Aberrant phase separation from a rare ABI3 mutation drives microglial dysfunction and Alzheimer's risk

Probably not a substantial claim: may not pass S3 (load-bearing). May combine two claims.

Elevated ABI3 expression has been reported in the brains of patients with AD and in amyloid/tau mouse models.

“Elevated ABI3 expression has been reported in the brains of patients with AD and in amyloid/tau mouse models, 18 , 41”

• cites [18] Ibanez KR, McFarland KN, Phillips J, et al. Deletion of Abi3/Gngt2 influences age‐progressive amyloid β and tau pathologies in distinctive ways. Alzheimers Res Ther. 2022; 14(1): 104. doi: 10.1186/s13195-022-01044-1 PMC9327202 35897046 sourceSupported
• cites [41] Satoh JI, Kino Y, Yanaizu M, et al. Microglia express ABI3 in the brains of Alzheimer's disease and Nasu‐Hakola disease. Intractable Rare Dis Res. 2017; 6(4): 262‐ 268. doi: 10.5582/irdr.2017.01073 PMC5735279 29259854 sourceDisputed

  The Results report ABI3-positive microglial clusters near amyloid plaques in AD brains, but the quantitative analysis found no significant difference in ABI3-immunolabeled area between AD brains and controls. Thus, this paper does not report elevated overall ABI3 expression in AD brains; it also does not report ABI3 expression in amyloid/tau mouse models. (LLM, conf=0.85)

  Checked by a reasoning model — may be wrong.

  Experimental — accuracy not yet measured

Probably not a substantial claim: may not pass S3 (load-bearing).

ABI3 is a component of the N-WASP complex.

“Although ABI3 is a component of the N‐WASP complex, 14”

• cites [14] Kim HG, Berdasco C, Nairn AC, Kim Y. The WAVE complex in developmental and adulthood brain disorders. Exp Mol Med. 2025; 57(1): 13‐ 29. doi: 10.1038/s12276-024-01386-w PMC11799376 39774290 sourceDisputed

  The Introduction distinguishes WASP/N-WASP from WAVE proteins and identifies ABI3 as a subunit of the WAVE complex. It therefore describes a different complex from the one named in the claim. (LLM, conf=0.85)

  Checked by a reasoning model — may be wrong.

  Experimental — accuracy not yet measured

Probably not a substantial claim: may not pass S3 (load-bearing). May combine two claims.

ABI3 is a component of the WAVE2 regulatory complex (WRC2) and has been implicated in regulating actin cytoskeleton dynamics.

“Abelson‐interactor (ABI) family member 3 (ABI3) is a component of the WASP‐family verprolin‐homologous protein 2 (WAVE2 regulatory complex (WRC2) and has been implicated in regulating actin cytoskeleton dynamics. 14”

• cites [14] Kim HG, Berdasco C, Nairn AC, Kim Y. The WAVE complex in developmental and adulthood brain disorders. Exp Mol Med. 2025; 57(1): 13‐ 29. doi: 10.1038/s12276-024-01386-w PMC11799376 39774290 sourceSupported

Probably not a substantial claim: may not pass S3 (load-bearing).

Many LOAD risk-associated genes, such as APOE4, TREM2, and PICALM, have been implicated in regulating microglial immunity and lipid metabolism.

“Many LOAD risk‐associated genes, such as Apolipoprotein E4 (APOE4), 9 , 10 Triggering receptor expressed on myeloid cells 2 (TREM2), 11 and Phosphatidylinositol‑binding clathrin assembly protein (PICALM), 12 have been implicated in regulating microglial immunity and lipid metabolism, 13”

• cites [9] Victor MB, Leary N, Luna X, et al. Lipid accumulation induced by APOE4 impairs microglial surveillance of neuronal‐network activity. Cell Stem Cell. 2022; 29(8): 1197‐ 1212.e8. doi: 10.1016/j.stem.2022.07.005 PMC9623845 35931030 sourcePartly supported

  The Results (page not provided) link APOE4 to altered microglial lipid processing and a pro-inflammatory state, supporting that example. The paper’s reference list mentions a TREM2–APOE pathway, but the passages do not establish the claim for PICALM; the cited paper therefore supports only part of the gene list. (LLM, conf=0.86)

  Experimental — accuracy not yet measured
• cites [10] Haney MS, Pálovics R, Munson CN, et al. APOE4/4 is linked to damaging lipid droplets in Alzheimer's disease microglia. Nature. 2024; 628(8006): 154‐ 161. doi: 10.1038/s41586-024-07185-7 PMC10990924 38480892 sourceSupported
• cites [11] Ulland TK, Song WM, Huang SC, et al. TREM2 maintains microglial metabolic fitness in Alzheimer's disease. Cell. 2017; 170(4): 649‐ 663.e13. doi: 10.1016/j.cell.2017.07.023 PMC5573224 28802038 sourcePartly supported

  The Introduction identifies TREM2 as a late-onset Alzheimer’s risk-associated microglial immune receptor, and the Discussion says it affects microglial metabolism. These passages support the TREM2 portion, but they do not establish the claim about APOE4 and PICALM or specifically show that all three regulate lipid metabolism. (LLM, conf=0.86)

  Experimental — accuracy not yet measured

Classified as probably not a substantial claim.

The IBA1 promoter paired with an artificial miRNA scaffold is a well-established strategy for microglia-specific gene manipulation.

