Biology subjects

Anderson, E. N.

Publications and source records attributed to Anderson, E. N..

6 recordsLinked to original sources

bioRxiv2026

Breaking β-sheets in FUS prion-like domain preserves phase separation and function but prevents aggregation and toxicity

The RNA-binding protein Fused in Sarcoma (FUS) undergoes phase separation associated with RNA processing. However, the prion-like low complexity (LC) domain of FUS forms solid-like aggregates in neurodegenerative diseases. Whether the formation of {beta}-sheet structure associated with pathology is also physiologically/functionally relevant is debated. Similarly, if mislocalization alone or concomitant aggregation is responsible for FUS gain-of-function toxicity remains to be probed. Here, we introduce {beta}-sheet breaking proline residues into FUS LC with the goal of preventing cross-{beta}-driven aggregation without disrupting essential functions and phase separation. {beta}-sheet-deficient FUS variants maintain native-like global motions, disorder, and phase separation, but no longer show a liquid-to-solid transition (LST). Biochemical partitioning, cellular localization, and auto- and cross-regulatory functions of FUS all remain essentially unchanged. Conversely, FUS-induced neurodegeneration in several Drosophila models is drastically reduced. These findings suggest a strategy for mitigating disease-related toxicity through backbone structure modulation to prevent prion-like domain protein aggregation. GRAPHICAL ABSTRACT O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=198 SRC="FIGDIR/small/706410v1_ufig1.gif" ALT="Figure 1"> View larger version (53K): org.highwire.dtl.DTLVardef@d15f63org.highwire.dtl.DTLVardef@1cd6221org.highwire.dtl.DTLVardef@e58126org.highwire.dtl.DTLVardef@181ec67_HPS_FORMAT_FIGEXP M_FIG C_FIG SUMMARYThe RNA-binding protein Fused in Sarcoma (FUS) undergoes phase separation as part of its physiological function but can aberrantly aggregate into solid-like assemblies in amyotrophic lateral sclerosis and frontotemporal dementia. To dissect the role of {beta}-sheets in both function and pathological transition, we engineered {beta}-sheet-preventing FUS variants via targeted proline residue insertions in the prion-like disordered region. These variants retained native structure, motions, and phase behavior yet showed dramatically reduced aggregation, both as an isolated prion-like domain and in full-length FUS. Crucially, these variants maintained a panel of FUS cellular functions that depend on FUS condensation but prevented FUS toxicity in fly models of neurodegeneration. Our findings implicate {beta}-sheets as key drivers of FUS condensate maturation and neuronal toxicity, highlighting {beta}-sheet modulation as a therapeutic strategy against FUS-related neurodegeneration. HIGHLIGHTSO_LITargeted proline additions disrupt {beta}-sheet formation in FUS without altering native conformations, dynamics, or phase separation behavior C_LIO_LI{beta}-sheet-deficient FUS variants prevent aggregation and liquid-to-solid transitions while retaining key biological functions C_LIO_LIIn vivo models reveal attenuated toxicity of {beta}-sheet-deficient FUS in Drosophila C_LIO_LI{beta}-sheets are identified as central drivers of condensate maturation and neuronal death, offering a therapeutic entry point for modulating prion-like domain pathology C_LI

biochemistry↗

bioRxiv2022

CLIP-Seq Analysis Enables the Design of Ribosomal RNA Bait Oligonucleotides That Protect Against C9orf72 ALS/FTD-Associated Poly-GR Pathophysiology

Amyotrophic lateral sclerosis and frontotemporal dementia patients with a hexanucleotide repeat expansion in C9ORF72 (C9-HRE) accumulate poly-GR and poly-PR aggregates. The pathogenicity of these arginine-rich dipeptide repeats (R-DPRs) is thought to be driven by their propensity to bind to low complexity domains of multivalent proteins. However, the ability of R-DPRs to bind native RNA and the significance of this interaction remains unclear. We used computational and experimental approaches to characterize the physicochemical properties of R-DPRs and their interaction with RNA. We find that poly-GR predominantly binds ribosomal RNA (rRNA) in cells and exhibits an interaction that is predicted to be energetically stronger than that for associated ribosomal proteins. Critically, modified rRNA "bait" oligonucleotides restore poly-GR-associated ribosomal deficits in cells and ameliorate poly-GR toxicity in patient neurons and Drosophila models. Our work strengthens the hypothesis that ribosomal function is impaired by R-DPRs, highlights a role for direct rRNA binding in mediating ribosomal disfunction, and presents a strategy for protecting against C9-HRE pathophysiological mechanisms.

