bims-axbals 2025-07-20 papers

bims-axbals

Biomed News

on Axonal biology and ALS

Issue of 2025–07–20
39 papers selected by
TJ Krzystek

Therapeutic Approaches for C9ORF72-Related ALS: Current Strategies and Future Horizons.
Derivation and Culture of Enriched Phrenic-Like Motor Neurons From Human iPSCs.
Long-read RNA sequencing unveils a novel cryptic exon in MNAT1 along with its full-length transcript structure in TDP-43 proteinopathy.
UBQLN2 in neurodegenerative disease: mechanistic insights and emerging therapeutic potential.
Human TDP-43 overexpression in zebrafish motor neurons triggers MND-like phenotypes through gain-of-function mechanism.
DCPS modulates TDP-43 mediated neurodegeneration through P-body regulation.
Inhibition of nonsense-mediated decay in TDP-43 deficient neurons reveals novel cryptic exons.
TDP-43 promotes efficient HSV-1 replication in human DRG-derived neurons.
TDP-43 toxic gain of function links ALS, FTD and Alzheimer's Disease through splicing dysregulation.
Emerging biomarkers in amyotrophic lateral sclerosis: from pathogenesis to clinical applications.
Nonsense-mediated decay masks cryptic splicing events caused by TDP-43 loss.
JIP4 deficiency causes a lysosomal storage disease arising from impaired cystine efflux.
LRRK2 kinase modulates glucose-stimulated insulin secretion via RAB8 phosphorylation and ciliogenesis.
Accumulation of LRRK2-associated phospho-Rab12 degenerative lysosomes in tauopathies.
Transcriptional modulation unique to vulnerable motor neurons predicts ALS across species and SOD1 mutations.
Live Cell Imaging to Monitor Axonal Pruning in Drosophila Motor Neurons.
Autophagy activators normalize aberrant Tau proteostasis and rescue synapses in human familial Alzheimer's disease iPSC-derived cortical organoids.
Downregulation of NEAT1 due to loss of TDP-43 function exacerbates motor neuron degeneration in amyotrophic lateral sclerosis.
Loss of the lysosomal lipid flippase ATP10B leads to progressive dopaminergic neurodegeneration and parkinsonian motor deficits.
Cytoskeletal scaffolding of NaVs and KVs in neocortical pyramidal neurons: Implications for neuronal signaling and plasticity.
KIF5A downregulation in spinal muscular atrophy links axonal regeneration defects with ALS.
A guide to selecting high-performing antibodies for Stearoyl-CoA desaturase (SCD1) (UniProt ID: O00767) for use in western blot, immunoprecipitation, and immunofluorescence.
Cholinergic neuron circadian clock mediates RNA-binding protein function and contributes to ALS disease phenotypes.
Neuronal branching in stem cell models of mitochondrial and neurological diseases.
Potassium channel clustering: mechanisms shaping axonal excitability.
The Role of Mitochondrial Dynamics in Doxorubicin-Induced Disease: Implications for Therapy.
NEDD4L induces mitochondrial dysfunction and neurodegeneration by promoting LIPT2 degradation in Huntington's disease.
The Ubiquitin E3 Ligase UBE3A Regulates GRIPAP1 and PACSIN1 Proteins Linked to the Endocytic Recycling of AMPA Receptors.
Novel Dual Mechanism GRT-X Agonist Acting on Kv7 Potassium Channel/Translocator Protein Receptor Prevents Motoneuron Degeneration Following Exposure to Mouse and Human Amyotrophic Lateral Sclerosis/Frontotemporal Dementia Astrocyte-Conditioned Media.
Modeling human retinal ganglion cell axonal outgrowth, development, and pathology using pluripotent stem cell-based microfluidic platforms.
Expanding the spectrum of annexin A11 proteinopathy in frontotemporal lobar degeneration and motor neuron disease.
The Rab3 family proteins in age-related neurodegeneration: unraveling molecular pathways and potential therapeutic targets.
GSK-3β coordinates axonal microtubule organisation through Shot and Tau.
Electroporation-Based Gene Delivery and Whole-Organoid Imaging in Human Retinal Organoids.
Low-dose IL-2 reinvigorates the immunoguardians of neurodegenerative diseases.
Enhanced axonal transport in large vertebrates: KIF5A adaptations in giraffes and pythons.
Treatment of age-related decreases in GTP levels restores endocytosis and autophagy.
Tau endo-lysosomal processing in human iPSC-derived microglia is impacted by tau aggregation state, but not by microglial activation status.
Mitochondria from huntington´s disease striatal astrocytes are hypermetabolic and compromise neuronal branching.

Int J Mol Sci. 2025 Jun 28. pii: 6268. [Epub ahead of print]26(13):

Therapeutic Approaches for C9ORF72-Related ALS: Current Strategies and Future Horizons.

Marco Cattaneo, Eleonora Giagnorio, Giuseppe Lauria, Stefania Marcuzzo.

  Amyotrophic lateral sclerosis (ALS) is a fatal neurodegenerative disease characterized by the loss of upper and lower motor neurons. One of its major genetic causes is C9ORF72, where mutations lead to hexanucleotide repeat expansions in the C9ORF72 gene. These expansions drive disease progression through mechanisms, including the formation of toxic RNAs and the accumulation of damaged proteins such as dipeptide repeats (DPRs). This review highlights these pathogenic mechanisms, focusing on RNA foci formation and the accumulation of toxic DPRs, which contribute to neuronal damage. It also discusses promising targeted therapies, including small molecules and biological drugs, designed to counteract these specific molecular events. Small molecules such as G-quadruplex stabilizers, proteasome and autophagy modulators, and RNase-targeting chimeras show potential in reducing RNA foci and DPR accumulation. Furthermore, targeting enzymes involved in repeat-associated non-AUG (RAN) translation and nucleocytoplasmic transport, which are crucial for disease pathogenesis, opens new therapeutic avenues. Even some anti-viral drugs show encouraging results in preclinical studies. Biological drugs, such as antisense oligonucleotides and gene-editing technologies like CRISPR-Cas, were explored for their potential to specifically target C9ORF72 mutations and modify the disease's molecular foundations. While preclinical and early clinical data show promise, challenges remain in optimizing delivery methods, ensuring long-term safety, and improving efficacy. This review concludes by emphasizing the importance of continued research and the potential for these therapies to alter the disease trajectory and improve patient outcomes.

Keywords:  C9ORF72; amyotrophic lateral sclerosis; biological drugs; small molecules; therapeutic strategies

DOI:  https://doi.org/10.3390/ijms26136268
Bio Protoc. 2025 Jul 05. 15(13): e5363

Derivation and Culture of Enriched Phrenic-Like Motor Neurons From Human iPSCs.

Louise Thiry, Julien Sirois, Thomas M Durcan, Stefano Stifani.

  The fatal motor neuron (MN) disease amyotrophic lateral sclerosis (ALS) is characterized by progressive degeneration of the phrenic MNs (phMNs) controlling the activity of the diaphragm, leading to death by respiratory failure. Human experimental models to study phMNs are lacking, hindering the understanding of the mechanisms of phMN degeneration in ALS. Here, we describe a protocol to derive phrenic-like MNs from human induced pluripotent stem cells (hiPSC-phMNs) within 30 days. During spinal cord development, phMNs emerge from specific MN progenitors located in the dorsalmost MN progenitor (pMN) domain at cervical levels, under the control of a ventral-to-dorsal gradient of Sonic hedgehog (SHH) signaling and a rostro-caudal gradient of retinoic acid (RA). The method presented here uses optimized concentrations of RA and the SHH agonist purmorphamine, followed by fluorescence-activated cell sorting (FACS) of the resulting MN progenitor cells (MNPCs) based on a cell-surface protein (IGDCC3) enriched in hiPSC-phMNs. The resulting cultures are highly enriched in MNs expressing typical phMN markers. This protocol enables the generation of hiPSC-phMNs and is highly reproducible using several hiPSC lines, offering a disease-relevant system to study mechanisms of respiratory MN dysfunction. While the protocol has been validated in the context of ALS research, it can be adopted to study human phrenic MNs in other research fields where these neurons are of interest. Key features • This protocol generates enriched hiPSC-derived phrenic motor neuron cultures. • The protocol can be used to develop models to study human respiratory motor neuron disease. • The protocol allows the generation of phrenic motor neuron preparations with potential for motor neuron replacement strategies. • The protocol requires experience in hiPSC culturing and FACS-based cell sorting for a successful outcome.

