Protein-based condensation mechanisms drive the assembly of RNA-rich P granules.
Ontology highlight
ABSTRACT:
SUBMITTER: Schmidt H
PROVIDER: S-EPMC8238508 | biostudies-literature |
REPOSITORIES: biostudies-literature
Similar Datasets
Project description:Stressed cells shut down translation, release mRNA molecules from polysomes, and form stress granules (SGs) via a network of interactions that involve G3BP. Here we focus on the mechanistic underpinnings of SG assembly. We show that, under non-stress conditions, G3BP adopts a compact auto-inhibited state stabilized by electrostatic intramolecular interactions between the intrinsically disordered acidic tracts and the positively charged arginine-rich region. Upon release from polysomes, unfolded mRNAs outcompete G3BP auto-inhibitory interactions, engendering a conformational transition that facilitates clustering of G3BP through protein-RNA interactions. Subsequent physical crosslinking of G3BP clusters drives RNA molecules into networked RNA/protein condensates. We show that G3BP condensates impede RNA entanglement and recruit additional client proteins that promote SG maturation or induce a liquid-to-solid transition that may underlie disease. We propose that condensation coupled to conformational rearrangements and heterotypic multivalent interactions may be a general principle underlying RNP granule assembly.
| S-EPMC7181197 | biostudies-literature
Project description:Cells form and use biomolecular condensates to execute biochemical reactions. The molecular properties of non-membrane-bound condensates are directly connected to the amino acid content of disordered protein regions. Lysine plays an important role in cellular function, but little is known about its role in biomolecular condensation. Here we show that protein disorder is abundant in protein/RNA granules and lysine is enriched in disordered regions of proteins in P-bodies compared to the entire human disordered proteome. Lysine-rich polypeptides phase separate into lysine/RNA-coacervates that are more dynamic and differ at the molecular level from arginine/RNA-coacervates. Consistent with the ability of lysine to drive phase separation, lysine-rich variants of the Alzheimer's disease-linked protein tau undergo coacervation with RNA in vitro and bind to stress granules in cells. Acetylation of lysine reverses liquid-liquid phase separation and reduces colocalization of tau with stress granules. Our study establishes lysine as an important regulator of cellular condensation.
| S-EPMC6606616 | biostudies-literature
Project description:The antimetabolite 5-fluorouracil is a widely used chemotherapeutic for the treatment of several solid cancers. However, resistance to 5-fluorouracil remains a major drawback in its clinical use. In this study we report that treatment of HeLa cells with 5-fluorouracil resulted in de novo assembly of stress granules. Moreover, we revealed that stress granule assembly under stress conditions as well as disassembly is altered in cells treated with 5-fluorouracil. Notably, we discovered that RACK1, a protein mediating cell survival and apoptosis, is a component of 5-fluorouracil-induced stress granules. To explore the mode of action of 5-fluorouracil accountable for de novo stress granule assembly, we analyzed 5-fluorouracil metabolites and noticed that stress granule assembly is caused by RNA, not DNA incorporating 5-fluorouracil metabolites. Interestingly, we observed that other RNA incorporating drugs also cause assembly of stress granules. Thus, our results suggest that incorporation of chemotherapeutics into RNA may result in stress granule assembly with potential significance in chemoresistance.
| S-EPMC4041438 | biostudies-literature
Project description:Human mitochondrial ribosomes are specialized in the synthesis of 13 proteins, which are fundamental components of the oxidative phosphorylation system. The pathway of mitoribosome biogenesis, the compartmentalization of the process, and factors involved remain largely unknown. Here, we have identified the DEAD-box protein DDX28 as an RNA granule component essential for the biogenesis of the mitoribosome large subunit (mt-LSU). DDX28 interacts with the 16S rRNA and the mt-LSU. RNAi-mediated DDX28 silencing in HEK293T cells does not affect mitochondrial mRNA stability or 16S rRNA processing or modification. However, it leads to reduced levels of 16S rRNA and mt-LSU proteins, impaired mt-LSU assembly, deeply attenuated mitochondrial protein synthesis, and consequent failure to assemble oxidative phosphorylation complexes. Our findings identify DDX28 as essential during the early stages of mitoribosome mt-LSU biogenesis, a process that takes place mainly near the mitochondrial nucleoids, in the compartment defined by the RNA granules.
| S-EPMC4534351 | biostudies-literature
Project description:Ribonucleoprotein (RNP) granules are RNA-protein assemblies that are involved in multiple aspects of RNA metabolism and are linked to memory, development, and disease. Some RNP granules form, in part, through the formation of intermolecular RNA-RNA interactions. In vitro, such trans RNA condensation occurs readily, suggesting that cells require mechanisms to modulate RNA-based condensation. We assess the mechanisms of RNA condensation and how cells modulate this phenomenon. We propose that cells control RNA condensation through ATP-dependent processes, static RNA buffering, and dynamic post-translational mechanisms. Moreover, perturbations in these mechanisms can be involved in disease. This reveals multiple cellular mechanisms of kinetic and thermodynamic control that maintain the proper distribution of RNA molecules between dispersed and condensed forms.
