular condensates. Pioneering studies from several laboratories [reviewed in] supported a model wherein biomolecular condensates associated with diverse biological processes form through the process of phase separation. These and other findings that followed have revolutionized our understanding of how biomolecules are organized in space and time within cells to perform myriad biological functions, including cell fate determination, signal transduction, endocytosis, regulation of gene expression and protein translation, and regulation of RNA metabolism.

Let's phase it: viruses are master architects of biomolecular condensates.

Viruses compartmentalize their replication and assembly machinery to both evade detection and concentrate the viral proteins and nucleic acids necessary for genome replication and virion production. Accumulating evidence suggests that diverse RNA and DNA viruses form replication organelles and nucleocapsid assembly sites using phase separation. In general, the biogenesis of these compartments is regulated by two types of viral protein, collectively known as antiterminators and nucleocapsid proteins, respectively.

Single-Molecule Tracking of RNA Polymerase In and Out of Condensates in Live Bacterial Cells.

Biomolecular condensates, first discovered in eukaryotic cells, were recently found in bacteria. The small size of these organisms presents unique challenges for identifying and characterizing condensates. Here, we describe a single-molecule approach for studying biomolecular condensates in live bacterial cells.

Dissecting the complexity of biomolecular condensates.

Biomolecular condensates comprise a diverse and ubiquitous class of membraneless organelles. Condensate assembly is often described by liquid-liquid phase separation. While this process explains many key features, it cannot account for the compositional or architectural complexity that condensates display in cells.

Evaluation of Lanthanide-Doped Upconverting Nanoparticles for and Applications.

Because of their unique physicochemical properties, lanthanide-doped upconverting nanoparticles (Ln-UCNPs) have exceptional potential for biological applications. However, the use in biological systems is hampered by the limited understanding of their bionano interactions. Our multidisciplinary study has generated these insights through in-depth and quantitative analyses.

Beyond Equilibrium Phase Diagrams: Enzymatic Activity Shakes Up Bacterial Condensates.

Phase separation is a thermodynamic process, but cells are inherently out of equilibrium. Guilhas et al. (2020) identify an active process through which an ATP-dependent motor controls the number and position of biomolecular condensates in bacteria.

Clusters of bacterial RNA polymerase are biomolecular condensates that assemble through liquid-liquid phase separation.

Once described as mere "bags of enzymes," bacterial cells are in fact highly organized, with many macromolecules exhibiting nonuniform localization patterns. Yet the physical and biochemical mechanisms that govern this spatial heterogeneity remain largely unknown. Here, we identify liquid-liquid phase separation (LLPS) as a mechanism for organizing clusters of RNA polymerase (RNAP) in Using fluorescence imaging, we show that RNAP quickly transitions from a dispersed to clustered localization pattern as cells enter log phase in nutrient-rich media.

Nucleolar Organization and Functions in Health and Disease.

The nucleolus is a prominent, membraneless compartment found within the nucleus of eukaryotic cells. It forms around ribosomal RNA (rRNA) genes, where it coordinates the transcription, processing, and packaging of rRNA to produce ribosomal subunits. Recent efforts to characterize the biophysical properties of the nucleolus have transformed our understanding of the assembly and organization of this dynamic compartment.

Evidence for and against Liquid-Liquid Phase Separation in the Nucleus.

Enclosed by two membranes, the nucleus itself is comprised of various membraneless compartments, including nuclear bodies and chromatin domains. These compartments play an important though still poorly understood role in gene regulation. Significant progress has been made in characterizing the dynamic behavior of nuclear compartments and liquid-liquid phase separation (LLPS) has emerged as a prominent mechanism governing their assembly.

Sequence-encoded material properties dictate the structure and function of nuclear bodies.

Concomitant with packaging the genome, the cell nucleus must also spatially organize the nucleoplasm. This complex mixture of proteins and nucleic acids partitions into a variety of phase-separated, membraneless organelles called nuclear bodies. Significant progress has been made in understanding the relationship between the material properties of nuclear bodies and their structural and functional consequences.

Hierarchical Size Scaling during Multicellular Growth and Development.

Multicellular organisms must regulate their growth across the diverse length scales of biological organization, but how this growth is controlled from organelle to body, while coordinating interdependent functions at each scale, remains poorly understood. We utilized the C. elegans worm intestine as a model system to identify distinct allometric scaling laws, revealing that the growth of individual structures is differentially regulated during development.

RNA transcription modulates phase transition-driven nuclear body assembly.

Nuclear bodies are RNA and protein-rich, membraneless organelles that play important roles in gene regulation. The largest and most well-known nuclear body is the nucleolus, an organelle whose primary function in ribosome biogenesis makes it key for cell growth and size homeostasis. The nucleolus and other nuclear bodies behave like liquid-phase droplets and appear to condense from the nucleoplasm by concentration-dependent phase separation.

Inverse size scaling of the nucleolus by a concentration-dependent phase transition.

Just as organ size typically increases with body size, the size of intracellular structures changes as cells grow and divide. Indeed, many organelles, such as the nucleus [1, 2], mitochondria [3], mitotic spindle [4, 5], and centrosome [6], exhibit size scaling, a phenomenon in which organelle size depends linearly on cell size. However, the mechanisms of organelle size scaling remain unclear.

Analytical tools to distinguish the effects of localization error, confinement, and medium elasticity on the velocity autocorrelation function.

Single particle tracking is a powerful technique for investigating the dynamic behavior of biological molecules. However, many of the analytical tools are prone to generate results that can lead to mistaken interpretations of the underlying transport process. Here, we explore the effects of localization error and confinement on the velocity autocorrelation function, Cυ.

Getting RNA and protein in phase.

Nonmembrane-bound organelles such as RNA granules behave like dynamic droplets, but the molecular details of their assembly are poorly understood. Several recent papers identify structural features that drive granule assembly, shedding light on how phase transitions functionally organize the cell and may lead to pathological protein aggregation.

Nonthermal ATP-dependent fluctuations contribute to the in vivo motion of chromosomal loci.

Chromosomal loci jiggle in place between segregation events in prokaryotic cells and during interphase in eukaryotic nuclei. This motion seems random and is often attributed to brownian motion. However, we show here that locus dynamics in live bacteria and yeast are sensitive to metabolic activity.

Bacterial chromosomal loci move subdiffusively through a viscoelastic cytoplasm.

Tracking of fluorescently labeled chromosomal loci in live bacterial cells reveals a robust scaling of the mean square displacement (MSD) as τ(0.39). We propose that the observed motion arises from relaxation of the Rouse modes of the DNA polymer within the viscoelastic environment of the cytoplasm.

Subdiffusive motion of a polymer composed of subdiffusive monomers.

We use Brownian dynamics simulations and analytical theory to investigate the physical principles underlying subdiffusive motion of a polymer. Specifically, we examine the consequences of confinement, self-interaction, viscoelasticity, and random waiting on monomer motion, as these physical phenomena may be relevant to the behavior of biological macromolecules in vivo. We find that neither confinement nor self-interaction alter the fundamental Rouse mode relaxations of a polymer.

Mu gets in the loop.

In this issue of Molecular Cell, Han and Mizuuchi present evidence for a possible Turing-like reaction-diffusion mechanism underlying target immunity by the bacteriophage Mu.