Keywords:  ALS; FACS-based cell sorting; Human iPSC; Motor neuron disease modeling; Motor neuron replacement therapy; Phrenic motor neurons; Respiratory motor neurons; iPSC-derived motor neurons

DOI:  https://doi.org/10.21769/BioProtoc.5363
Commun Biol. 2025 Jul 16. 8(1): 1056

Long-read RNA sequencing unveils a novel cryptic exon in MNAT1 along with its full-length transcript structure in TDP-43 proteinopathy.

Yoshihisa Tanaka, Naohiro Sunamura, Rei Kajitani, Marie Ikeguchi, Ryo Kunimoto.

Understanding the role of transcript isoforms is essential for elucidating disease mechanisms. TDP-43 regulates RNA splicing, and its dysfunction in neurons is a hallmark of some neurodegenerative diseases, including amyotrophic lateral sclerosis (ALS) and frontotemporal degeneration (FTD). While an association between TDP-43-dependent cryptic exons and disease pathogenesis has been suggested, an approach to investigate how cryptic exons disrupt transcript isoforms has yet to be established. In this study, we developed IsoRefiner, a novel method for identifying full-length transcript structures using long-read RNA-seq. Leveraging this method, we performed long-read RNA-seq, guided by prior short-read RNA-seq, to comprehensively determine the full-length structures of aberrant transcripts due to TDP-43 dysregulation in human iPSC-derived motor neurons. We identified a novel TDP-43-dependent cryptic exon in the MNAT1 gene, along with its full-length transcript structure. Furthermore, we confirmed the presence of the MNAT1 cryptic exon in patients with ALS and FTD. Our findings deepen understanding of TDP-43 proteinopathy and advance splicing research.

DOI: https://doi.org/10.1038/s42003-025-08463-4
Biochem Soc Trans. 2025 Jul 14. pii: BST20253053. [Epub ahead of print]

UBQLN2 in neurodegenerative disease: mechanistic insights and emerging therapeutic potential.

Autumn M Matthews, Alexandra M Whiteley.

  Ubiquilins (UBQLNs) regulate cellular protein turnover by shuttling proteins, or 'clients', to the proteasome or autophagy pathways for degradation. Of the five different UBQLN genes in humans, UBQLN2 is the most highly expressed in the nervous system and muscle tissue and has been linked to multiple neurodegenerative diseases. In particular, point mutations of UBQLN2 cause an X-linked, dominant form of amyotrophic lateral sclerosis (ALS), ALS with frontotemporal dementia (ALS/FTD), or FTD. Failed protein degradation is a hallmark of many neurodegenerative diseases, including ALS and FTD; however, it is not clear exactly how ALS/FTD-associated UBQLN2 mutations contribute to pathogenesis. Recent studies have revealed the complexity of UBQLN2 biology and allow deeper understanding as to how UBQLN2 dysfunction may contribute to neurodegenerative disease. UBQLN2 is necessary for mitochondrial protein degradation and for regulating mitochondrial turnover, both of which are essential for motor neurons and have been implicated in the pathogenesis of ALS. Stress granule (SG) formation and regulation are also affected by UBQLN2 mutations, and their dysregulation may contribute to the toxic protein aggregation and SG changes observed in neurodegenerative disease. Finally, there are compelling links connecting UBQLN2 dysfunction with changes to downstream neuronal morphology, function, and behavior. This review will detail the emerging consensus on how UBQLN2 protects against neurodegenerative disease and will provide insights into potential therapeutic approaches.

Keywords:  ALS; PEG10; Ubiquilin 2; mitochondria; neurodegenerative disease; protein degradation; stress granules

DOI:  https://doi.org/10.1042/BST20253053
bioRxiv. 2025 Jul 07. pii: 2025.07.06.663393. [Epub ahead of print]

Human TDP-43 overexpression in zebrafish motor neurons triggers MND-like phenotypes through gain-of-function mechanism.

Alison L Hogan, Madison Kane, Patrick Chiu, Grant Richter, Cindy Maurel, Sharlynn Wu, Natalie M Scherer, Emily K Don, Albert Lee, Ian Blair, Roger Chung, Marco Morsch.

Dysregulation of the TAR DNA-binding protein 43 (TDP-43), including intraneuronal cytoplasmic mislocalisation and aggregation is a feature of multiple neurodegenerative diseases including amyotrophic lateral sclerosis (ALS), frontotemporal lobar dementia (FTLD), limbic-predominant age-related TDP-43 encephalopathy (LATE) and alzheimers disease (AD). Unravelling the causes and functional consequences of TDP-43 dysregulation is paramount to understanding disease mechanisms as well as identifying effective therapeutic targets. Here we present a comprehensive in vivo characterisation of three stable transgenic zebrafish models that express human TDP-43 variants in motor neurons. We demonstrate that overexpression of predominantly nuclear wildtype TDP-43, cytoplasm-targeted TDP-43, and an ALS-linked variant (G294V) each induce toxic gain-of-function effects, leading to impaired motor function, motor neuron loss, and muscle atrophy. Importantly, these models reveal distinct phenotypes, with the ALS-linked mutant exhibiting axonal transport deficits and neuromuscular junction disruption, while cytoplasmic mislocalised TDP-43 heightened susceptibility to oxidative stress. Two FDA-approved drugs used to treat ALS, edaravone and riluzole, were examined in these models and revealed that edaravone, but not riluzole, was effective in rescuing motor deficits associated with cytoplasmic TDP-43 expression and, to a lesser extent, mutant TDP-43 G294V . Collectively, these findings reveal distinct pathological consequences of TDP-43 dysregulation, providing neuron-centric mechanistic insights, and establish the humanised TDP-43 zebrafish as an efficient system for preclinical therapeutic testing.
Graphical abstract:

DOI: https://doi.org/10.1101/2025.07.06.663393
bioRxiv. 2025 Jun 15. pii: 2025.06.13.659508. [Epub ahead of print]

DCPS modulates TDP-43 mediated neurodegeneration through P-body regulation.

Yingzhi Ye, Zhe Zhang, Yu Xiao, Chengzhang Zhu, Noelle Wright, Julie Asbury, Yongxin Huang, Weiren Wang, Laura Gomez-Isaza, Juan C Troncoso, Chuan He, Shuying Sun.

The proteinopathy of the RNA-binding protein TDP-43, characterized by nuclear clearance and cytoplasmic inclusion, is a hallmark of multiple neurodegenerative diseases, including amyotrophic lateral sclerosis (ALS), frontotemporal dementia (FTD), and Alzheimer's disease (AD). Through CRISPR interference (CRISPRi) screening in human neurons, we identified the decapping enzyme scavenger (DCPS) as a novel genetic modifier of TDP-43 loss-of-function (LOF)-mediated neurotoxicity. Our findings reveal that TDP-43 LOF leads to aberrant mRNA degradation, via disrupting the properties and function of processing bodies (P-bodies). TDP-43 interacts with P-body component proteins, potentially influencing their dynamic equilibrium and assembly into ribonucleoprotein (RNP) granules. Reducing DCPS restores P-body integrity and RNA turnover, ultimately improving neuronal survival. Overall, this study highlights a novel role of TDP-43 in RNA processing through P-body regulation and identifies DCPS as a potential therapeutic target for TDP-43 proteinopathy-related neurodegenerative diseases.

DOI: https://doi.org/10.1101/2025.06.13.659508
bioRxiv. 2025 Jun 29. pii: 2025.06.28.661837. [Epub ahead of print]

Inhibition of nonsense-mediated decay in TDP-43 deficient neurons reveals novel cryptic exons.

Irika R Sinha, Yingzhi Ye, Yini Li, Parker S Sandal, Philip C Wong, Shuying Sun, Jonathan P Ling.

TAR DNA-binding protein 43 kDa (TDP-43) is an essential splicing repressor whose loss of function underlies the pathophysiology of amyotrophic lateral sclerosis and frontotemporal dementia (ALS-FTD). Nuclear clearance of TDP-43 disrupts its function and leads to the inclusion of aberrant cryptic exons. These cryptic exons frequently introduce premature termination codons resulting in the degradation of affected transcripts through nonsense-mediated mRNA decay (NMD). Conventional RNA sequencing approaches thus may fail to detect cryptic exons that are efficiently degraded by NMD, precluding identification of potential therapeutic targets. We generated a comprehensive set of neuronal targets of TDP-43 in human iPSC-derived i 3 Neurons (i 3 N) by combining TDP-43 knockdown with inhibition of multiple factors essential for NMD, revealing novel cryptic targets. We then restored expression of selected NMD targets in TDP-43 deficient i 3 Ns and determined which genes improved neuronal viability. Our findings highlight the role of NMD in masking cryptic splicing events and identify novel potential therapeutic targets for TDP-43-related neurodegenerative disorders.

