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The fatal adult motor neuron disease amyotrophic lateral sclerosis (ALS) shares some clinical and pathological overlap with frontotemporal dementia (FTD), an early-onset neurodegenerative disorder. The RNA/DNA-binding proteins fused in sarcoma (FUS; also known as TLS) and TAR DNA binding protein-43 (TDP-43) have recently been shown to be genetically and pathologically associated with familial forms of ALS and FTD. It is currently unknown whether perturbation of these proteins results in disease through mechanisms that are independent of normal protein function or via the pathophysiological disruption of molecular processes in which they are both critical. Here, we report that Drosophila mutants in which the homolog of FUS is disrupted exhibit decreased adult viability, diminished locomotor speed, and reduced life span compared with controls. These phenotypes were fully rescued by wild-type human FUS, but not ALS-associated mutant FUS proteins. A mutant of the Drosophila homolog of TDP-43 had similar, but more severe, deficits. Through cross-rescue analysis, we demonstrated that FUS acted together with and downstream of TDP-43 in a common genetic pathway in neurons. Furthermore, we found that these proteins associated with each other in an RNA-dependent complex. Our results establish that FUS and TDP-43 function together in vivo and suggest that molecular pathways requiring the combined activities of both of these proteins may be disrupted in ALS and FTD.

Biomolecular condensation partitions cellular contents and has important roles in stress responses, maintaining homeostasis, development and disease. Many nuclear and cytoplasmic condensates are rich in RNA and RNA-binding proteins (RBPs), which undergo liquid–liquid phase separation (LLPS). Whereas the role of RBPs in condensates has been well studied, less attention has been paid to the contribution of RNA to LLPS. In this Review, we discuss the role of RNA in biomolecular condensation and highlight considerations for designing condensate reconstitution experiments. We focus on RNA properties such as composition, length, structure, modifications and expression level. These properties can modulate the biophysical features of native condensates, including their size, shape, viscosity, liquidity, surface tension and composition. We also discuss the role of RNA–protein condensates in development, disease and homeostasis, emphasizing how their properties and function can be determined by RNA. Finally, we discuss the multifaceted cellular functions of biomolecular condensates, including cell compartmentalization through RNA transport and localization, supporting catalytic processes, storage and inheritance of specific molecules, and buffering noise and responding to stress.Recent studies have highlighted the contribution of RNA to cellular liquid–liquid phase separation and condensate formation. RNA features modulate the composition and biophysical properties of RNA–protein condensates, which have various cellular functions, including RNA transport and localization, supporting catalytic processes and responding to stress.

Ferroptosis, an iron-dependent nonapoptotic cell death, is a highly regulated tumor suppressing process. However, functions and mechanisms of RNA-binding proteins in regulation of evasion of ferroptosis during lung cancer progression are still largely unknown. Here, we report that the RNA-binding protein RBMS1 participates in lung cancer development via mediating ferroptosis evasion. Through an shRNA-mediated systematic screen, we discovered that RBMS1 is a key ferroptosis regulator. Clinically, RBMS1 was elevated in lung cancer and its high expression was associated with reduced patient survival. Conversely, depletion of RBMS1 inhibited lung cancer progression both in vivo and in vitro. Mechanistically, RBMS1 interacted with the translation initiation factor eIF3d directly to bridge the 3'- and 5'-UTR of SLC7A11. RBMS1 ablation inhibited the translation of SLC7A11, reduced SLC7A11-mediated cystine uptake, and promoted ferroptosis. In a drug screen that targeted RBMS1, we further uncovered that nortriptyline hydrochloride decreased the level of RBMS1, thereby promoting ferroptosis. Importantly, RBMS1 depletion or inhibition by nortriptyline hydrochloride sensitized radioresistant lung cancer cells to radiotherapy. Our findings established RBMS1 as a translational regulator of ferroptosis and a prognostic factor with therapeutic potential and clinical value.

