RESUMEN
Prion-like proteins can assume distinct conformational and physical states in the same cell. Sequence analysis suggests that prion-like proteins are prevalent in various species; however, it remains unclear what functional space they occupy in multicellular organisms. Here, we report the identification of a prion-like protein, Herzog (CG5830), through a multimodal screen in Drosophila melanogaster. Herzog functions as a membrane-associated phosphatase and controls embryonic patterning, likely being involved in TGF-ß/BMP and FGF/EGF signaling pathways. Remarkably, monomeric Herzog is enzymatically inactive and becomes active upon amyloid-like assembly. The prion-like domain of Herzog is necessary for both its assembly and membrane targeting. Removal of the prion-like domain impairs activity, while restoring assembly on the membrane using a heterologous prion-like domain and membrane-targeting motif can restore phosphatase activity. This study provides an example of a prion-like domain that allows an enzyme to gain essential functionality via amyloid-like assembly to control animal development.
Asunto(s)
Proteínas Amiloidogénicas/metabolismo , Proteínas de Drosophila/metabolismo , Drosophila melanogaster/embriología , Desarrollo Embrionario , Fosfoproteínas Fosfatasas/metabolismo , Monoéster Fosfórico Hidrolasas/metabolismo , Proteínas Amiloidogénicas/química , Proteínas Amiloidogénicas/genética , Animales , Proteínas de Drosophila/química , Proteínas de Drosophila/genética , Drosophila melanogaster/genética , Drosophila melanogaster/metabolismo , Fosfoproteínas Fosfatasas/química , Fosfoproteínas Fosfatasas/genética , Monoéster Fosfórico Hidrolasas/química , Monoéster Fosfórico Hidrolasas/genética , Priones/química , Dominios ProteicosRESUMEN
Model organisms are widely used in research as accessible and convenient systems to study a particular area or question in biology. Traditionally only a handful of organisms have been widely studied, but modern research tools are enabling researchers to extend the set of model organisms to include less-studied and more unusual systems. This Forum highlights a range of 'non-model model organisms' as emerging systems for tackling questions across the whole spectrum of biology (and beyond), the opportunities and challenges, and the outlook for the future.
Asunto(s)
Biología , Eucariontes , Modelos Animales , Animales , PlantasRESUMEN
Biomolecular condensates organize biochemistry, yet little is known about how cells control the position and scale of these structures. In cells, condensates often appear as relatively small assemblies that do not coarsen into a single droplet despite their propensity to fuse. Here, we report that ribonucleoprotein condensates of the glutamine-rich protein Whi3 interact with the endoplasmic reticulum, which prompted us to examine how membrane association controls condensate size. Reconstitution revealed that membrane recruitment promotes Whi3 condensation under physiological conditions. These assemblies rapidly arrest, resembling size distributions seen in cells. The temporal ordering of molecular interactions and the slow diffusion of membrane-bound complexes can limit condensate size. Our experiments reveal a trade-off between locally enhanced protein concentration at membranes, which favours condensation, and an accompanying reduction in diffusion, which restricts coarsening. Given that many condensates bind endomembranes, we predict that the biophysical properties of lipid bilayers are key for controlling condensate sizes throughout the cell.
Asunto(s)
Ribonucleoproteínas , Ribonucleoproteínas/genéticaRESUMEN
Glutamine-rich tracts, also known as polyQ domains, have received a great deal of attention for their role in multiple neurodegenerative diseases, including Huntington's disease (HD), spinocerebellar ataxia (SCA), and others [22], [27]. Expansions in the normal polyQ tracts are thus commonly linked to disease, but polyQ domains themselves play multiple important functional roles in cells that are being increasingly appreciated. The biochemical nature of these domains allows them to adopt a number of different structures and form large assemblies that enable environmental responsiveness, localized signaling, and cellular memory. In many cases, these involve the formation of condensates that have varied material states. In this review, we highlight known and emerging functional roles for polyQ tracts in normal cell physiology.
Asunto(s)
Enfermedad de Huntington , Péptidos/química , HumanosRESUMEN
Biomolecular condensation is a way of organizing cytosol in which proteins and nucleic acids coassemble into compartments. In the multinucleate filamentous fungus Ashbya gossypii, the RNA-binding protein Whi3 regulates the cell cycle and cell polarity through forming macromolecular structures that behave like condensates. Whi3 has distinct spatial localizations and mRNA targets, making it a powerful model for how, when, and where specific identities are established for condensates. We identified residues on Whi3 that are differentially phosphorylated under specific conditions and generated mutants that ablate this regulation. This yielded separation of function alleles that were functional for either cell polarity or nuclear cycling but not both. This study shows that phosphorylation of individual residues on molecules in biomolecular condensates can provide specificity that gives rise to distinct functional identities in the same cell.
Asunto(s)
Ciclo Celular/genética , Polaridad Celular/genética , Eremothecium/metabolismo , Proteínas Fúngicas/metabolismo , Procesamiento Proteico-Postraduccional , Proteínas de Unión al ARN/metabolismo , Alelos , Secuencia de Bases , Compartimento Celular/genética , Citosol/metabolismo , Citosol/ultraestructura , Eremothecium/genética , Eremothecium/ultraestructura , Proteínas Fúngicas/genética , Expresión Génica , Calor , Mutación , Fosforilación , ARN de Hongos/genética , ARN de Hongos/metabolismo , ARN Mensajero/genética , ARN Mensajero/metabolismo , Proteínas de Unión al ARN/genética , Estrés Fisiológico/genéticaRESUMEN
RNA promotes liquid-liquid phase separation (LLPS) to build membraneless compartments in cells. How distinct molecular compositions are established and maintained in these liquid compartments is unknown. Here, we report that secondary structure allows messenger RNAs (mRNAs) to self-associate and determines whether an mRNA is recruited to or excluded from liquid compartments. The polyQ-protein Whi3 induces conformational changes in RNA structure and generates distinct molecular fluctuations depending on the RNA sequence. These data support a model in which structure-based, RNA-RNA interactions promote assembly of distinct droplets and protein-driven, conformational dynamics of the RNA maintain this identity. Thus, the shape of RNA can promote the formation and coexistence of the diverse array of RNA-rich liquid compartments found in a single cell.
Asunto(s)
Péptidos/química , Transición de Fase , ARN Mensajero/química , Proteínas de Unión al ARN/química , Proteínas de Saccharomyces cerevisiae/química , Secuencia de Bases , Ciclinas/química , Conformación de Ácido NucleicoRESUMEN
Fluorescence microscopy is a powerful approach for studying subcellular dynamics at high spatiotemporal resolution; however, conventional fluorescence microscopy techniques are light-intensive and introduce unnecessary photodamage. Light-sheet fluorescence microscopy (LSFM) mitigates these problems by selectively illuminating the focal plane of the detection objective by using orthogonal excitation. Orthogonal excitation requires geometries that physically limit the detection objective numerical aperture (NA), thereby limiting both light-gathering efficiency (brightness) and native spatial resolution. We present a novel live-cell LSFM method, lateral interference tilted excitation (LITE), in which a tilted light sheet illuminates the detection objective focal plane without a sterically limiting illumination scheme. LITE is thus compatible with any detection objective, including oil immersion, without an upper NA limit. LITE combines the low photodamage of LSFM with high resolution, high brightness, and coverslip-based objectives. We demonstrate the utility of LITE for imaging animal, fungal, and plant model organisms over many hours at high spatiotemporal resolution.