DOI: https://doi.org/10.1101/2025.06.28.661837
bioRxiv. 2025 Jul 08. pii: 2025.07.08.662712. [Epub ahead of print]

TDP-43 promotes efficient HSV-1 replication in human DRG-derived neurons.

Shirley E Braspenning, Denise Ohnezeit, Olivia A DeGulis, Angus C Wilson, Ian J Mohr.

TAR DNA-binding protein 43 (TDP-43) is a versatile nuclear RNA-binding protein that performs important functions in RNA localization, processing and stability. In the neurodegenerative disease amyotrophic lateral sclerosis (ALS) TDP-43 forms toxic, insoluble cytoplasmic aggregates that ultimately lead to neuronal loss. Although TDP-43 is expressed in every cell type, its function and subcellular localization are particularly important for neuronal homeostasis. However, it is unknown if TDP-43 has a role during herpesvirus infection. Herpes simplex virus type-1 (HSV-1), a ubiquitous neurotropic pathogen, is considered a contributing factor to neurodegenerative disorders. In this study, we tested the requirement for TDP-43 during HSV-1 infection in neuronal and non-neuronal cells. HSV-1 infection of epithelial cells and primary fibroblasts did not change overall TDP-43 abundance, nor did TDP-43 depletion detectably alter HSV-1 replication in a multicycle growth experiment. By contrast, when TDP-43 was depleted in neuronally derived, matured HD10.6 cells, HSV-1 infectious virus production was significantly reduced in both single- and multicycle growth experiments. Notably, TDP-43 depletion restricts viral lytic gene expression at the immediate-early phase. Through nanopore direct RNA-sequencing we uncovered enhanced intron retention in two essential viral genes upon TDP-43 depletion. Thus, while depletion of TDP-43 does not affect replication in epithelial cells and fibroblasts, TDP-43 is required for efficient replication in HD10.6 cells through modifying the abundance and splicing of viral mRNAs.
IMPORTANCE: Herpes simplex virus type-1 is a widespread neurotropic pathogen that can cause life-threatening infections of the brain and is increasingly linked to neurodegenerative disease. However, due to the lack of scalable in vitro human neuronal models or small animal models that recapitulate disease, little is known about virus-host interactions in neurons specifically. Using human epithelial cells, primary fibroblasts and a human neuron-derived cell line, we uncovered a cell type specific TDP-43 requirement for efficient HSV-1 virus replication. TDP-43 is a critical neuronal disease gene, and we showed it promotes virus gene expression and splicing of viral mRNAs in neuron-derived cells. This work provides valuable insights into the possible etiology of neurodegenerative disease and highlights the importance of studying virus-host interactions in relevant model systems.

DOI: https://doi.org/10.1101/2025.07.08.662712
bioRxiv. 2025 May 05. pii: 2025.04.20.648873. [Epub ahead of print]

TDP-43 toxic gain of function links ALS, FTD and Alzheimer's Disease through splicing dysregulation.

Welmoed Van Zuiden, Thea D Meimoun, Chen Bar, Aviad Siany, Lihi Moshe, Nancy S Yacovzada, Eviatar Weizman, Manuela Neumann, Aron S Buchman, Yanling Wang, David A Bennett, Jonathan D Glass, Adam N Trautwig, Nicholas T Seyfried, Johnathan Cooper-Knock, Eran Hornstein.

Loss of nuclear TDP-43 splicing activity is a common feature across neurodegenerative diseases including amyotrophic lateral sclerosis (ALS) and frontotemporal lobar degeneration (FTLD), but its relevance to Alzheimer's disease (AD) remains unclear. Here, we show that TDP-43 pathology in AD is broadly associated with splicing abnormalities, including aberrant splicing of amyloid precursor protein (APP). TDP-43 drives the formation of elongated APP isoforms, disrupting alternative splicing across ALS, FTLD-TDP and AD, providing a compelling mechanism for a long-standing observation of APP isoform dysregulation. We further establish a mechanistic link between TDP-43, APP splicing, and A-beta pathology. Surprisingly, the disruption to alternative APP splicing is mediated by a toxic gain of cytoplasmic TDP-43 function, rather than loss of its nuclear role. Using proximity proteomics and base editing in human iPSC-derived neurons, we show that TDP-43 pathology causes cytoplasmic co-sequestration of splicing regulators SCAF11, SRSF5, and TIAL1. Knockdown of these regulators also results in APP mis-splicing and increased A-beta burden, without affecting other TDP-43 targets such as STMN2 or UNC13A. Together, our findings suggest that TDP-43-mediated splicing dysfunction upstream of APP contributes to the pathogenesis of seemingly disparate neurodegenerative diseases, uniting AD and ALS/FTLD-TDP through a shared molecular mechanism.

DOI: https://doi.org/10.1101/2025.04.20.648873
Front Mol Biosci. 2025 ;12 1608853

Emerging biomarkers in amyotrophic lateral sclerosis: from pathogenesis to clinical applications.

Farah Anjum, Maha Bakhuraysah, Abdulaziz Alsharif, Taj Mohammad, Anas Shamsi, Md Imtaiyaz Hassan.

  Amyotrophic lateral sclerosis (ALS) is a severe neurodegenerative condition marked by the gradual loss of motor neurons in the brain and spinal cord. As the most common adult-onset motor neuron disease, ALS manifests through gradually worsening muscle weakness that ultimately progresses to complete paralysis. The disease presents in both sporadic and familial forms. Diagnosis is often delayed until substantial and irreversible motor neuron damage has already occurred. Clinical outcomes in ALS have only been defined through large-scale clinical trials with lengthy follow-up periods due to the disease's inherent heterogeneity and the absence of disease-specific biomarkers. Current biomarker detection methods, such as invasive cerebrospinal fluid (CSF) analysis or advanced imaging, are impractical for routine use, particularly in late-stage ALS. Several blood-based biomarkers have shown promise, including neurofilament levels, cryptic RNA-derived peptides, and immune-mediated changes, which may enable non-invasive monitoring. Nevertheless, the development of these methods is hindered by technical challenges, such as blood matrix interference and low analyte abundance. Among the emerging biomarkers, neurofilament light chain (NfL) appears to be the most promising, as its concentrations change in line with disease progression and distinguish clinically relevant groups. NfL facilitates patient stratification based on clinical progression rates (e.g., rapid vs slow progressors), while cryptic exon-derived peptides, such as UNC13A-derived peptides, enable genetic stratification by identifying molecular subtypes linked to TDP-43 pathology (e.g., C9orf72 vs sporadic ALS). These biomarkers hold promise to optimize clinical trial design through enriched cohort selection and accelerating therapeutic translation by monitoring target engagement. In this review, we have summarized recent developments in ALS biomarker studies, focusing on neurofilaments in each biofluid, transcriptomic signatures, and neuroinflammatory biomarkers, emphasizing technical challenges surrounding reproducibility in measurement. Finally, we discussed the potential integration of these biomarkers into clinical practice to advance drug development through precision medicine, thereby enabling shorter and more targeted clinical trials.

Keywords:  amyotrophic lateral sclerosis; biomarkers; molecular basis of neurodegeneration; neuroinflammation markers; therapeutic targets

DOI:  https://doi.org/10.3389/fmolb.2025.1608853
bioRxiv. 2025 Jul 09. pii: 2025.07.09.664014. [Epub ahead of print]

Nonsense-mediated decay masks cryptic splicing events caused by TDP-43 loss.

Yi Zeng, Odilia Sianto, Anastasiia Lovchykova, Chang Liu, Tetsuya Akiyama, Leonard Petrucelli, Aaron D Gitler.

In frontotemporal dementia and amyotrophic lateral sclerosis, the RNA-binding protein TDP-43 is lost from the nucleus, leading to cryptic exon inclusion events in dozens of neuronal genes. Here, we show that many cryptic splicing events have been missed by standard RNA-sequencing analyses because they are substrates for nonsense-mediated decay. By inhibiting nonsense-mediated decay in neurons we unmask hundreds of novel cryptic splicing events caused by TDP-43 depletion, providing a new picture to TDP-43 loss of function in neurons.

DOI: https://doi.org/10.1101/2025.07.09.664014
bioRxiv. 2025 Jun 08. pii: 2025.06.06.657909. [Epub ahead of print]

JIP4 deficiency causes a lysosomal storage disease arising from impaired cystine efflux.

Layla M Nassar, Xiaojian Shi, Agnes Roczniak-Ferguson, Hongying Shen, Shawn M Ferguson.