RNA-binding protein (RBP) has a highly dynamic spatiotemporal regulation process and important biological functions. They are critical to maintain the transcriptome through post-transcriptionally controlling the processing and transportation of RNA, including regulating RNA splicing, polyadenylation, mRNA stability, mRNA localization, and translation. Alteration of each process will affect the RNA life cycle, produce abnormal protein phenotypes, and thus lead to the occurrence and development of tumors. Here, we summarize RBPs involved in tumor progression and the underlying molecular mechanisms whereby they are regulated and exert their effects. This analysis is an important step towards the comprehensive characterization of post-transcriptional gene regulation involved in tumor progression.

The PIWI-interacting RNA (piRNA) pathway protects genome integrity in part through establishing repressive heterochromatin at transposon loci. Silencing requires piRNA-guided targeting of nuclear PIWI proteins to nascent transposon transcripts, yet the subsequent molecular events are not understood. Here, we identify SFiNX (silencing factor interacting nuclear export variant), an interdependent protein complex required for Piwi-mediated cotranscriptional silencing in Drosophila. SFiNX consists of Nxf2–Nxt1, a gonad-specific variant of the heterodimeric messenger RNA export receptor Nxf1–Nxt1 and the Piwi-associated protein Panoramix. SFiNX mutant flies are sterile and exhibit transposon derepression because piRNA-loaded Piwi is unable to establish heterochromatin. Within SFiNX, Panoramix recruits heterochromatin effectors, while the RNA binding protein Nxf2 licenses cotranscriptional silencing. Our data reveal how Nxf2 might have evolved from an RNA transport receptor into a cotranscriptional silencing factor. Thus, NXF variants, which are abundant in metazoans, can have diverse molecular functions and might have been coopted for host genome defense more broadly.

Stress granules (SGs) are membraneless cell compartments formed in response to different stress stimuli, wherein translation factors, mRNAs, RNA-binding proteins (RBPs) and other proteins coalesce together. SGs assembly is crucial for cell survival, since SGs are implicated in the regulation of translation, mRNA storage and stabilization and cell signalling, during stress. One defining feature of SGs is their dynamism, as they are quickly assembled upon stress and then rapidly dispersed after the stress source is no longer present. Recently, SGs dynamics, their components and their functions have begun to be studied in the context of human diseases. Interestingly, the regulated protein self-assembly that mediates SG formation contrasts with the pathological protein aggregation that is a feature of several neurodegenerative diseases. In particular, aberrant protein coalescence is a key feature of polyglutamine (PolyQ) diseases, a group of nine disorders that are caused by an abnormal expansion of PolyQ tract-bearing proteins, which increases the propensity of those proteins to aggregate. Available data concerning the abnormal properties of the mutant PolyQ disease-causing proteins and their involvement in stress response dysregulation strongly suggests an important role for SGs in the pathogenesis of PolyQ disorders. This review aims at discussing the evidence supporting the existence of a link between SGs functionality and PolyQ disorders, by focusing on the biology of SGs and on the way it can be altered in a PolyQ disease context.

Most N6-methyladenosine (m 6 A) associated regulatory proteins (i.e., m 6 A writer, eraser, and reader proteins) are involved in the pathogenesis of various cancers, mostly in m 6 A-dependent manners. As a component in the m 6 A ‘writers’, KIAA1429 is reported to be an RNA-binding protein and involved in the m 6 A modification, mRNA splicing and processing. Till now, the functions of KIAA1429 in tumorigenesis and related mechanism have not been reported. In the present study, we found KIAA1429 was highly expressed in breast cancer tissues, but frequently down-regulated in non-cancerous breast tissues. The overall survival of breast cancer patients with high-expression KIAA1429 was significantly shorter than those with low-expression KIAA1429. Then, we demonstrated that KIAA1429 was associated with breast cancer proliferation and metastasis in vivo and in vitro. The potential targeting genes of KIAA1429 in breast cancer were identified by RNA immunoprecipitation sequencing. One of these genes is cyclin-dependent kinase 1 (CDK1), which plays an oncogenic role in cancers. Furthermore, we confirmed that KIAA1429 played its oncogenic role in breast cancer by regulating CDK1 by an m 6 A-independent manner. 5′-fluorouracil was found to be very effective in reducing the expression of KIAA1429 and CDK1 in breast cancer. These findings indicated that KIAA1429 could promote breast cancer progression and was correlated with pathogenesis. It may represent a promising therapeutic strategy on breast cancer, especially in combination with CDK1 treatment.