“This promoter, paired with an artificial miRNA scaffold embedded in the miR‐RNA backbone, is a well‐established strategy for microglia‐specific gene manipulation. 33 , 34 , 35”

• cites [33] Serrano C, Cananzi S, Shen T, et al. Simple and highly specific targeting of resident microglia with adeno‐associated virus. iScience. 2024; 27(9): 110706. doi: 10.1016/j.isci.2024.110706 PMC11407971 39297168 sourcePartly supported

  The Results section on miR124T reports that the hIBA1a promoter combined with miR124 target sites produced highly microglia-specific expression. It does not describe an artificial miRNA scaffold embedded in a miR-RNA backbone; the Introduction presents this promoter-based approach as a finding of the study rather than a well-established strategy. Page numbers are unavailable in our copy. (LLM, conf=0.88)

  Experimental — accuracy not yet measured
• cites [34] Okada Y, Hosoi N, Matsuzaki Y, et al. Development of microglia‐targeting adeno‐associated viral vectors as tools to study microglial behavior in vivo. Commun Biol. 2022; 5(1): 1224. doi: 10.1038/s42003-022-04200-3 PMC9652230 36369525 sourcePartly supported

  The Methods (Vector construction; page unavailable) describe pairing the Iba1 promoter with miRNA target sequences, and the Results report microglial targeting with the resulting AAV vector. They do not describe an artificial miRNA scaffold embedded in a miR-RNA backbone. The Discussion also cautions that the Iba1 promoter is not strictly microglia-specific, so the paper does not establish the claimed pairing as a well-established strategy. (LLM, conf=0.88)

  Experimental — accuracy not yet measured
• cites [35] Aoki R, Konno A, Hosoi N, et al. AAV vectors for specific and efficient gene expression in microglia. Cell Rep Methods. 2025; 5(8): 101116. doi: 10.1016/j.crmeth.2025.101116 PMC12461629 40744011 sourcePartly supported

  The Results section (‘The Iba1 promoter is indispensable for transgene expression in microglia’; page not provided) supports the role of the mIba1 promoter in microglia-selective expression. But the Methods describe pairing it with miRNA target sequences, not an artificial miRNA scaffold embedded in a miRNA backbone. This paper therefore does not support the specific pairing claimed. (LLM, conf=0.89)

  Experimental — accuracy not yet measured

Probably not a substantial claim: may not pass S3 (load-bearing).

GWASs have provided accumulating evidence that gene polymorphisms increase the risk of the more common late-onset AD (LOAD).

“genome‐wide association studies (GWASs) have provided accumulating evidence that gene polymorphisms increase the risk of the more common late‐onset AD (LOAD). 6 , 7”

• cites [6] Bellenguez C, Küçükali F, Jansen IE, et al. New insights into the genetic etiology of Alzheimer's disease and related dementias. Nat Genet. 2022; 54(4): 412‐ 436. doi: 10.1038/s41588-022-01024-z PMC9005347 35379992 sourceSupported
• cites [7] Wightman DP, Jansen IE, Savage JE, et al. A genome‐wide association study with 1,126,563 individuals identifies new risk loci for Alzheimer's disease. Nat Genet. 2021; 53(9): 1276‐ 1282. doi: 10.1038/s41588-021-00921-z PMC10243600 34493870 sourceSupported

Probably not a substantial claim: may not pass S3 (load-bearing).

LLPS is often driven by proteins containing intrinsically disordered regions (IDRs), which facilitate multivalent interactions and dynamic condensate assembly.

“LLPS is often driven by proteins containing intrinsically disordered regions (IDRs), which facilitate multivalent interactions and dynamic condensate assembly. 21”

• cites [21] Ahmed R, Forman‐Kay JD. Aberrant phase separation: linking IDR mutations to disease. Cell Res. 2023; 33(8): 583‐ 584. doi: 10.1038/s41422-023-00804-4 PMC10397348 37016021 sourceSupported

Probably not a substantial claim: may not pass S3 (load-bearing).

Mutations in APP and PSEN account for only a small proportion of early-onset familial AD cases.

“Although mutations in amyloid precursor protein (APP) and presenilins (PSEN1, PSEN2) account for only a small proportion of early‐onset familial AD cases, 4 , 5”

• cites [4] Hunter S, Brayne C. Understanding the roles of mutations in the amyloid precursor protein in Alzheimer disease. Mol Psychiatry. 2018; 23(1): 81‐ 93. doi: 10.1038/mp.2017.218 29112196 sourceNot checked — no full text of the source
• cites [5] Kelleher RJ 3rd, Shen J. Presenilin‐1 mutations and Alzheimer's disease. Proc Natl Acad Sci U S A. 2017; 114(4): 629‐ 631. doi: 10.1073/pnas.1619574114 PMC5278466 28082723 sourceCould not verify

  The opening passage (page not available) calls PSEN1 mutations the most common cause of familial Alzheimer’s disease, and a later passage discusses onset ages for people with PSEN or APP mutations. Neither gives the proportion of early-onset familial cases attributable to APP and presenilin mutations. Searches for that proportion found no passage addressing it; 'most common cause' does not establish or refute that the combined proportion is small. (LLM, conf=0.73)

  Experimental — accuracy not yet measured

May combine two claims.

ABI3 is highly abundant in both human AD brains and APP/PS1 mice.

“Consistent with previous scRNA‐seq studies, ABI3 is expressed predominantly in microglia and highly abundant in both human AD brains and APP/PS1 mice. 39 , 40”

• cites [39] Olah M, Menon V, Habib N, et al. Single cell RNA sequencing of human microglia uncovers a subset associated with Alzheimer's disease. Nat Commun. 2020; 11(1): 6129. doi: 10.1038/s41467-020-19737-2 PMC7704703 33257666 sourceCould not verify

  The Results section on AD-associated microglial subsets discusses cluster-defining genes and changes in human AD tissue, but does not identify ABI3 or report its abundance. Searches for ABI3 and APP/PS1 mice found no passage in our copy addressing the claimed expression in either human AD brains or that mouse model. (LLM, conf=0.72)

  Experimental — accuracy not yet measured
• cites [40] Turner AK, Shaw BC, Simpson JF, Estus S. Identification and quantitation of novel ABI3 isoforms relative to Alzheimer's disease genetics and neuropathology. Genes (Basel). 2022; 13(9): 1607. doi: 10.3390/genes13091607 PMC9498898 36140776 sourcePartly supported

  Results §3.2 reports that ABI3 expression is modestly associated with AD neuropathology in human brain samples, while §3.3 reports robust Abi3 expression in APP/PS1 mouse microglia, especially disease-associated microglia. These findings support expression in both settings, but do not establish that ABI3 is highly abundant throughout human AD brains or APP/PS1 mice. (LLM, conf=0.83)

  Experimental — accuracy not yet measured

Probably not a substantial claim: may not pass S3 (load-bearing).

