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1. Splicing regulation through biomolecular condensates and membraneless organelles

   https://doi.org/10.1038/s41580-024-00739-7  

   Giudice, Jimena; Jiang, Hao (September 2024, Nature Reviews Molecular Cell Biology)

   Zlotorynski, Eytan (Ed.)

   Biomolecular condensates, sometimes also known as membraneless organelles (MLOs), can form through weak multivalent intermolecular interactions of proteins and nucleic acids, a process often associated with liquid-liquid phase separation. Biomolecular condensates are emerging as sites and regulatory platforms of vital cellular functions, including transcription and RNA processing. In the first part of this Review, we comprehensively discuss how alternative splicing regulates the formation and properties of condensates, and conversely the roles of biomolecular condensates in splicing regulation. In the second part, we focus on the spatial connection between splicing regulation and nuclear MLOs such as transcriptional condensates, splicing condensates and nuclear speckles. We then discuss key studies showing how splicing regulation through biomolecular condensates is implicated in human pathologies such as neurodegenerative diseases, different types of cancer, developmental disorders and cardiomyopathies, and conclude with a discussion of outstanding questions pertaining to the roles of condensates and MLOs in splicing regulation and how to experimentally study them. 

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2. The condensation of HP1α/Swi6 imparts nuclear stiffness

   https://doi.org/10.1016/j.celrep.2024.114373  

   Williams, Jessica F; Surovtsev, Ivan V; Schreiner, Sarah M; Chen, Ziyuan; Raiymbek, Gulzhan; Nguyen, Hang; Hu, Yan; Biteen, Julie S; Mochrie, Simon GJ; Ragunathan, Kaushik; et al (July 2024, Cell Reports)

   Biomolecular condensates have emerged as major drivers of cellular organization. It remains largely unexplored, however, whether these condensates can impart mechanical function(s) to the cell. The heterochromatin protein HP1α (Swi6 in Schizosaccharomyces pombe) crosslinks histone H3K9 methylated nucleosomes and has been proposed to undergo condensation to drive the liquid-like clustering of heterochromatin domains. Here, we leverage the genetically tractable S. pombe model and a separation-of-function allele to elucidate a mechanical function imparted by Swi6 condensation. Using single-molecule imaging, force spectroscopy, and high-resolution live-cell imaging, we show that Swi6 is critical for nuclear resistance to external force. Strikingly, it is the condensed yet dynamic pool of Swi6, rather than the chromatin-bound molecules, that is essential to imparting mechanical stiffness. Our findings suggest that Swi6 condensates embedded in the chromatin meshwork establish the emergent mechanical behavior of the nucleus as a whole, revealing that biomolecular condensation can influence organelle and cell mechanics. 

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3. A model for organization and regulation of nuclear condensates by gene activity

   https://doi.org/10.1038/s41467-023-39878-4  

   Schede, Halima_H; Natarajan, Pradeep; Chakraborty, Arup_K; Shrinivas, Krishna (July 2023, Nature Communications)

   Abstract Condensation by phase separation has recently emerged as a mechanism underlying many nuclear compartments essential for cellular functions. Nuclear condensates enrich nucleic acids and proteins, localize to specific genomic regions, and often promote gene expression. How diverse properties of nuclear condensates are shaped by gene organization and activity is poorly understood. Here, we develop a physics-based model to interrogate how spatially-varying transcription activity impacts condensate properties and dynamics. Our model predicts that spatial clustering of active genes can enable precise localization and de novo nucleation of condensates. Strong clustering and high activity results in aspherical condensate morphologies. Condensates can flow towards distant gene clusters and competition between multiple clusters lead to stretched morphologies and activity-dependent repositioning. Overall, our model predicts and recapitulates morphological and dynamical features of diverse nuclear condensates and offers a unified mechanistic framework to study the interplay between non-equilibrium processes, spatially-varying transcription, and multicomponent condensates in cell biology. 

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4. Overexpression of the microtubule-binding protein CLIP-170 induces a +TIP network superstructure consistent with a biomolecular condensate

   https://doi.org/10.1371/journal.pone.0260401  

   Wu, Yueh-Fu O.; Bryant, Annamarie T.; Nelson, Nora T.; Madey, Alexander G.; Fernandes, Gail F.; Goodson, Holly V. (December 2021, PLOS ONE)

   Acharya, Bipul R. (Ed.)

   Proper regulation of microtubule (MT) dynamics is critical for cellular processes including cell division and intracellular transport. Plus-end tracking proteins (+TIPs) dynamically track growing MTs and play a key role in MT regulation. +TIPs participate in a complex web of intra- and inter- molecular interactions known as the +TIP network. Hypotheses addressing the purpose of +TIP:+TIP interactions include relieving +TIP autoinhibition and localizing MT regulators to growing MT ends. In addition, we have proposed that the web of +TIP:+TIP interactions has a physical purpose: creating a dynamic scaffold that constrains the structural fluctuations of the fragile MT tip and thus acts as a polymerization chaperone. Here we examine the possibility that this proposed scaffold is a biomolecular condensate (i.e., liquid droplet). Many animal +TIP network proteins are multivalent and have intrinsically disordered regions, features commonly found in biomolecular condensates. Moreover, previous studies have shown that overexpression of the +TIP CLIP-170 induces large “patch” structures containing CLIP-170 and other +TIPs; we hypothesized that these structures might be biomolecular condensates. To test this hypothesis, we used video microscopy, immunofluorescence staining, and Fluorescence Recovery After Photobleaching (FRAP). Our data show that the CLIP-170-induced patches have hallmarks indicative of a biomolecular condensate, one that contains +TIP proteins and excludes other known condensate markers. Moreover, bioinformatic studies demonstrate that the presence of intrinsically disordered regions is conserved in key +TIPs, implying that these regions are functionally significant. Together, these results indicate that the CLIP-170 induced patches in cells are phase-separated liquid condensates and raise the possibility that the endogenous +TIP network might form a liquid droplet at MT ends or other +TIP locations. 

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5. Decoding the physical principles of two-component biomolecular phase separation

   https://doi.org/10.7554/eLife.62403  

   Zhang, Yaojun; Xu, Bin; Weiner, Benjamin G; Meir, Yigal; Wingreen, Ned S (March 2021, eLife)

   null (Ed.)

   Cells possess a multiplicity of non-membrane-bound compartments, which form via liquid-liquid phase separation. These condensates assemble and dissolve as needed to enable central cellular functions. One important class of condensates is those composed of two associating polymer species that form one-to-one specific bonds. What are the physical principles that underlie phase separation in such systems? To address this question, we employed coarse-grained molecular dynamics simulations to examine how the phase boundaries depend on polymer valence, stoichiometry, and binding strength. We discovered a striking phenomenon – for sufficiently strong binding, phase separation is suppressed at rational polymer stoichiometries, which we termed the magic-ratio effect. We further developed an analytical dimer-gel theory that confirmed the magic-ratio effect and disentangled the individual roles of polymer properties in shaping the phase diagram. Our work provides new insights into the factors controlling the phase diagrams of biomolecular condensates, with implications for natural and synthetic systems. 

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