Liquid-liquid phase separation is thought to underly gene transcription, through the condensation of the large-scale nucleolus, or in smaller assemblies known as transcriptional hubs or condensates. However, phase separation has not yet been directly linked with transcriptional output, and our biophysical understanding of transcription dynamics is poor. Here, we utilize an optogenetic approach to control condensation of key FET-family transcriptional regulators, particularly TAF15. We show that amino acid sequence-dependent phase separation of TAF15 is enhanced significantly due to strong nuclear interactions with the C-terminal domain (CTD) of RNA Pol II. Nascent CTD clusters at primed genomic loci lower the energetic barrier for nucleation of TAF15 condensates, which in turn further recruit RNA Pol II to drive transcriptional output. These results suggest a model in which positive feedback between key transcriptional components drives intermittent dynamics of localized phase separation, to amplify gene expression.Cells organize complex biochemical reactions through compartmentalization into various organelles. Many of these organelles are membrane-less condensates, which form through liquid-liquid phase separation (LLPS), in which biomolecules condense into dynamic liquid droplets [1][2][3][4] . Nuclear condensates are thought to play key roles in regulating the flow of genetic information and include a diverse set of structures ranging from large scale assemblies such as nuclear speckles and the nucleolus, to structures that form on smaller length scales such as Cajal bodies, promyelocytic leukemia (PML) bodies, and gems.The concept that LLPS can drive transcriptional activity has recently received intense attention. The nucleolus is a large nuclear condensate which assembles around actively transcribing ribosomal RNA (rRNA) loci 5,6 , and is thought to facilitate the processing and maturation of rRNA into pre-ribosomal particles. Smaller transcriptional condensates have been proposed to assemble throughout the genome to facilitate transcription of non-ribosomal genes through the RNA polymerase II (Pol II) machinery [7][8][9][10] . These nanoscale transcriptional condensates have been hypothesized to facilitate enhancer-promoter interactions [11][12][13] , possibly through the generation of localized force through surface-tension driven droplet coalescence 14 ..
The abundant accumulation of inclusion bodies containing polyglutamine-expanded mutant huntingtin (mHTT) aggregates is considered as the key pathological event in Huntington’s disease (HD). Here, we demonstrate that FKBP12, an isomerase that exhibits reduced expression in HD, decreases the amyloidogenicity of mHTT, interrupts its oligomerization process, and structurally promotes the formation of amorphous deposits. By combining fluorescence-activated cell sorting with multiple biophysical techniques, we confirm that FKBP12 reduces the amyloid property of these ultrastructural-distinct mHTT aggregates within cells. Moreover, the neuroprotective effect of FKBP12 is demonstrated in both cellular and nematode models. Finally, we show that FKBP12 also inhibit the fibrillization process of other disease-related and aggregation-prone peptides. Our results suggest a novel function of FKBP12 in ameliorating the proteotoxicity in mHTT, which may shed light on unraveling the roles of FKBP12 in different neurodegenerative diseases and developing possible therapeutic strategies.
Liquid-liquid phase separation is thought to underly gene transcription, through the condensation of the large-scale nucleolus, or in smaller assemblies known as transcriptional hubs or condensates. However, phase separation has not yet been directly linked with transcriptional output, and our biophysical understanding of transcription dynamics is poor. Here, we utilize an optogenetic approach to control condensation of key FET-family transcriptional regulators, particularly TAF15. We show that amino acid sequence-dependent phase separation of TAF15 is enhanced significantly due to strong nuclear interactions with the C-terminal domain (CTD) of RNA Pol II. Nascent CTD clusters at primed genomic loci lower the energetic barrier for nucleation of TAF15 condensates, which in turn further recruit RNA Pol II to drive transcriptional output. These results suggest a model in which positive feedback between key transcriptional components drives intermittent dynamics of localized phase separation, to amplify gene expression.Cells organize complex biochemical reactions through compartmentalization into various organelles. Many of these organelles are membrane-less condensates, which form through liquid-liquid phase separation (LLPS), in which biomolecules condense into dynamic liquid droplets 1-4 . Nuclear condensates are thought to play key roles in regulating the flow of genetic information and include a diverse set of structures ranging from large scale assemblies such as nuclear speckles and the nucleolus, to structures that form on smaller length scales such as Cajal bodies, promyelocytic leukemia (PML) bodies, and gems.The concept that LLPS can drive transcriptional activity has recently received intense attention. The nucleolus is a large nuclear condensate which assembles around actively transcribing ribosomal RNA (rRNA) loci 5, 6 , and is thought to facilitate the processing and maturation of rRNA into pre-ribosomal particles. Smaller transcriptional condensates have been proposed to assemble throughout the genome to facilitate transcription of non-ribosomal genes through the RNA polymerase II (Pol II) machinery 7-10 . These nanoscale transcriptional condensates have been hypothesized to facilitate enhancer-promoter interactions 11-13 , possibly through the generation of localized force through surface-tension driven droplet coalescence 14 .
SummaryBiomolecular condensates assemble in living cells through phase separation and related phase transitions. An underappreciated feature of these dynamic molecular assemblies is that they form interfaces with cellular structures, including membranes, cytoskeleton, DNA and RNA, and other membraneless compartments. These interfaces are expected to give rise to capillary forces, but there are few ways of quantifying and harnessing these forces in living cells. Here, we introduce VECTOR (ViscoElasticChromatinTethering andORganization), which uses light-inducible biomolecular condensates to generate capillary forces at targeted DNA loci. VECTOR can be utilized to programmably reposition genomic loci on a timescale of seconds to minutes, quantitatively revealing local heterogeneity in the viscoelastic material properties of chromatin. These synthetic condensates are built from components that naturally form liquid-like structures in living cells, highlighting the potential role for native condensates to generate forces and do work to reorganize the genome and impact chromatin architecture.
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