In AD, tau's mutations or abnormal PTMs induce aberrant and excess LLPS, driving fibril formation and neurotoxicity.

“In AD, tau's mutations or abnormal post‐translational modifications (PTMs) induce aberrant and excess LLPS, driving fibril formation and neurotoxicity. 25”

• cites [25] Boyko S, Surewicz WK. Tau liquid‐liquid phase separation in neurodegenerative diseases. Trends Cell Biol. 2022; 32(7): 611‐ 623. doi: 10.1016/j.tcb.2022.01.011 PMC9189016 35181198 sourcePartly supported

  In “The regulatory role of LLPS in tau aggregation” (page unavailable), the review reports that pathogenic mutations accelerate fibril formation within tau droplets. But “The effect of disease-associated mutation on tau LLPS” says the tested mutations do not significantly increase tau’s intrinsic LLPS propensity. The PTM section describes effects that vary by modification, while the cellular-context section suggests a possible toxicity mechanism; neither establishes the claimed mutation/PTM-to-excess-LLPS-to-neurotoxicity pathway in AD. (LLM, conf=0.88)

  Experimental — accuracy not yet measured

Probably not a substantial claim: may not pass S3 (load-bearing).

ABI3 is an important regulator of actin assembly.

“ABI3 is an important regulator of actin assembly 27”

• cites [27] Han KA, Ko J. Orchestration of synaptic functions by WAVE regulatory complex‐mediated actin reorganization. Exp Mol Med. 2023; 55(6): 1065‐ 1075. doi: 10.1038/s12276-023-01004-1 PMC10318009 37258575 sourceSupported

Probably not a substantial claim: may not pass S3 (load-bearing). May combine two claims.

LLPS plays key roles in fundamental biological processes such as cytoskeletal regulation, synaptic signaling, and stress granule formation.

“In addition, LLPS plays key roles in fundamental biological processes such as cytoskeletal regulation, 26 synaptic signaling, 49 and stress granule formation. 50 , 51”

• cites [26] Cheng X, Case LB. Condensates control the actin cytoskeleton. Dev Cell. 2025; 60(11): 1519‐ 1520. doi: 10.1016/j.devcel.2025.04.015 40494278 sourceNot checked — no full text of the source
• cites [49] Chen X, Wu X, Wu H, Zhang M. Phase separation at the synapse. Nat Neurosci. 2020; 23(3): 301‐ 310. doi: 10.1038/s41593-019-0579-9 32015539 sourceNot checked — no full text of the source
• cites [50] Meng F, Li H, Huang Y, et al. RIOK1 phase separation restricts PTEN translation via stress granules activating tumor growth in hepatocellular carcinoma. Nat Cancer. 2025; 6(7): 1223‐ 1241. doi: 10.1038/s43018-025-00984-5 40467995 sourceNot checked — no full text of the source
• cites [51] Yang P, Mathieu C, Kolaitis RM, et al. G3BP1 is a tunable switch that triggers phase separation to assemble stress granules. Cell. 2020; 181(2): 325‐ 345.e28. doi: 10.1016/j.cell.2020.03.046 PMC7448383 32302571 sourceSupported

May combine two claims.

Other studies using germline Abi3 knockout mice have yielded partially conflicting results: one reported increased Aβ aggregation and impaired phagocytosis, whereas another observed reduced Aβ deposition but exacerbated tau pathology, and neither assessed cognitive deficits.

“Other studies using germline Abi3 knockout mice have yielded partially conflicting results: one reported increased Aβ aggregation and impaired phagocytosis, 17 whereas another observed reduced Aβ deposition but exacerbated tau pathology, 18 and neither assessed cognitive deficits.”

• cites [17] Karahan H, Smith DC, Kim B, et al. Deletion of Abi3 gene locus exacerbates neuropathological features of Alzheimer's disease in a mouse model of Aβ amyloidosis. Sci Adv. 2021; 7(45): eabe3954. doi: 10.1126/sciadv.abe3954 PMC8565913 34731000 sourceSupported
• cites [18] Ibanez KR, McFarland KN, Phillips J, et al. Deletion of Abi3/Gngt2 influences age‐progressive amyloid β and tau pathologies in distinctive ways. Alzheimers Res Ther. 2022; 14(1): 104. doi: 10.1186/s13195-022-01044-1 PMC9327202 35897046 sourceSupported

May combine two claims.

The rare ABI3 S209F variant has recently been genetically associated with increased AD risk, with heritability comparable to that of well-known LOAD susceptibility genes such as TREM2.

“Notably, the rare ABI3 S209F variant has recently been genetically associated with increased AD risk, with heritability comparable to that of well‐known LOAD susceptibility genes such as TREM2. 15”

• cites [15] Sims R, van der Lee SJ, et al. Rare coding variants in PLCG2, ABI3, and TREM2 implicate microglial‐mediated innate immunity in Alzheimer's disease. Nat Genet. 2017; 49(9): 1373‐ 1384. doi: 10.1038/ng.3916 PMC5669039 28714976 sourcePartly supported

  The supplied passage labeled Abstract (page unavailable) reports that ABI3 p.S209F is associated with increased late-onset Alzheimer’s disease risk and identifies TREM2 as a known susceptibility gene. It gives association statistics for variants in both genes, but does not establish that their contributions to heritability are comparable. (LLM, conf=0.87)

  Experimental — accuracy not yet measured

Probably not a substantial claim: may not pass S3 (load-bearing).