Lysosomes break down macromolecules, clear cellular waste and recycle nutrients such as cystine. We describe a novel mechanism whereby JIP4 regulates lysosomal cystine storage by controlling the abundance of cystinosin (CTNS), the transporter responsible for lysosomal cystine efflux. To this end, JIP4, previously characterized as a motor adaptor and kinase signaling scaffold, suppresses TMEM55B-dependent ubiquitylation of CTNS. Loss of JIP4 reduces CTNS protein levels, leading to lysosomal cystine accumulation and lysosomal storage defects that phenocopy loss of CTNS in both human cells and the renal proximal tubules of JIP4 knockout mice. These phenotypes mirror cystinosis, the lysosomal storage disease caused by CTNS loss-of-function. Our findings thus reveal a fundamental process that controls the efflux of lysosomal cystine and has relevance to understanding human disease arising from JIP4 mutations.

DOI: https://doi.org/10.1101/2025.06.06.657909
Cell Mol Life Sci. 2025 Jul 17. 82(1): 276

LRRK2 kinase modulates glucose-stimulated insulin secretion via RAB8 phosphorylation and ciliogenesis.

Nevia Dule, Algerta Marku, Alessandra Galli, Francesca Pischedda, Adriano Lama, Michela Castagna, Paola Marciani, Federico Bertuzzi, Giovanni Piccoli, Carla Perego.

  Leucine-rich repeat kinase 2 (LRRK2) encodes a multidomain protein whose mutations have been identified as genetic risk factors for Parkinson's disease (PD), an age-related neurodegenerative disorder. Outside the nervous system, LRRK2 is expressed in multiple tissues, including the endocrine pancreas, but its role here is unknown. Using pharmacological and molecular approaches, we show that LRRK2 kinase activity regulates stimulated insulin secretion by influencing secretory granule trafficking. The PD-associated LRRK2 mutant G2019S, characterized by enhanced kinase activity, increases the basal insulin release in complementary in vitro models and affects the metabolic profile in transgenic mice. Mechanistically, we demonstrate that LRRK2 kinase activity influences the formation of the primary cilium, an antenna-like structure acting as signaling platform to regulate hormones secretion. Specifically, LRRK2 phosphorylates RAB8 in a glucose-dependent manner, facilitating its recruitment to the primary cilium. These findings identify LRRK2 as a regulator of insulin secretion in pancreatic β-cells. Given the role of insulin signaling and glucose homeostasis in the nervous system, our data suggest that LRRK2 may also contribute to PD development through peripheral action.

Keywords:  Insulin; LRRK2; β-cell; RABs; Parkinson disease; Primary cilium

DOI:  https://doi.org/10.1007/s00018-025-05810-w
bioRxiv. 2025 Jun 09. pii: 2025.06.06.658328. [Epub ahead of print]

Accumulation of LRRK2-associated phospho-Rab12 degenerative lysosomes in tauopathies.

Silas A Buck, Tuyana Malankhanova, Eileen B Ma, Sarah Yim, Harrison W Pratt, John Ervin, Shih-Hsiu J Wang, Todd J Cohen, Andrew B West, Laurie H Sanders.

Parkinson's disease (PD) pathogenic mutations in leucine-rich repeat kinase 2 ( LRRK2 ) are associated with endolysosomal dysfunction across cell types, and carriers of LRRK2 mutations variably present with phosphorylated tau and α-synuclein deposits in post-mortem analysis. LRRK2 mutations increase the phosphorylation of Rab substrates including Rab12. Rab12 is expressed in neuronal and non-neuronal cells with localization to membranes in the endolysosomal compartment. Under lysosomal stress, LRRK2 interaction with Rab12 upregulates LRRK2 kinase activity. In this study, using a recently developed monoclonal antibody directed to the LRRK2-mediated phosphorylation site on Rab12 at amino acid Ser106 (pS106-Rab12), we test whether aberrant LRRK2 phosphorylation is associated with tau and/or α-synuclein pathology across clinically distinct neurodegenerative diseases. Analysis of brain tissue lysates and immunohistochemistry of pathology-susceptible brain regions demonstrate that pS106-Rab12 levels are increased in Dementia with Lewy bodies (DLB), Alzheimer's disease (AD), and PD, and in LRRK2 mutation carriers. In early pathological stages, phosphorylated Rab12 localizes to granulovacuolar degeneration bodies (GVBs), which are thought to be active lysosomal-like structures, in neurons. pS106-Rab12-positive GVBs accumulate with pathological tau across brain tissues in DLB, AD, and PD, and in LRRK2 mutation carriers. In a mouse model of tauopathy, pS106-Rab12 localizes to GVBs during early tau deposition in an age-dependent manner. While GVBs are largely absent in neurons with mature protein pathology, subsets of both tau and α-synuclein inclusions appear to incorporate pS106-Rab12 at later pathological stages. Finally, pS106-Rab12 labels GVBs in neurons and shows widespread co-pathology with tau inclusions in primary tauopathies including Pick's disease, progressive supranuclear palsy and corticobasal degeneration. These results implicate LRRK2 kinase activity and Rab phosphorylation in endolysosomal dysfunction in both tau and α-synuclein-associated neurodegenerative diseases.

DOI: https://doi.org/10.1101/2025.06.06.658328
Genome Res. 2025 Jul 17. pii: gr.279501.124. [Epub ahead of print]

Transcriptional modulation unique to vulnerable motor neurons predicts ALS across species and SOD1 mutations.

Irene Mei, Susanne Nichterwitz, Melanie Leboeuf, Jik Nijssen, Isadora Lenoel, Dirk Repsilber, Christian S Lobsiger, Eva Hedlund.

Amyotrophic lateral sclerosis (ALS) is characterized by the progressive loss of motor neurons (MNs) that innervate skeletal muscles. However, certain MN groups including ocular MNs, are relatively resilient. To reveal key drivers of resilience versus vulnerability in ALS, we investigate the transcriptional dynamics of four distinct MN populations in SOD1G93A ALS mice using LCM-seq and single molecule fluorescent in situ hybridization. We find that resilient ocular MNs regulate few genes in response to disease. Instead, they exhibit high baseline gene expression of neuroprotective factors including En1, Pvalb, Cd63 and Gal, some of which vulnerable MNs upregulate during disease. Vulnerable motor neuron groups upregulate both detrimental and regenerative responses to ALS and share pathway activation, indicating that breakdown occurs through similar mechanisms across vulnerable neurons, albeit with distinct timing. Meta-analysis across four rodent mutant SOD1 MN transcriptome datasets identify a shared vulnerability code of 39 genes including Atf4, Nupr1, Ddit3, and Penk, involved in apoptosis as well as proregenerative and anti-apoptotic signature consisting of Atf3, Vgf, Ina, Sprr1a, Fgf21, Gap43, Adcyap1, and Mt1 Machine learning using genes upregulated in SOD1G93A spinal MN predicts disease in human stem cell-derived SOD1E100G MNs, and shows that dysregulation of VGF, INA, and PENK are strong disease-predictors across species and SOD1 mutations. Our study reveals MN population-specific gene expression and temporal disease-induced regulation that together provide a basis to explain ALS selective vulnerability and resilience and that can be used to predict disease.

DOI: https://doi.org/10.1101/gr.279501.124
Bio Protoc. 2025 Jul 05. 15(13): e5367

Live Cell Imaging to Monitor Axonal Pruning in Drosophila Motor Neurons.

Keyao Long, Wanyue Xu, Xun Miao, Su Wang, Menglong Rui.

  Over the lifespan of an individual, brain function requires adjustments in response to environmental changes and learning experiences. During early development, neurons overproduce neurite branches, and neuronal pruning removes the unnecessary neurite branches to make a more accurate neural circuit. Drosophila motoneurons prune their intermediate axon bundles rather than the terminal neuromuscular junction (NMJ) by degeneration, which provides a unique advantage for studying axon pruning. The pruning process of motor axon bundles can be directly analyzed by real-time imaging, and this protocol provides a straightforward method for monitoring the developmental process of Drosophila motor neurons using live cell imaging. Key features • Long-range projecting axon bundles of Drosophila motor neurons extending from soma on the ventral nerve cord (VNC) undergo degeneration rather than retraction during metamorphosis. • The pruning process of motor axon bundles can be directly observed by real-time live-cell imaging. • The complete clearance of axon bundles occurs approximately 22 h after pupal formation (22 h APF). • Mushroom body (MB) γ neuron axon pruning regulatory genes are conserved for motor neurons.

Keywords:  Axonal pruning; Drosophila melanogaster; Live cell imaging; Motor neurons; Neurodevelopment

DOI:  https://doi.org/10.21769/BioProtoc.5367
bioRxiv. 2025 Jul 08. pii: 2025.06.25.661453. [Epub ahead of print]

Autophagy activators normalize aberrant Tau proteostasis and rescue synapses in human familial Alzheimer's disease iPSC-derived cortical organoids.

