TIA-1 is an RNA-binding protein that sequesters target RNA into stress granules under conditions of cellular stress. Promotion of stress granule formation by TIA-1 depends upon self-association of its prion-like domain that facilitates liquid-liquid phase separation and is thought to be enhanced via RNA binding. However, the mechanisms underlying the influence of RNA on TIA-1 self-association have not been previously demonstrated. Here we have investigated the self-associating properties of full-length TIA-1 in the presence of designed and native TIA-1 nucleic acid binding sites in vitro, monitoring phase separation, fibril formation and shape. We show that single stranded RNA and DNA induce liquid-liquid phase separation of TIA-1 in a multisite, sequence-specific manner and also efficiently promote formation of amyloid-like fibrils. Although RNA binding to a single site induces a small conformational change in TIA-1, this alone does not enhance phase separation of TIA-1. Tandem binding sites are required to enhance phase separation of TIA-1 and this is finely tuned by the protein:binding site stoichiometry rather than nucleic acid length. Native tandem TIA-1 binding sites within the 3′ UTR of p53 mRNA also efficiently enhance phase separation of TIA-1 and thus may potentially act as potent nucleation sites for stress granule assembly.
Stress granules are non-membrane bound RNA-protein granules essential for survival during acute cellular stress. TIA-1 is a key protein in the formation of stress granules that undergoes liquid-liquid phase separation by association with specific RNAs and protein-protein interactions. However, the fundamental properties of the TIA-1 protein that enable phase-separation also render TIA-1 susceptible to the formation of irreversible fibrillar aggregates. Despite this, within physiological stress granules, TIA-1 is not present as fibrils, pointing to additional factors within the cell that prevent TIA-1 aggregation. Here we show that heterotypic interactions with stress granule co-factors Zn2+ and RGG-rich regions from FUS each act together with nucleic acid to induce the liquid-liquid phase separation of TIA-1. In contrast, these co-factors do not enhance nucleic acid induced fibril formation of TIA-1, but rather robustly inhibit the process. NMR titration experiments revealed specific interactions between Zn2+ and H94 and H96 in RRM2 of TIA-1. Strikingly, this interaction promotes multimerization of TIA-1 independently of the prion-like domain. Thus, through different molecular mechanisms, these stress granule co-factors promote TIA-1 liquid-liquid phase separation and suppress fibrillar aggregates, potentially contributing to the dynamic nature of stress granules and the cellular protection that they provide.
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.
hi@scite.ai
10624 S. Eastern Ave., Ste. A-614
Henderson, NV 89052, USA
Copyright © 2024 scite LLC. All rights reserved.
Made with 💙 for researchers
Part of the Research Solutions Family.