Classical phase-separating proteins such as FUS, TDP-43, tau, and α-synuclein exhibit abnormal accumulation and aggregation that are closely linked to neuronal dysfunction.

“Classical phase‐separating proteins such as FUS, 46 TDP‐43, 47 tau, 25 , 29 and α‐synuclein 48 exhibit abnormal accumulation and aggregation that are closely linked to neuronal dysfunction.”

• cites [46] Murthy AC, Dignon GL, Kan Y, et al. Molecular interactions underlying liquid‐liquid phase separation of the FUS low‐complexity domain. Nat Struct Mol Biol. 2019; 26(7): 637‐ 648. doi: 10.1038/s41594-019-0250-x PMC6613800 31270472 sourceSupported
• cites [47] Gruijs da Silva LA, Simonetti F, Hutten S, et al. Disease‐linked TDP‐43 hyperphosphorylation suppresses TDP‐43 condensation and aggregation. EMBO J. 2022; 41(8): e108443. doi: 10.15252/embj.2021108443 PMC9016352 35112738 sourceSupported
• cites [25] Boyko S, Surewicz WK. Tau liquid‐liquid phase separation in neurodegenerative diseases. Trends Cell Biol. 2022; 32(7): 611‐ 623. doi: 10.1016/j.tcb.2022.01.011 PMC9189016 35181198 sourceSupported
• cites [29] Wegmann S, Eftekharzadeh B, Tepper K, et al. Tau protein liquid‐liquid phase separation can initiate tau aggregation. EMBO J. 2018; 37(7): e98049. doi: 10.15252/embj.201798049 PMC5881631 29472250 sourceSupported
• cites [48] Ray S, Singh N, Kumar R, et al. α‐Synuclein aggregation nucleates through liquid‐liquid phase separation. Nat Chem. 2020; 12(8): 705‐ 716. doi: 10.1038/s41557-020-0465-9 32514159 sourceSupported

Probably not a substantial claim: may not pass S3 (load-bearing).

Most current LLPS studies have focused on aggregation-prone proteins, which tend to form excessive condensates under pathological conditions and subsequently undergo a liquid-to-solid transition, thereby exacerbating disease progression.

“However, most current LLPS studies have focused on these aggregation‐prone proteins, which tend to form excessive condensates under pathological conditions and subsequently undergo a liquid‐to‐solid transition, thereby exacerbating disease progression. 24”

• cites [24] Zbinden A, Pérez‐Berlanga M, De Rossi P, Polymenidou M. Phase separation and neurodegenerative diseases: a disturbance in the force. Dev Cell. 2020; 55(1): 45‐ 68. doi: 10.1016/j.devcel.2020.09.014 33049211 sourcePartly supported

  The review discusses aggregation-prone proteins and says that loss of condensate dynamics may trigger immobile aggregates (indexed Abstract passages; page unavailable). But it presents the link between LLPS and aggregation as a hypothesis, says a direct link is still missing, and notes that the role of aggregates in disease is debated. It does not establish that most LLPS studies focus on these proteins or that their liquid-to-solid transition generally exacerbates disease progression. (LLM, conf=0.86)

  Experimental — accuracy not yet measured

May combine two claims.

ABI3 and S209F condensates are not associated with membrane-bound organelles and are dissolved by 1,6-HD, confirming LLPS dependence.

“co‐localization studies with organelle markers confirmed that ABI3‐EGFP and ABI3 S209F‐EGFP condensates were not associated with membrane‐bound organelles (Figure S9A–F).”

• Also stated in: § ABI3 S209F mutation impairs its LLPS capacity in live cells · boosted

  “Treatment with 1,6‐hexanediol (1,6‐HD), a disruptor of hydrophobic interactions, induced gradual dissolution of both ABI3 and ABI3 S209F condensates (Figure 4G, Figure S10B), confirming their LLPS dependence.”

May combine two claims.

Primary microglia overexpressing WT ABI3 exhibit strongest migration, whereas S209F shows weakest migration.

“Compared with the control group, primary microglia overexpressing WT ABI3 exhibited the strongest migration capacity, whereas those expressing ABI3 S209F mutant showed the weakest migration ability (Figure 3C,D).”

May combine two claims.

ABI3 deficiency disrupts normal microglial migration, phagocytosis, and clustering around Aβ plaques.

“Here, we found that both ABI3 deficiency and the S209F mutation disrupt normal microglial migration, phagocytosis, and clustering around Aβ plaques.”

May combine two claims.

In Morris water maze, 5×FAD+ABI3 mice show faster acquisition and better probe trial performance, whereas S209F mice show impaired recall.

“During training, 5×FAD+ABI3 mice displayed faster acquisition, with shorter escape latencies and path lengths (Figure 2H, Figure S6G).”

• Also stated in: § ABI3 overexpression improves spatial learning and memory in 5×FAD mice, and this effect is abolished by the S209F mutation · plain

  “In probe trials, these mice showed increased target quadrant occupancy and distance traveled within the target zone compared with 5×FAD+Ctrl. By contrast, 5×FAD+ ABI3 S209F mice exhibited impaired recall, with fewer platform crossings, reduced target quadrant dwell time, and shorter travel distance.”

May combine two claims.

With PEG 8000, ABI3 condensates are more abundant, larger, and more regular than S209F, which forms hydrogel-like structures at high concentrations.

“With PEG 8000, higher protein concentration increased the number and size of ABI3 condensates, whereas the ABI3 S209F variant formed hydrogel‐like structures at high concentrations. ABI3 condensates were more abundant, larger, and more regular in shape compared with those formed by ABI3 S209F (Figure 5B,C, Figure S13).”

The quote only partly states this claim.