Sergio R Labra, Jadon Compher, Akhil Prabhavalkar, Mireya Almaraz, Claudia Cedeño Kwong, Christine Baal, Maria Talantova, Nima Dolatabadi, Julian Piña-Sanz, Yubo Wang, Leonard Yoon, Swagata Ghatak, Zi Gao, Yuting Zhang, Dorit Trudler, Lynee Massey, Wei Lin, Anthony Balistreri, Michael Bula, Nicholas J Schork, Tony S Mondala, Steven R Head, Jeffery W Kelly, Stuart A Lipton.

Alzheimer's disease (AD) is the most common form of dementia worldwide. Despite extensive progress, the cellular and molecular mechanisms of AD remain incompletely understood, partially due to inadequate disease models. To illuminate the earliest changes in hereditary (familial) Alzheimer's disease, we developed an isogenic AD cerebrocortical organoid (CO) model. Our refined methodology produces COs containing excitatory and inhibitory neurons alongside glial cells, utilizing established isogenic wild-type and diseased human induced pluripotent stem cells (hiPSCs) carrying heterozygous familial AD mutations, namely PSEN1 ΔE9/WT , PSEN1 M146V/WT , or APP swe/WT . Our CO model reveals time-progressive accumulation of amyloid beta (Aβ) species, loss of monomeric Tau, and accumulation of aggregated high-molecular-weight (HMW) phospho(p)-Tau species. This is accompanied by neuronal hyperexcitability, as observed in early human AD cases on electroencephalography (EEG), and synapse loss. Single-cell RNA-sequencing analyses reveal significant differences in molecular abnormalities in excitatory vs. inhibitory neurons, helping explain AD clinical phenotypes. Finally, we show that chronic dosing with autophagy activators, including a novel CNS-penetrant mTOR inhibitor-independent drug candidate, normalizes pathologic accumulation of Aβ and HMW p-Tau, normalizes hyperexcitability, and rescues synaptic loss in COs. Collectively, our results demonstrate these COs are a useful human AD model suitable for assessing early features of familial AD etiology and for testing drug candidates that ameliorate or prevent molecular AD phenotypes.

DOI: https://doi.org/10.1101/2025.06.25.661453
Brain Commun. 2025 ;7(4): fcaf261

Downregulation of NEAT1 due to loss of TDP-43 function exacerbates motor neuron degeneration in amyotrophic lateral sclerosis.

Yu Kawakami, Yohei Iguchi, Jiayi Li, Yoshinobu Amakusa, Takashi Yoshimura, Ryo Chikuchi, Satoshi Yokoi, Madoka Iida, Yuichi Riku, Yasushi Iwasaki, Tetsuro Hirose, Shinichi Nakagawa, Masahisa Katsuno.

  TAR DNA-binding protein 43 (TDP-43) is of particular interest in the pathogenesis of amyotrophic lateral sclerosis (ALS). It has been speculated that loss of nuclear TDP-43 and its cytoplasmic aggregation contributes to neurodegeneration. Although considerable attention has been paid to RNA metabolism in TDP-43 function, TDP-43 is also known to act as a transcription factor. This study found that the expression of Nuclear-enriched abundant transcript 1 (NEAT1), a long-non-coding RNA, was substantially downregulated in motor neurons with nuclear TDP-43 loss, but upregulated in those with preserved nuclear TDP-43, in the postmortem spinal cords of patients with sporadic ALS. TDP-43 depletion induced Neat1 downregulation in Neuro2a cells, primary cortical neurons, and mouse spinal motor neurons. Furthermore, TDP-43 was found to positively regulate NEAT1 at the transcriptional level. Finally, Neat1 knockout exacerbates neurodegeneration of hSOD1G93A mice accompanied by increased misfolded superoxide dismutase 1 (SOD1) aggregations. Transcriptome analysis revealed that Neat1 knockout reduced protein folding-related genes, such as heat shock protein family A member 1A (Hspa1a), in the spinal cords of hSOD1G93A mice. Our results indicated that the loss of TDP-43 function enhances ALS neurodegeneration by losing the protective effect of NEAT1.

Keywords:  NEAT1; TDP-43; amyotrophic lateral sclerosis; mouse model; neurodegeneration

DOI:  https://doi.org/10.1093/braincomms/fcaf261
Acta Neuropathol. 2025 Jul 17. 150(1): 5

Loss of the lysosomal lipid flippase ATP10B leads to progressive dopaminergic neurodegeneration and parkinsonian motor deficits.

María Sanchiz-Calvo, Elena Coccia, Christopher Cawthorne, Gustavo Morrone Parfitt, Teresa Torre-Muruzabal, George Tsafaras, Koen Van Laere, Diego Cabezudo, Ana Cascalho, Chris Van den Haute, Peter Vangheluwe, Joel Blanchard, Eduard Bentea, Veerle Baekelandt.

  ATP10B, a transmembrane lipid flippase located in late endosomes and lysosomes, facilitates the export of glucosylceramide and phosphatidylcholine by coupling this process to ATP hydrolysis. Recently, loss-of-function mutations in the ATP10B gene have been identified in Parkinson's disease patients, pointing to ATP10B as a candidate genetic risk factor. Previous studies have shown compromised lysosomal functionality upon ATP10B knockdown in human cell lines and primary cortical neurons. To investigate the role of ATP10B in Parkinson's disease neuropathology, specifically in the nigrostriatal dopaminergic system, we induced ATP10B knockdown specifically in substantia nigra pars compacta neurons of rats using viral vector technology. Additionally, midbrain neuronal cultures derived from ATP10B knock-out human induced pluripotent stem cells clones were used to study the impact of ATP10B loss in dopaminergic neurons in a more translational model. Atp10b knockdown in rat brain induced parkinsonian motor deficits, and longitudinal striatal dopamine transporter 18F-FE-PE2I PET imaging revealed a progressive decrease in binding potential. Immunohistochemical analysis conducted one year post-injection confirmed the loss of dopaminergic terminals in the striatum, alongside a loss of dopaminergic neurons in the substantia nigra pars compacta. The expression of LAMP1, LAMP2a, cathepsin B and glucocerebrosidase was studied in dopaminergic neurons. A decrease in lysosomal numbers and an increase in lysosomal volume were observed more consistently in one of the knockdown constructs. The vulnerability of dopaminergic neurons to ATP10B loss-of-function was also observed in midbrain neuronal cultures derived from ATP10B knock-out human induced pluripotent stem cells clones, which showed a significant reduction in TH-positive neurons. Taken together, our findings demonstrate that ATP10B depletion detrimentally impacts the viability of dopaminergic neurons both in vivo and in vitro. Moreover, a broader impact on the functionality of the nigrostriatal pathway was evidenced as rats with Atp10b knockdown exhibited motor impairments similar to those observed in Parkinson's disease patients.

Keywords:  ATP10B; Behavior; Dopaminergic neurons; In vivo; Lysosomes; Nigrostriatal pathway; PET; Parkinson’s disease

DOI:  https://doi.org/10.1007/s00401-025-02908-0
Curr Opin Cell Biol. 2025 Jul 16. pii: S0955-0674(25)00108-5. [Epub ahead of print]96 102570

Cytoskeletal scaffolding of NaVs and KVs in neocortical pyramidal neurons: Implications for neuronal signaling and plasticity.

Carina C Elvira, Paul M Jenkins.

The initiation and propagation of action potentials (APs) depend on the precise localization of voltage-gated sodium (NaV) and potassium (KV) channels in neurons. In neocortical pyramidal neurons, NaV1.2 and NaV1.6 are key at the axon initial segment (AIS) and nodes of Ranvier (noR), driving AP initiation and propagation. NaV1.2 also supports AP backpropagation in the soma and dendrites. Ankyrin-G anchors these channels at the AIS and noR, while new findings reveal that ankyrin-B scaffolds NaV1.2 in dendrites. This review highlights how ankyrins stabilize NaV and KV channels across neuronal domains, ensuring proper function crucial for excitability, synaptic plasticity, and signaling. Recent findings explore how ankyrins differentially localize NaV1.2 and NaV1.6, with implications for understanding neurological disorders linked to disrupted channel localization.

DOI: https://doi.org/10.1016/j.ceb.2025.102570
bioRxiv. 2025 Jul 13. pii: 2025.07.11.664426. [Epub ahead of print]

KIF5A downregulation in spinal muscular atrophy links axonal regeneration defects with ALS.

Tetsuya Akiyama, Yi Zeng, Caiwei Guo, Olivia Gautier, Lauren S Koepke, Juliane Bombosch, Odilia Sianto, Jay P Ross, Phuong T Hoang, Luke Yuchen Zhao, Cole Spencer, Michelle Monje, John W Day, Aaron D Gitler.