neuroscience↗

bioRxiv2022

Axon guidance pathways modulate neurotoxicity of ALS-associated UBQLN2

Mutations in the ubiquitin (Ub) chaperone Ubiquilin 2 (UBQLN2) cause X-linked forms of amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD) through unknown mechanisms. Here we show that aggregation-prone, ALS-associated mutants of UBQLN2 (UBQLN2ALS) trigger heat stress-dependent neurodegeneration in Drosophila. A genetic modifier screen implicated endolysosomal and axon guidance genes, including the netrin receptor, Unc-5, as key modulators of UBQLN2 toxicity. Reduced gene dosage of Unc-5 or its coreceptor Dcc/frazzled diminished neurodegenerative phenotypes, including motor dysfunction, neuromuscular junction defects, and shortened lifespan, in flies expressing UBQLN2ALS alleles. Induced pluripotent stem cells (iPSCs) harboring UBQLN2ALS knockin mutations exhibited lysosomal defects while inducible motor neurons (iMNs) expressing UBQLN2ALS alleles exhibited cytosolic UBQLN2 inclusions, reduced neurite complexity, and growth cone defects that were partially reversed by silencing of UNC5B and DCC. The combined findings suggest that altered growth cone dynamics are a conserved pathomechanism in UBQLN2-associated ALS/FTD.

neuroscience↗

bioRxiv2022

WNK kinases sense molecular crowding and rescue cell volume via phase separation

When challenged by hypertonicity, dehydrated cells must defend their volume to survive. This process requires the phosphorylation-dependent regulation of SLC12 cation chloride transporters by WNK kinases, but how these kinases are activated by cell shrinkage remains unknown. Within seconds of cell exposure to hypertonicity, WNK1 concentrates into membraneless droplets, initiating a phosphorylation-dependent signal that drives net ion influx via the SLC12 cotransporters to rescue volume. The formation of WNK1 condensates is driven by its intrinsically disordered C-terminus, whose evolutionarily conserved signatures are necessary for efficient phase separation and volume recovery. This disorder-encoded phase behavior occurs within physiological constraints and is activated in vivo by molecular crowding rather than changes in cell size. This allows WNK1 to bypass a strengthened ionic milieu that favors kinase inactivity and reclaim cell volume through condensate-mediated signal amplification. Thus, WNK kinases are physiological crowding sensors that phase separate to coordinate a cell volume rescue response.

cell biology↗

bioRxiv2019

Small molecules for modulating protein driven liquid-liquid phase separation in treating neurodegenerative disease

Neurodegenerave diseases such as amyotrophic lateral sclerosis (ALS) are oten associated with mutaons in proteins that are associated with stress granules. Stress granules are condensates formed by liquid-liquid phase separaon which, when aberrant, can lead to altered condensaon behaviours and disease phenotypes. Here, we identified lipoamide, a small molecule which specifically prevents cytoplasmic condensaon of stress granule proteins. Thermal proteome profiling showed that lipoamide preferentially stabilises intrinsically disordered domain-containing proteins. These include SRSF1 and SFPQ, stress granule proteins necessary for lipoamide activity. The redox state of SFPQ correlates with its condensate-dissolving behaviour, in concordance with the importance of the dithiolane ring for lipoamide activity. In animals, lipoamide ameliorates aging-associated aggregaon of a stress granule reporter, improves neuronal morphology, and recovers motor defects caused by expression of ALS-associated FUS and TDP-43 mutants. In conclusion, lipoamide is a well-tolerated small molecule modulator of stress granule condensaon and dissection of its molecular mechanism identified a cellular pathway for redox regulaon of stress granule formaon.

cell biology↗

bioRxiv2017

Alternative cleavage of a bone morphogenetic protein (BMP) produces ligands with distinct developmental functions and receptor preference

TGF-{beta} and Bone Morphogenetic Protein (BMP) family proteins are made as proprotein dimers, which are cleaved by proprotein convertases to release the active C-terminal ligand dimer. Multiple proteolytic processing sites in Glass bottom boat (Gbb), the Drosophila BMP7 ortholog, can produce distinct forms of active ligand. Cleavage at the S1 or atypical S0 site produces Gbb15, the conventional small BMP ligand, while cleavage at the NS site produces the larger Gbb38 ligand (1, 2). Here, we found that blocking NS cleavage increased association of the full length prodomain with Gbb15 resulting in a concomitant decrease in signaling activity. NS cleavage is required in vivo for Gbb-Decapentaplegic (Dpp) heterodimer-mediated wing vein patterning but not in cell culture to enable Gbb15-Dpp het-erodimer activity. Gbb NS cleavage is also required in vivo for the regulation of pupal ecdysis and viability that is dependent on the type II receptor Wishful thinking (Wit). We found that the ability of Gbb38 to signal requires the expression of either Wit or the type I receptor, Saxophone (Sax). Finally, we discovered that the production of Gbb38 in 3rd instar larvae results when processing at the S1/S0 site is blocked by O-linked glycosylation. Our findings demonstrate that BMP prodomain cleavage can ensure that the mature ligand is not inhibited by the prodomain. Furthermore, alternative processing of BMP proproteins produces ligand types that signal preferentially through different receptors and exhibit specific developmental functions.

biochemistry↗