WT ABI3 overexpression increases phagocytic capacity compared with S209F in primary microglia and HMC3 cells.

“Twenty‐four hours post‐transfection, cells overexpressing WT ABI3 exhibited significantly higher phagocytic capacity compared with those expressing the S209F variant (Figures 3E,F).”

• Also stated in: § ABI3 overexpression enhances microglial migration and phagocytosis, which are impaired by the S209F mutation · plain

  “Confocal microscopy revealed that ABI3‐overexpressing cells engulfed significantly more beads than controls, whereas ABI3 S209F‐overexpressing cells showed reduced bead uptake versus ABI3 overexpression in HMC3 cells (Figure 3G,H).”

May combine two claims.

ABI3 ΔIDR2-FUS IDR and ABI3 S209F ΔIDR4-FUS IDR showed increased droplet number but with reduced diameter and area compared with ABI3 ΔIDR2 alone.

“By contrast, ABI3 ΔIDR2‐FUS IDR and ABI3 S209F ΔIDR4‐FUS IDR showed increased droplet number but with reduced diameter and area compared with ABI3 ΔIDR2 alone”

May combine two claims.

S209F impairs microglial functions, including migration, phagocytosis, and clustering around Aβ plaques.

“thereby impairing microglial functions.”

• Also stated in: § BACKGROUND · plain

  “Here, we found that both ABI3 deficiency and the S209F mutation disrupt normal microglial migration, phagocytosis, and clustering around Aβ plaques.”

• Also stated in: § RESEARCH IN CONTEXT · plain

  “This defect compromises key microglial functions, including migration, phagocytosis, and clustering around amyloid beta (Aβ) plaques.”

May combine two claims.

S209F abolished the effects of ABI3 on microglial clustering, Aβ clearance, and cognitive decline.

“whereas S209F abolished these effects.”

May combine two claims.

Deletion of IDR2 or SH3 alters condensate properties in WT ABI3, with IDR2 loss most pronounced; in S209F background, IDR4 deletion disrupts droplet formation.

“For WT ABI3, deletion of IDR2 or the SH3 domain significantly altered condensate properties, producing fewer but larger droplets, with IDR2 loss exerting the most pronounced effect. In the S209F background deletion of IDR4 substantially disrupted droplet formation, affecting number, diameter, and area (Figure 6G–J).”

May combine two claims.

Following Aβ stimulation, ABI3 increases wound healing rate and relative wound density, whereas S209F decreases wound closure.

“Following 100 nM Aβ stimulation, the ABI3 group displayed a marked increase in wound healing rate and relative wound density over time compared to vector controls, whereas the ABI3 S209F group showed a significant time‐dependent decrease in wound closure versus ABI3 (Figure S7A,B).”

May combine two claims. The quote only partly states this claim.

ABI3 depletion in APP/PS1 mice aggravates amyloid pathology and impairs microglial recruitment and morphology.

“At 12 months of age, ABI3 depletion markedly aggravated amyloid pathology in APP/PS1 mice.”

• Also stated in: § Microglia‐specific ABI3 knockdown exacerbates amyloid pathology and impairs microglial migration and phagocytosis · plain

  “Confocal imaging revealed increased Aβ plaque number, larger mean plaque diameter, and greater plaque area in AD+ABI3 KD mice compared to AD+Ctrl mice (Figure S3C,D).”

• Also stated in: § Microglia‐specific ABI3 knockdown exacerbates amyloid pathology and impairs microglial migration and phagocytosis · boosted

  “Morphometric analyses demonstrated profound defects in plaque‐associated microglia following ABI3 knockdown.”

May combine two claims. The quote only partly states this claim.

With Ficoll 400, ABI3 maintains droplets, whereas S209F transitions to rosary-like assemblies and fibrous structures.

“Using Ficoll 400 as an alternative crowding agent, ABI3 maintained droplet structures with increasing concentration, whereas ABI3 S209F transitioned to rosary‐like assemblies (Figure S14).”

• Also stated in: § ABI3 S209F mutation impairs its LLPS capacity in vitro · plain

  “Notably, ABI3 S209F developed irregular hydrogel networks at high concentrations and underwent transitions from droplets to fibrous structures over time (Figure 5F, Figure S14–S15).”

May combine two claims. The quote only partly states this claim.

Endogenous ABI3 forms cytoplasmic droplet-like condensates in HEK293T, HMC3 cells, and mouse hippocampal tissue.

“We observed that endogenous ABI3 formed distinct cytoplasmic droplet‐like condensates in both HEK293T and HMC3 cells (Figure 4A).”

• Also stated in: § ABI3 S209F mutation impairs its LLPS capacity in live cells · plain

  “Furthermore, similar droplet‐like condensates of ABI3 were also detected in the hippocampal region of mouse brain tissue (Figure 4B).”

May combine two claims.

25 μM ABI3 or S209F induces turbidification, and turbidity changes are time- and concentration-dependent.

“we observed that 25 µM ABI3 or ABI3 S209F induced turbidification, a hallmark of LLPS (Figure S12C).”

• Also stated in: § ABI3 S209F mutation impairs its LLPS capacity in vitro · plain

  “Turbidity assays at λ = 600 nm confirmed that these changes occurred in both time‐ and concentration‐dependent manners (Figure S12D).”

May combine two claims.

ABI3 overexpression enhances wound closure in HMC3 cells, whereas S209F reduces wound healing rates.

“The ABI3 overexpression group showed a time‐dependent enhancement of wound closure compared to vector controls, whereas the ABI3 S209F overexpression group exhibited significantly reduced wound healing rates versus ABI3 overexpression (Figure 3B).”

May combine two claims.

ABI3 S209F expression fails to attenuate Aβ plaque deposition and impairs microglial migration in 5×FAD mice.