Interferon-induced transmembrane proteins (IFITMs 1, 2 and 3) have emerged as important innate immune effectors that prevent diverse virus infections in vertebrates. However, the cellular mechanisms and live-cell imaging of these small membrane proteins have been challenging to evaluate during viral entry of mammalian cells. Using CRISPR–Cas9-mediated IFITM-mutant cell lines, we demonstrate that human IFITM1, IFITM2 and IFITM3 act cooperatively and function in a dose-dependent fashion in interferon-stimulated cells. Through site-specific fluorophore tagging and live-cell imaging studies, we show that IFITM3 is on endocytic vesicles that fuse with incoming virus particles and enhances the trafficking of this pathogenic cargo to lysosomes. IFITM3 trafficking is specific to restricted viruses, requires S-palmitoylation and is abrogated with loss-of-function mutants. The site-specific protein labeling and live-cell imaging approaches described here should facilitate the functional analysis of host factors involved in pathogen restriction as well as their mechanisms of regulation. Live-cell imaging and virus trafficking studies show that the host innate immune receptor IFITM3 localizes with endocytic vesicles that fuse with incoming viruses to ultimately enhance their traffic to lysosomes.

Key messageFunctions of most splice isoforms that are generated by alternative splicing are unknown. We show that two splice variants that encode proteins differing in only eight amino acids have distinct functions in a stress response.Serine/arginine-rich (SR) and SR-like proteins, a conserved family of RNA binding proteins across eukaryotes, play important roles in pre-mRNA splicing and other post-transcriptional processes. Pre-mRNAs of SR and SR-like proteins undergo extensive alternative splicing in response to diverse stresses and produce multiple splice isoforms. However, the functions of most splice isoforms remain elusive. Alternative splicing of pre-mRNA of Arabidopsis SR45, which encodes an SR-like splicing regulator, generates two isoforms (long—SR45.1 and short—SR45.2). The proteins encoded by these two isoforms differ in eight amino acids. Here, we investigated the role of SR45 and its splice variants in salt stress tolerance. The loss of SR45 resulted in enhanced sensitivity to salt stress and changes in expression and splicing of genes involved in regulating salt stress response. Interestingly, only the long isoform (SR45.1) rescued the salt-sensitive phenotype as well as the altered gene expression and splicing patterns in the mutant. These results suggest that SR45 positively regulates salt tolerance. Furthermore, only the long isoform is required for SR45-mediated salt tolerance.

TDP-43 regulates alternative splicing of the cystic fibrosis transmembrane regulator (CFTR) and is found in cytosolic granules associated with several neurodegenerative disorders. A new solution structure of the tandem RNA-recognition motifs (RRMs) that mediate interactions with its UG-rich RNA targets reveals a new RRM arrangement critical for TDP-43 function. TDP-43 encodes an alternative-splicing regulator with tandem RNA-recognition motifs (RRMs). The protein regulates cystic fibrosis transmembrane regulator ( CFTR ) exon 9 splicing through binding to long UG-rich RNA sequences and is found in cytoplasmic inclusions of several neurodegenerative diseases. We solved the solution structure of the TDP-43 RRMs in complex with UG-rich RNA. Ten nucleotides are bound by both RRMs, and six are recognized sequence specifically. Among these, a central G interacts with both RRMs and stabilizes a new tandem RRM arrangement. Mutations that eliminate recognition of this key nucleotide or crucial inter-RRM interactions disrupt RNA binding and TDP-43–dependent splicing regulation. In contrast, point mutations that affect base-specific recognition in either RRM have weaker effects. Our findings reveal not only how TDP-43 recognizes UG repeats but also how RNA binding–dependent inter-RRM interactions are crucial for TDP-43 function.