Spinal muscular atrophy (SMA) is a devastating neuromuscular disorder caused by mutations in the Survival Motor Neuron 1 (SMN1) gene, leading to decreased SMN levels and motor neuron dysfunction. SMN-restoring therapies offer clinical benefit, but the downstream molecular consequences of SMN reduction remain incompletely understood. Here, we demonstrate that SMN deficiency results in downregulation of KIF5A in human neurons and in a mouse model of SMA. We provide evidence that reduced SMN levels impair axon regeneration, which is rescued by KIF5A overexpression and that the RNA-binding protein SMN functions to stabilize KIF5A mRNA. These findings provide evidence of a molecular link between SMA and ALS pathophysiology, highlighting KIF5A as a new SMN target. Our findings suggest SMN-independent interventions targeting KIF5A could represent a complementary therapeutic approach for SMA and other motor neuron diseases.

DOI: https://doi.org/10.1101/2025.07.11.664426
F1000Res. 2025 ;14 10

A guide to selecting high-performing antibodies for Stearoyl-CoA desaturase (SCD1) (UniProt ID: O00767) for use in western blot, immunoprecipitation, and immunofluorescence.

NeuroSGC/YCharOS/EDDU collaborative group

  The enzyme stearoyl-CoA desaturase (SCD1) is a modulator of lipid metabolism by catalyzing the biosynthesis of mono-unsaturated fatty acids from saturated fatty acids. Understanding the specific mechanisms by which SCD1 plays in health and disease can provide novel insides in therapeutic targets, a process that would be facilitated by the availability of high-quality antibodies. Here we have characterized nine SCD1 commercial antibodies for western blot, immunoprecipitation, and immunofluorescence using a standardized experimental protocol based on comparing read-outs in knockout cell lines and isogenic parental controls. These studies are part of a larger, collaborative initiative seeking to address antibody reproducibility issues by characterizing commercially available antibodies for human proteins and publishing the results openly as a resource for the scientific community. While use of antibodies and protocols vary between laboratories, we encourage readers to use this report as a guide to select the most appropriate antibodies for their specific needs.

Keywords:  O00767; SCD; SCD1; Steroyl-CoA desaturase; antibody characterization; antibody validation; immunofluorescence; immunoprecipitation; western blot

DOI:  https://doi.org/10.12688/f1000research.160217.2
bioRxiv. 2025 Jun 23. pii: 2025.05.06.652480. [Epub ahead of print]

Cholinergic neuron circadian clock mediates RNA-binding protein function and contributes to ALS disease phenotypes.

Sharon B Tam, Nathan J Waldeck, Matthew Wright, Erin M Baker, Evangelos Kiskinis, Joseph Bass, Robert G Kalb.

Circadian clocks are encoded by a transcription-translation feedback loop that aligns physiological processes with the solar cycle. Previous work linking the circadian clock to the regulation of RNA-binding proteins (RBPs) and alternative splicing provides a foundation for the vital examination of their mechanistic connections in the context of amyotrophic lateral sclerosis (ALS)-a fatal neurodegenerative disease characterized by disrupted RBP function. Here, we reveal enrichment of genes associated with ALS and other neurodegenerative diseases in the spinal cord cholinergic neuron rhythmic transcriptome. We demonstrate that there is circadian regulation of ALS-linked RBPs and rhythmic alternative splicing of genes involved in intracellular transport ( Aftph and Mvb12a ), microtubule cytoskeleton organization ( Limch1 and Drc3 ), and synaptic function ( Sipa1l2 ) in this neuronal sub-type. Further, we show that the cholinergic neuron clock regulates sporadic ALS-associated changes in cytoskeleton and neuromuscular junction synapse gene expression. Finally, we report that cell-type-specific Bmal1 -deletion (i) increases lumbar spinal cord motor neuron loss and sciatic nerve axon degeneration, (ii) drives alternative splicing of genes encoding ALS-linked RBPs ( Matr3 and Srsf7 ), and (iii) drives alternative splicing of genes associated with microtubule transport and postsynaptic organization. Our results establish a role for the cholinergic neuron circadian clock in RBP function and ALS disease phenotypes.

DOI: https://doi.org/10.1101/2025.05.06.652480
Stem Cells. 2025 Jul 16. pii: sxaf050. [Epub ahead of print]

Neuronal branching in stem cell models of mitochondrial and neurological diseases.

Selene Lickfett, Carmen Menacho, Sidney Cambridge, Alessandro Prigione.

  Neuronal branching, the extension and arborization of neurites, is critical for establishing and maintaining functional neural circuits. Emerging evidence suggests that mitochondria play an important role in regulating this process. In this review, we explore how the use of human induced pluripotent stem cell (iPSC)-derived neuronal models in two dimensions (2D) and three dimensions (3D) could help uncover possible mechanisms linking mitochondrial function and dysfunction to neuronal branching capacity. We highlight examples of iPSC-based models of mitochondrial and neurological diseases where aberrant neurite growth has been observed and discuss the potential therapeutic implications. Additionally, we review current methodologies for assessing neurite outgrowth in 2D and 3D neuronal models, addressing their strengths and limitations. Insights gained from these models emphasize the significance of mitochondrial health in neuronal branching and demonstrate the potential of iPSC-derived neurons and brain organoids for studying disrupted neuronal morphology. Harnessing these human stem cell models to devise phenotypic drug discovery platforms can eventually pave the way for innovative therapeutic interventions, particularly in the context of disorders with poorly understood genetic mechanisms and limited therapeutic options.

Keywords:  iPSCs; mitochondria; mitochondrial diseases; neurodegeneration; neuronal branching; neurons

DOI:  https://doi.org/10.1093/stmcls/sxaf050
Front Cell Neurosci. 2025 ;19 1627517

Potassium channel clustering: mechanisms shaping axonal excitability.

Gabriel Escobedo, Matthew N Rasband.

  The precise clustering of ion channels at axon initial segments (AIS) and nodes of Ranvier is essential for axonal excitability and rapid action potential propagation. Among the axonal ion channels, voltage-gated potassium channels (Kv) and two-pore domain potassium (K2P) leak channels are key regulators of AIS and nodal excitability. Kv7 and Kv1 channels contribute to action potential threshold and repolarization at the AIS, and membrane repolarization in axons has historically been attributed to Kv channels. However, recent studies suggest that at nodes of Ranvier K2P channels, particularly TRAAK and TREK-1, play a dominant role in action potential repolarization. The interaction of Kv and K2P channels with diverse scaffolding proteins ensures their precise localization at AIS and nodes. Mislocalization or dysfunction of axonal Kv and K2P channels can cause epilepsy and neurodevelopmental disorders. This review explores the diversity of potassium channels and the mechanisms responsible for their clustering at AIS and nodes of Ranvier. Understanding these processes will be essential for therapeutic strategies aimed at treating diseases characterized by abnormal potassium channel expression, clustering, and function in neurons.

Keywords:  axon; axon initial segment; ion channel; node of Ranvier; scaffold

DOI:  https://doi.org/10.3389/fncel.2025.1627517
J Cell Physiol. 2025 Jul;240(7): e70064

The Role of Mitochondrial Dynamics in Doxorubicin-Induced Disease: Implications for Therapy.

Huan Yue, Yousheng Chen, Junxiao Feng, Weiying Yue, Xingjuan Shi.

  As an anthracycline chemotherapy drug, doxorubicin (Dox) is generally prescribed to treat a variety of malignant tumors. Nevertheless, Dox exhibited toxicity at a high dosage, which might eventually lead to injury of the body. Mitochondrial dynamics, including mitochondrial fission and fusion, regulates mitochondrial homeostasis and cellular function. Mounting evidence has demonstrated that imbalance in mitochondrial dynamics, manifested by increased mitochondrial fission or decreased mitochondrial fusion, is associated with the development of Dox-induced diseases. In this paper, we will elaborate the role of mitochondrial dynamics in Dox-induced diseases, and discuss the regulatory mechanism of mitochondrial dynamics in Dox-induced diseases, including apoptosis, fibrosis, myocardial atrophy and inflammation. Elucidating these issues may provide important value in the diagnosis and potential therapeutic strategies for Dox-induced diseases through regulation of mitochondria dynamics.

Keywords:  Dox‐induced diseases; apoptosis; fibrosis; mitochondrial dynamics; mitochondrial fission; mitochondrial fusion

DOI:  https://doi.org/10.1002/jcp.70064
Proc Natl Acad Sci U S A. 2025 Jul 22. 122(29): e2503342122

NEDD4L induces mitochondrial dysfunction and neurodegeneration by promoting LIPT2 degradation in Huntington's disease.

Pan Fan, Yaqing Liu, Chunyue Liu, Hao Yao, Shibo Xu, Yueqing Jiang, Yang Wu, Yan Liu, Xing Guo.