“In contrast, expression of ABI3 S209F failed to attenuate Aβ plaque deposition in 5×FAD mice (Figure 2B,C).”

• Also stated in: § ABI3 overexpression mitigates amyloid pathology in 5×FAD mice, and this effect is abolished by the S209F mutation · plain

  “whereas ABI3 S209F overexpression impaired this microglial migration (Figure 2D,E).”

May combine two claims.

Molecular dynamics modeling shows S209A similar to S209F, while S209D increases polarity and interacts with Arg201.

“Molecular dynamics modeling showed that the S209A variant exhibits RMSD fluctuations and hydrophobic interactions comparable to those of S209F. In contrast, the S209D mutant increases side‐chain polarity and interacts with the nearby positively charged Arg201, forming a stable electrostatic structure (Figure S11).”

May combine two claims.

FRAP shows both proteins maintain liquid-like properties, but S209F recovers faster.

“Consistent with cellular observations, FRAP analysis demonstrated that both proteins maintain liquid‐like properties, but ABI3 S209F condensates recovered fluorescence more rapidly than ABI3, suggesting altered molecular dynamics in vitro (Figure 5D,E).”

May combine two claims.

WT ABI3 contains IDR1 (89-132) and IDR2 (161-303); S209F splits IDR2 into IDR3 (161-200) and IDR4 (207-303).

“PONDR analysis revealed that WT ABI3 contains two IDRs: IDR1 (89‐132) and IDR2 (161‐303), and the S209F mutation splits IDR2 (161‐303) into two discrete regions: IDR3 (161‐200) and IDR4 (207‐303) (Figure 6A).”

May combine two claims.

ABI3 overexpression rescues Y-maze and novel arm defects in 5×FAD mice, whereas S209F worsens them.

“ABI3 overexpression partially rescued this defect, whereas ABI3 S209F further decreased alternation rate (Figure 2F).”

• Also stated in: § ABI3 overexpression improves spatial learning and memory in 5×FAD mice, and this effect is abolished by the S209F mutation · plain

  “ABI3 overexpression enhanced novel arm exploration and reduced familiar arm preference, whereas ABI3 S209F produced the opposite effect (Figure 2G).”

May combine two claims.

Both ABI3 and S209F condensates show liquid-like behavior (movement, fusion, fission).

“Live‐cell imaging revealed rapid movement, fusion (Figure 4J), and fission (Figure 4K) of both ABI3 and ABI3 S209F condensates, further supporting their liquid‐like behavior.”

May combine two claims.

5×FAD+Ctrl mice exhibit impaired working memory in Y-maze and novel arm exploration.

“In the Y‐maze, 5×FAD+Ctrl mice exhibited reduced spontaneous alternation compared with WT+Ctrl, consistent with impaired working memory.”

• Also stated in: § ABI3 overexpression improves spatial learning and memory in 5×FAD mice, and this effect is abolished by the S209F mutation · plain

  “In the novel arm exploration test, 5×FAD+Ctrl mice showed fewer novel arm entries and lower exploration percentage relative to WT+Ctrl.”

May combine two claims.

S209A increases droplet number but reduces diameter/area, whereas S209D decreases number and promotes larger condensates.

“Confocal imaging revealed that the S209A mutation increased droplet number but reduced condensate diameter and area, whereas the S209D mutation decreased droplet number while promoting larger condensates (Figure 6C–E).”

May combine two claims. The quote only partly states this claim.

No group differences were observed in open field, novel object recognition, or swimming speed.

“In the open field test, no group differences were observed in locomotor activity or anxiety‐related parameters, indicating preserved basal motor function (Figure S6A–D).”

• Also stated in: § ABI3 overexpression improves spatial learning and memory in 5×FAD mice, and this effect is abolished by the S209F mutation · plain

  “In novel object recognition, no significant difference was observed in the percentage of recognition index (Figure S6E).”

• Also stated in: § ABI3 overexpression improves spatial learning and memory in 5×FAD mice, and this effect is abolished by the S209F mutation · plain

  “In the Morris water maze, swimming speed was comparable across all groups (Figure S6F).”

May combine two claims.

ABI3 overexpression in 5×FAD mice reduces Aβ burden and enhances microglial recruitment to plaques.

“Confocal imaging of Aβ‐stained hippocampal sections revealed that ABI3 overexpression reduced Aβ burden, with significantly fewer plaque numbers, shorter mean diameter, and smaller plaque area compared with 5×FAD controls.”

• Also stated in: § ABI3 overexpression mitigates amyloid pathology in 5×FAD mice, and this effect is abolished by the S209F mutation · boosted

  “Notably, quantification of Aβ‐associated microglia clustering demonstrated enhanced microglial recruitment of Iba1+ cells to plaques in 5×FAD+ABI3 mice, whereas ABI3 S209F overexpression impaired this microglial migration (Figure 2D,E).”

May combine two claims.

At 15 μM with PEG 8000, both ABI3 and S209F condensates show rapid movement, fusion, and fission.

“Moreover, at 15 µM protein incubation with PEG 8000, both ABI3 and ABI3 S209F condensates showed rapid movement, fusion (Figure S12F), and fission (Figure S12G), further supporting their liquid‐like behavior in vitro.”

May combine two claims.

Treatment with 1,6-HD phenocopied the S209F mutation, significantly impairing both migration and phagocytosis of red fluorescent latex beads to levels comparable to those observed in S209F-expressing cells.

“Treatment with 1,6‐HD phenocopied the S209F mutation, significantly impairing both migration (HMC3: Figure S20A–S21B; BV2: Figure S22A–S23B) and phagocytosis of red fluorescent latex beads (HMC3: Figure S20C–21D; BV2: Figure S22C–23D) to levels comparable to those observed in S209F‐expressing cells.”

May combine two claims.

Blue light stimulation induced robust increases in both phagocytosis and migration selectively in cells expressing the optoABI3 IDR construct, whereas control cells showed no response.