| S-EPMC7211619 | biostudies-literature
Project description:Proteins with low complexity, disordered sequences are receiving increasing attention due to their central roles in the biogenesis and regulation of membraneless organelles. In eukaryotic organisms, a substantial fraction of disordered proteins reside in the nucleus, thereby facilitating the formation of nuclear bodies, nucleolus, and chromatin compartmentalization. The heterochromatin family of proteins (HP1) is an important player in driving the formation of gene silenced mesoscopic heterochromatin B compartments and pericentric regions. Recent experiments have shown that the HP1a sequence of Drosophila melanogaster can undergo liquid-liquid phase separation under both in vitro and in vivo conditions, induced by changes of the monovalent salt concentration. While the phase separation of HP1a is thought to be the mechanism underlying chromatin compartmentalization, the molecular level mechanistic picture of salt-driven phase separation of HP1a has remained poorly understood. The disordered hinge region of HP1a is seen as the driver of salt-induced condensation because of its charge enriched sequence and post-translational modifications. Here, we set out to decipher the mechanisms of salt-induced condensation of HP1a through a systematic study of salt-dependent conformations of single chains and fuzzy dimers of disordered HP1a hinge sequences. Using multiple independent all-atom simulations with and without enhanced sampling, we carry out detailed characterization of conformational ensembles of disordered HP1a chains under different ionic conditions using various polymeric and structural measures. We show that the mobile ion release, enhancement of local transient secondary structural elements, and side-chain exposure to solvent are robust trends that accompany fuzzy dimer formation. Furthermore, we find that salt-induced changes in the ensemble of conformations of HP1a disordered hinge sequence fine-tune the inter-chain vs. self-chain interactions in ways that favor fuzzy dimer formation under low salt conditions in the agreement with condensation trends seen in experiments.
| S-EPMC8234964 | biostudies-literature
Project description:The germline originates from primordial embryonic germ cells which give rise to sperm and egg cells and consequently, to the next generation. Germ cells of many organisms contain electron-dense granules that comprise RNA and proteins indispensable for germline development. Here we review recent reports that provide important insights into the structure and function of crucial RNA and protein components of the granules, including DEAD-box helicases, Tudor domain proteins, Piwi/Argonaute proteins and piRNA. Collectively, these components function in translational control, remodeling of ribonucleoprotein complexes and transposon silencing. Furthermore, they interact with each other by means of conserved structural modules and post-translationally modified amino acids. These data suggest a widespread use of several protein motifs in germline development and further our understanding of other ribonucleoprotein structures, for example, processing bodies and neuronal granules.
| S-EPMC2929181 | biostudies-literature
Project description:Germ granules are biomolecular condensates that promote germ cell totipotency in animals. In C. elegans, MEG-3 and MEG-4 function redundantly to assemble germ granules in germline blastomeres. Here, we show that meg-3/4 mutant animals exhibit defects in RNA interference (RNAi) that are transgenerationally disconnected from the meg-3/4 genotype. Similar non-Mendelian inheritance is associated with other mutations disrupting germ granule formation, indicating that loss of germ granules is the likely cause of the observed disconnects between genotype and phenotype. meg-3/4 animals produce aberrant siRNAs that are propagated for ?10 generations in wild-type descendants of meg-3/4 ancestors. Aberrant siRNAs inappropriately and heritably silence germline-expressed genes including the RNAi gene sid-1, suggesting that transgenerational silencing of sid-1 underlies inherited defects in RNAi. We conclude that one function of germ granules is to organize RNA-based epigenetic inheritance pathways and that germ granule loss has consequences that persist for many generations.
| S-EPMC7316138 | biostudies-literature
Project description:Stress granules are condensates of non-translating mRNAs and proteins involved in the stress response and neurodegenerative diseases. Stress granules form in part through intermolecular RNA-RNA interactions, and to better understand how RNA-based condensation occurs, we demonstrate that RNA is effectively recruited to the surfaces of RNA or RNP condensates in vitro. We demonstrate that, through ATP-dependent RNA binding, the DEAD-box protein eIF4A reduces RNA condensation in vitro and limits stress granule formation in cells. This defines a function for eIF4A to limit intermolecular RNA-RNA interactions in cells. These results establish an important role for eIF4A, and potentially other DEAD-box proteins, as ATP-dependent RNA chaperones that limit the condensation of RNA, analogous to the function of proteins like HSP70 in combatting protein aggregates.
| S-EPMC7194247 | biostudies-literature
Project description:Regulated transport and local translation of mRNA in neurons are critical for modulating synaptic strength, maintaining proper neural circuitry, and establishing long term memory. Neuronal RNA granules are ribonucleoprotein particles that serve to transport mRNA along microtubules and control local protein synthesis in response to synaptic activity. Studies suggest that neuronal RNA granules share similar structures and functions with somatic P-bodies. We recently reported that the Huntington disease protein huntingtin (Htt) associates with Argonaute (Ago) and localizes to cytoplasmic P-bodies, which serve as sites of mRNA storage, degradation, and small RNA-mediated gene silencing. Here we report that wild-type Htt associates with Ago2 and components of neuronal granules and co-traffics with mRNA in dendrites. Htt was found to co-localize with RNA containing the 3'-untranslated region sequence of known dendritically targeted mRNAs. Knockdown of Htt in neurons caused altered localization of mRNA. When tethered to a reporter construct, Htt down-regulated reporter gene expression in a manner dependent on Ago2, suggesting that Htt may function to repress translation of mRNAs during transport in neuronal granules.
| S-EPMC2857123 | biostudies-literature