  Impairment of mitochondrial protein stability is associated with neurodegeneration in Huntington's disease (HD). However, the E3 ligase responsible for maintaining mitochondrial protein homeostasis in HD remains poorly understood. In this study, we demonstrate that NEDD4L protein levels are elevated in human striatal organoids (hSOs) derived from induced pluripotent stem cells of patients as well as in a mouse model of HD. Overexpression of NEDD4L leads to degeneration and cell death of medium spiny neurons (MSNs), along with a reduction in motor activities. Conversely, deletion of NEDD4L restores abnormal MSN morphology, corrects deficits in calcium signaling, alleviates neurodegeneration in HD-hSOs, and improves motor dysfunction observed in YAC128 mice. Mechanistically, NEDD4L disrupts mitochondrial function by binding to lipoyl(octanoyl) transferase 2 (LIPT2) and promoting its degradation through ubiquitination and lysosomal pathways. This process impairs lipoic acid biosynthesis and the lipoylation of E2 subunits of alpha-ketoglutarate dehydrogenase (α-KGDH E2). Furthermore, either overexpressing LIPT2 or administering lipoic acid mitigates neurodegeneration and rectifies deficits in motor coordination activity. These findings unveil a molecular mechanism underlying the regulation of lipoic acid metabolism and underscore the potential therapeutic role of protein lipoylation in the treatment of HD.

Keywords:  Huntington’s disease; LIPT2; NEDD4L; neurodegeneration; ubiquitin

DOI:  https://doi.org/10.1073/pnas.2503342122
Mol Cell Biol. 2025 Jul 16. 1-16

The Ubiquitin E3 Ligase UBE3A Regulates GRIPAP1 and PACSIN1 Proteins Linked to the Endocytic Recycling of AMPA Receptors.

Laura Drebushenko, Natalya Belous, Fritz W Lischka, Qiong Zhou, Ayse Malci, Michael S Sidorov, Barrington Burnett, Martin L Doughty.

  Angelman syndrome (AS) is a neurodevelopmental disorder characterized by cognitive and language impairments, seizures, reduced or fragmented sleep, motor ataxia, and a characteristic happy affect. AS arises due to the neuronal loss of UBE3A, an E3 ligase that regulates protein abundance through the addition of lysine 48 (K48)-linked polyubiquitin chains to proteins targeted for degradation by the ubiquitin proteasome system (UPS). Using a dual SMAD inhibition protocol to derive cortical neurons from human induced pluripotent stem cells, we examined UBE3A deletion effects on the neuronal proteome by liquid chromatography tandem mass spectrometry (LC-MS/MS). LC-MS/MS identified 645 proteins differentially abundant between UBE3A knockout (KO) and isogenic UBE3A wild-type control cortical neurons. Proteins with increased abundance with UBE3A loss of function include GRIPAP1 and PACSIN1, synaptic proteins implicated in AMPA receptor recycling. We provide evidence UBE3A polyubiquitinates PACSIN1 and GRIPAP1 to regulate protein turnover, with potential implications for impaired activity-dependent synaptic plasticity observed in AS.

Keywords:  AMPA receptor; GRIPAP1; PACSIN1; UBE3A; cortical neuron; human induced pluripotent stem cell; synapse; ubiquitin

DOI:  https://doi.org/10.1080/10985549.2025.2470431
ACS Chem Neurosci. 2025 Jul 17.

Novel Dual Mechanism GRT-X Agonist Acting on Kv7 Potassium Channel/Translocator Protein Receptor Prevents Motoneuron Degeneration Following Exposure to Mouse and Human Amyotrophic Lateral Sclerosis/Frontotemporal Dementia Astrocyte-Conditioned Media.

Vera M Masegosa, Elsa Fritz, Daniela Corvalan, Fabiola Rojas, Polett Garcés, Xavier Navarro, Petra Bloms-Funke, Brigitte van Zundert, Mireia Herrando-Grabulosa.

  Amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD) form a continuous spectrum of aggressive neurodegenerative diseases affecting primarily motoneurons (MNs) and cortical frontotemporal neurons. Noncell autonomous mechanisms contribute to ALS/FTD, wherein astrocytes release toxic factor(s) detrimental to MNs. Because of the multifactorial nature of ALS, single-pathway-focused therapies have limited effectiveness in improving ALS. Therefore, novel combinatorial therapies are currently being pursued. Here, we evaluated whether the simultaneous activation of two complementary targets, the voltage-gated potassium channels 7.2/3 (Kv7.2/3) and the mitochondrial translocator protein (TSPO), by a novel synthesized compound (GRT-X) is an effective neuroprotective treatment in ALS in vitro models. We exposed primary rat ventral spinal cord neuronal cultures and rat spinal cord organotypic cultures to astrocyte-conditioned medium derived from primary mouse ALS astrocytes expressing mutant human SOD1 (SOD1G93A-ACM) or from human-induced pluripotent stem cell (iPSC)-derived astrocytes carrying an ALS-causing mutation in SOD1 (SOD1D90A-ACM) or an ALS/FTD-causing mutation in TDP-43 (TDP43A90 V-ACM). We report that the diverse human and mouse ALS/FTD-ACMs compromise the MN viability. Remarkably, GRT-X led to consistent protection of MNs. Moreover, ALS/FTD-ACM increases oxidative stress levels, which are prevented with GRT-X treatment. Together, we show that the complementary activation of TSPO and Kv7.2/3 may offer a novel therapeutic strategy for ALS/FTD due to its capacity to protect MNs from noncell-autonomous toxicity induced by diseased astrocytes.

Keywords:  GRT-X; amyotrophic lateral sclerosis; astrocyte conditioned medium; frontotemporal dementia; motoneuron death; oxidative and excitotoxity stress

DOI:  https://doi.org/10.1021/acschemneuro.5c00197
bioRxiv. 2025 May 08. pii: 2025.05.02.651934. [Epub ahead of print]

Modeling human retinal ganglion cell axonal outgrowth, development, and pathology using pluripotent stem cell-based microfluidic platforms.

Cátia Gomes, Kang-Chieh Huang, Sailee S Lavekar, Jade Harkin, Carson Prosser, Yue Fang, Claire Kalem, Adrian Oblak, Chi Zhang, Jason S Meyer.

Retinal ganglion cells (RGCs) are highly compartmentalized cells, with long axons serving as the sole connection between the eye and the brain. RGC degeneration in injury and/or disease also occurs in a compartmentalized manner, with distinct injury responses in axonal and somatodendritic compartments. Thus, the goal of this study was to establish a novel microfluidic-based platform for the analysis of RGC compartmentalization in health and disease states. Human pluripotent stem cell (hPSC)-derived RGCs were seeded into microfluidics, enabling the recruitment and isolation of axons apart from the somatodendritic compartment. Initial studies explored axonal outgrowth and compartmentalization of axons and dendrites. We then compared the differential response of RGCs differentiated from hPSCs carrying the OPTN(E50K) glaucoma mutation with isogenic control RGCs in their respective axonal and somatodendritic compartments, followed by analysis of axonal transport. Further, we explored the axonal transcriptome via RNA-seq, focusing on disease-related axonal differences. Finally, we established models to uniquely orient astrocytes along the axonal compartment combined with modulation of astrocyte reactivity as a pathological feature of neurodegeneration. Overall, RGC culture within microfluidic chips allowed enhanced cell growth and maturation, including long-distance axonal projections and proper compartmentalization, while patient-specific RGCs exhibited axonal outgrowth deficits as well as decreased rate of axonal transport. Finally, the induction of astrocyte reactivity uniquely along the proximal region of RGC axons led to the onset of neurodegenerative phenotypes in RGCs. These results represent the first study to effectively recapitulate the highly compartmentalized properties of hPSC-derived RGCs in healthy and disease states, providing a more physiologically relevant in vitro model for neuronal development and degeneration.

DOI: https://doi.org/10.1101/2025.05.02.651934
bioRxiv. 2025 Jun 28. pii: 2025.06.26.661831. [Epub ahead of print]

Expanding the spectrum of annexin A11 proteinopathy in frontotemporal lobar degeneration and motor neuron disease.

Nikhil B Ghayal, Richard J Crook, Angita Jain, Gunveen Sachdeva, Shanu F Roemer, Hiroaki Sekiya, Michael A DeTure, Matthew Baker, Wouter De Coster, Bjorn E Oskarsson, Keith A Josephs, Rosa Rademakers, Marka van Blitterswijk, Dennis W Dickson.