“Blue light stimulation induced robust increases in both phagocytosis (HMC3: Figures S26A,B; BV2: Figures S27A,B) and migration (HMC3: Figures S26C–E; BV2: Figures S27C–E) selectively in cells expressing the optoABI3 IDR construct, whereas control cells showed no response.”

May combine two claims.

Fusing the FUS-IDR to ABI3 S209F effectively rescues condensate formation as well as microglial migration and phagocytosis.

“Notably, fusing the FUS‐IDR to ABI3 S209F effectively rescues condensate formation as well as microglial migration and phagocytosis”

May combine two claims.

No altered body weight or metabolic gene expression was detected.

“No altered body weight or metabolic gene expression was detected”

May combine two claims.

The S209F mutant protein was expressed at levels comparable to that of WT, ruling out a simple loss-of-expression mechanism.

“As shown in Figure S28, the mutant protein was expressed at levels comparable to that of WT, ruling out a simple loss‐of‐expression mechanism.”

May combine two claims.

Microglial ABI3 overexpression mitigates amyloid pathology by promoting microglial recruitment to plaques under AD pathology, whereas the AD-associated ABI3 S209F variant abolishes these protective effects.

“Altogether, these findings revealed that microglial ABI3 overexpression mitigates amyloid pathology by promoting microglial recruitment to plaques under AD pathology, whereas the AD‐associated ABI3 S209F variant abolished these protective effects, failing to alleviate plaque burden or restore microglial responses.”

• AssumptionRests on an assumption: The ABI3 S209F variant is associated with Alzheimer's disease.

Probably not a substantial claim: may not pass S3 (load-bearing).

FRAP experiments were repeated five times.

“The FRAP experiment was repeated five times.”

• Also stated in: § FRAP assay in HEK293T cells · plain

  “The experiment was repeated five times, and GraphPad Prism 10.1.2 was used to analyze the data and plot FRAP curves.”

Classified as probably not a substantial claim. May combine two claims.

For all in vivo experiments, the mouse served as the biological replicate; individual cells or fields were not treated as independent replicates.

“For all in vivo experiments, the mouse served as the biological replicate. For each mouse, 5–6 coronal sections (20 µm) spanning the hippocampus were selected at regular intervals (every 240 µm). Within each section, 3–5 fields were acquired in hippocampal subregions. Values were averaged across all fields and sections to generate a single per‐mouse average, which was used for statistical comparis”

Probably not a substantial claim: may not pass S3 (load-bearing).

ABI3 mean fluorescence intensity was quantified by two blinded observers.

“ABI3 mean fluorescence intensity was quantified in Iba1+ (microglia), NeuN+ (neurons), and GFAP+ (astrocytes) cells with DAPI+ nuclei by two blinded observers.”

Classified as probably not a substantial claim.

Purity of primary microglia was assessed by immunostaining for Iba1 and was consistently >95%.

“Purity was assessed by immunostaining for Iba1 and was consistently >95%.”

May combine two claims.

Microglial-specific ABI3 depletion exacerbates amyloid plaque burden and disrupts microglial recruitment and phagocytic capacity, highlighting a critical role for ABI3 in maintaining microglial response to Aβ pathology.

“Together, these findings demonstrated that microglial‐specific ABI3 depletion exacerbates amyloid plaque burden and disrupts microglial recruitment and phagocytic capacity, highlighting a critical role for ABI3 in maintaining microglial response to Aβ pathology.”

Classified as probably not a substantial claim. May combine two claims.

Phase separation regulates the cytoskeleton, which is closely linked to cell migration.

“Because phase separation has been reported to regulate the cytoskeleton, 26 which is closely linked to cell migration”

• cites [26] Cheng X, Case LB. Condensates control the actin cytoskeleton. Dev Cell. 2025; 60(11): 1519‐ 1520. doi: 10.1016/j.devcel.2025.04.015 40494278 sourceNot checked — no full text of the source

Probably not a substantial claim: may not pass S3 (load-bearing). May combine two claims.

Consistent microscope settings ensured reliable results, and a 360-4095 gray threshold removed background noise.

“Consistent microscope settings ensured reliable results, and a 360‐4095 gray threshold removed background noise.”

May combine two claims.

ABI3 undergoes phase separation and S209F perturbs condensate formation and dynamics, providing a mechanistic link to AD pathogenesis.

“Altogether, these findings establish that ABI3 undergoes phase separation and demonstrate that the S209F mutation perturbs condensate formation and dynamics, providing a mechanistic link to its role in AD pathogenesis.”

• AssumptionRests on an assumption: The ABI3 S209F variant is associated with Alzheimer's disease.

Probably not a substantial claim: may not pass S3 (load-bearing).

All animal experiments were approved by the Ethics Committee for Experimental Animals of the Affiliated Hospital of Guangdong Medical University.

“All animal experiments described in this article were approved by the Ethics Committee for Experimental Animals of the Affiliated Hospital of Guangdong Medical University (License number: SYXK (Guangdong) 2022‐0286).”

May combine two claims.

The AD-associated ABI3 S209F variant drives AD pathogenesis by disrupting LLPS, causing microglial dysfunction.

“The AD‐associated ABI3 S209F variant drives pathogenesis by disrupting LLPS, causing microglial dysfunction.”

Probably not a substantial claim: may not pass S3 (load-bearing).

Many proteins involved in actin cytoskeletal dynamics undergo LLPS to concentrate effectors and facilitate signal transduction.

“Notably, many proteins involved in actin cytoskeletal dynamics undergo LLPS to concentrate effectors and facilitate signal transduction. 26”

• cites [26] Cheng X, Case LB. Condensates control the actin cytoskeleton. Dev Cell. 2025; 60(11): 1519‐ 1520. doi: 10.1016/j.devcel.2025.04.015 40494278 sourceNot checked — no full text of the source

May combine two claims.

ABI3 overexpression improves spatial learning and memory in 5×FAD mice, whereas S209F fails to improve cognitive dysfunction.

“Together, these results indicated that microglial ABI3 overexpression ameliorates spatial learning and memory defects in 5×FAD mice, whereas the S209F variant fails to improve cognitive dysfunction.”

Probably not a substantial claim: may not pass S3 (load-bearing). May combine two claims.

All procedures followed NIH guidelines and were approved by the Affiliated Hospital of Guangdong Medical University Animal Care Committee.

“All procedures followed National Institutes of Health (NIH) guidelines and were approved by the Affiliated Hospital of Guangdong Medical University Animal Care Committee.”

Classified as probably not a substantial claim.

LLPS regulates diverse processes in health and disease by dynamically compartmentalizing proteins and nucleic acids without membranes.

“By dynamically compartmentalizing proteins and nucleic acids without membranes, LLPS regulates diverse processes in health and disease. 22 , 23”

• cites [22] Shimobayashi SF, Ronceray P, Sanders DW, Haataja MP, Brangwynne CP. Nucleation landscape of biomolecular condensates. Nature. 2021; 599(7885): 503‐ 506. doi: 10.1038/s41586-021-03905-5 34552246 sourceNot checked — no full text of the source
• cites [23] Xie F, Zhou X, Ran Y, et al. Targeting FOXM1 condensates reduces breast tumour growth and metastasis. Nature. 2025; 638(8052): 1112‐ 1121. doi: 10.1038/s41586-024-08421-w 39814884 sourceNot checked — no full text of the source

May combine two claims.

ABI3 possesses stronger intrinsic LLPS capacity, whereas S209F disrupts condensate formation and promotes aberrant phase transitions.

“Collectively, these in vitro results demonstrated that ABI3 possesses stronger intrinsic LLPS capacity, whereas the S209F mutation disrupts condensate formation and phase separation dynamics, promoting aberrant phase transitions under diverse crowding and ionic environments.”

May combine two claims.

Microglia-specific ABI3 knockdown was efficient and selective.

“Immunofluorescence showed selective ABI3 reduction in microglia, but not in neurons or astrocytes, with regional and single‐cell confirmation (Figure S2C–E).”

• Also stated in: § Microglia‐specific ABI3 knockdown exacerbates amyloid pathology and impairs microglial migration and phagocytosis · boosted

  “Hippocampal mRNA and protein levels were also reduced (Figure S3A,B), collectively demonstrating efficient microglia‐specific ABI3 KD.”

May combine two claims.

ABI3 promotes phagocytosis and migration in HMC3 cells, whereas S209F impairs these functions, highlighting its role in dysregulated microglial responses in AD.

“These results confirm that ABI3 promotes both phagocytosis and migration in HMC3 cells, whereas the ABI3 S209F mutation impairs these functions, highlighting its role in dysregulated microglial responses in AD.”

• AssumptionRests on an assumption: The ABI3 S209F variant is associated with Alzheimer's disease.

Classified as probably not a substantial claim.

Biomolecular condensates formed through LLPS have emerged as critical organizers of cellular biochemistry.

“Biomolecular condensates formed through liquid–liquid phase separation (LLPS) have emerged as critical organizers of cellular biochemistry.”

Classified as probably not a substantial claim.

Microglia are the resident macrophages of the central nervous system that exert protective function by phagocytosing and clearing pathological protein aggregates.

“Microglia are the resident macrophages of the central nervous system that exert protective function by phagocytosing and clearing pathological protein aggregates. 8”

• cites [8] Leng F, Edison P. Neuroinflammation and microglial activation in Alzheimer disease: where do we go from here?. Nat Rev Neurol. 2021; 17(3): 157‐ 172. doi: 10.1038/s41582-020-00435-y 33318676 sourceNot checked — no full text of the source

May combine two claims.

Our AAV-mediated, hippocampus-specific microglial ABI3 knockdown avoids global gene deletion and hypothalamic impairment.

“Our AAV‐mediated, hippocampus‐specific microglial ABI3 knockdown avoids global gene deletion and hypothalamic impairment.”

May combine two claims.

The S209F mutation impairs microglial migration and neuroprotective function by disrupting ABI3 LLPS.

“Therefore, the S209F mutation impairs microglial migration and neuroprotective function by disrupting ABI3 LLPS.”

Probably not a substantial claim: may not pass S3 (load-bearing).

The exclusive use of male mice constitutes a limitation of this study, limiting the generalizability of our conclusions to female animals and human populations.

“We acknowledge that the exclusive use of male mice constitutes a limitation of this study, limiting the generalizability of our conclusions to female animals and human populations.”

May combine two claims.

ABI3 IDR2 improves migration and phagocytosis.

“demonstrating that the ABI3 IDR2 improves migration and phagocytosis.”

May combine two claims.

The S209F variant exerts its pathogenic effects through altered phosphorylation and disrupted LLPS.

“our data further suggest that the S209F variant exerts its pathogenic effects through altered phosphorylation and disrupted LLPS”

Probably not a substantial claim: may not pass S3 (load-bearing).

We were unable to identify an anti-ABI3 antibody that produces specific and reliable signal for flow cytometric detection of endogenous mouse Abi3.

“despite extensive testing of multiple commercially available anti‐ABI3 antibodies, we were unable to identify one that produces specific and reliable signal for flow cytometric detection of endogenous mouse Abi3.”

Probably not a substantial claim: may not pass S3 (load-bearing).

These discrepancies may arise from differences in models, sex, or pathological background.

“These discrepancies may arise from differences in models, sex, or pathological background.”

May combine two claims.

ABI3 is a non-aggregative phase-separating protein in AD neuroprotection.

“reveals the unique role of ABI3 as a non‐aggregative phase‐separating protein in AD neuroprotection.”