Aggregation of TAR DNA-binding protein 43 (TDP-43) is strongly associated with frontotemporal lobar degeneration (FTLD-TDP), motor neuron disease (MND-TDP), and overlap disorders like FTLD-MND. Three major forms of motor neuron disease are recognized and include primary lateral sclerosis (PLS), amyotrophic lateral sclerosis (ALS), and progressive muscular atrophy (PMA). Annexin A11 (ANXA11) is understood to aggregate in amyotrophic lateral sclerosis (ALS-TDP) associated with pathogenic variants in ANXA11 , as well as in FTLD-TDP type C. Given these observations and recent reports of ANXA11 variants in patients with semantic variant frontotemporal dementia (svFTD) and FTD-MND presentations, we sought to characterize ANXA11 proteinopathy in an autopsy cohort of 379 cases with FTLD-TDP, as well as FTLD-MND and MND-TDP cases subclassified neuropathologically into PLS, ALS, and PMA. All FTLD-TDP type C cases had ANXA11 proteinopathy. However, ANXA11 proteinopathy was present in over 40% of FTLD-MND and in 38 out of 40 FTLD-PLS cases (95%), of which 80% had TDP type B or an unclassifiable TDP-43 proteinopathy and 15% had TDP type C. Genetic analyses excluded pathogenic ANXA11 variants in all ANXA11-positive cases. We thus demonstrated novel forms of ANXA11 proteinopathy strongly associated with FTLD-PLS, but not with TDP type C or pathogenic ANXA11 variants. Given the emerging relationship of ANXA11 in TDP-43 proteinopathies, we propose that TDP-43 and ANXA11 proteinopathy (TAP) comprises the molecular pathology of cases with abundant inclusions that are co-immunoreactive for both proteins and we subclassify three types of TAP based on distinct clinical and neuropathologic features.

DOI: https://doi.org/10.1101/2025.06.26.661831
NPJ Aging. 2025 Jul 14. 11(1): 64

The Rab3 family proteins in age-related neurodegeneration: unraveling molecular pathways and potential therapeutic targets.

Haijun He, Ruixue Ai, Evandro Fei Fang, Konstantinos Palikaras.

The Rab3 protein family is composed of a series of small GTP-binding proteins, including Rab3a, Rab3b, Rab3c, and Rab3d, termed Rab3s. They play crucial roles in health, including in brain function, such as through the regulation of synaptic transmission and neuronal activities. In the high-energy-demanding and high-traffic neurons, the Rab3s regulate essential cellular processes, including trafficking of synaptic vesicles and lysosomal positioning, which are pivotal for the maintenance of synaptic integrity and neuronal physiology. Emerging findings suggest that alterations in Rab3s expression are associated with age-related neurodegenerative pathologies, including Alzheimer's disease, Parkinson's disease, and Huntington's disease, among others. Here, we provide an overview of how Rab3s dysregulation disrupts neuronal homeostasis, contributing to impaired autophagy, synaptic dysfunction, and eventually leading to neuronal death. We highlight emerging questions on how Rab3s safeguards the brain and how their dysfunction contributes to the different neurodegenerative diseases. We propose fine-tuning the Rab3s signaling directly or indirectly, such as via targeting their upstream protein AMPK, holding therapeutic potential.

DOI: https://doi.org/10.1038/s41514-025-00257-6
bioRxiv. 2025 May 01. pii: 2024.09.08.611864. [Epub ahead of print]

GSK-3β coordinates axonal microtubule organisation through Shot and Tau.

Andre Voelzmann, Lubna Nuhu-Soso, Alex Roof, Sanjai Patel, Hayley Bennett, Antony Adamson, Marvin Bentley, Gareth J O Evans, Ines Hahn.

Glycogen Synthase Kinase 3β (GSK-3β) is a key coordinator of neuronal development and maintenance; hyperactive GSK-3β is linked to neurodevelopmental and -degenerative diseases and therefore a promising therapeutic target. In neurons, GSK-3β coordinates the cytoskeleton by phosphorylating microtubule-binding proteins. In this study, we found that tight regulation of GSK-3β kinase activity is required for the maintenance of parallel microtubule bundles in Drosophila and rat axons. Up- or down-regulation of GSK-3β led to axons forming pathological swellings in which microtubule bundles disintegrated into disorganised, curled microtubules. We identified the microtubule bundling proteins Shot and Tau as key GSK-3β targets and found that GSK-3β exerted its regulatory effect on microtubule bundling through them. GSK-3β regulates the ability of Shot and Tau to attach to microtubules and/or the plus-end protein Eb1. Mis-regulation of GSK-3β leads to the loss of Eb1-Shot-mediated guidance of polymerising microtubules into parallel bundles, thus causing disorganisation. We propose microtubule disorganisation as a new explanation for how GSK-3β hyperactivity leads to neurodegeneration and why global inhibition of GSK-3β has not been successful in clinical trials for neuronal disorders.

DOI: https://doi.org/10.1101/2024.09.08.611864
bioRxiv. 2025 Jun 11. pii: 2025.06.07.658463. [Epub ahead of print]

Electroporation-Based Gene Delivery and Whole-Organoid Imaging in Human Retinal Organoids.

Keevon Flohr, Michael Janecek, Lingyun Wang, Vicente Valle, Shaohua Pi, Rui T Peixoto, Susana da Silva.

Human retinal organoids (hRetOrg) derived from human induced pluripotent stem cells (hiPSCs) have emerged as powerful in vitro systems for studying retinal development, modeling retinal diseases, and evaluating therapeutic strategies. However, current genetic manipulation approaches, such as stable hiPSC line generation and viral transduction, are laborious, costly, and inefficient, with limited spatial specificity and high variability. Here, we report a rapid, scalable, and spatially precise electroporation-based platform for efficient plasmid-based gene delivery in early-stage hRetOrg. This method enables tunable and region-specific transfection of retinal progenitor cells without viral vectors or clonal selection. Coupled with resonant-scanning two-photon microscopy, this approach allows fast live cell imaging of whole organoids with subcellular resolution. This versatile system supports high-throughput genetic manipulation and imaging in intact hRetOrg, advancing studies of human retinal development, gene function, and disease.
Motivation: hRetOrgs offer an unprecedented platform for functional genetic studies of human retinal development and disease. However, existing methods for gene manipulation in hRetOrg are limited by low throughput, inefficiency, and lack of scalability, hindering systematic analysis of gene function and regulatory elements. To address these limitations, we developed a streamlined, high-efficiency pipeline that enables spatially targeted electroporation of hRetOrg during early retinogenesis, combined with fast, high-resolution imaging of whole organoids using two-photon microscopy, allowing studies at both tissue and subcellular scales.

DOI: https://doi.org/10.1101/2025.06.07.658463
Trends Immunol. 2025 Jul 12. pii: S1471-4906(25)00171-1. [Epub ahead of print]

Low-dose IL-2 reinvigorates the immunoguardians of neurodegenerative diseases.

Yunbo Wei, Yu Zhang, Di Yu.

  Amyotrophic lateral sclerosis (ALS) is a life-threatening neurodegenerative disease caused by motor neuron loss. In a recent Phase 2b trial, Bensimon and colleagues report that the addition of low-dose interleukin 2 (LD-IL-2) immunotherapy to standard of care (SOC) shows promise in enhancing immune tolerance and improving survival in individuals with slower disease progression.

Keywords:  Treg cells; amyotrophic lateral sclerosis (ALS); biomarker; immunomodulation; low-dose IL-2

DOI:  https://doi.org/10.1016/j.it.2025.07.004
bioRxiv. 2025 May 05. pii: 2025.05.04.652084. [Epub ahead of print]

Enhanced axonal transport in large vertebrates: KIF5A adaptations in giraffes and pythons.

Taketoshi Kambara, Lu Rao, Yosuke Yamagishi, Kazuho Ikeda, Daisuke Taniguchi, Tsuyoshi Imasaki, Hideki Shigematsu, Naoki Sakai, Arne Gennerich, Ryo Nitta, Yasushi Okada.

Efficient axonal transport is essential for neuronal function, particularly in species with exceptionally long axons. The Kinesin-1 motor protein KIF5A plays a key role in this process, but whether and how it adapts to the transport demands of large vertebrates remains unclear. Here, we show that KIF5A from giraffes (GcKIF5A) and pythons (PbKIF5A), moves 25% faster than its mouse counterpart (MmKIF5A) on neuronal microtubules in vitro and within axons of cultured mouse hippocampal neurons. This enhanced velocity, driven by three unique amino acid substitutions (R114Q, S155A and Y309F), facilitates long-distance transport in these species and represents a case of convergent evolution. Our structural analysis reveals that accelerated ADP release underlies the increased speed of GcKIF5A. Despite exhibiting a reduced force generation, GcKIF5A maintains efficient cargo transport under load. Furthermore, GcKIF5A exerts less drag in mixed-motor environments, an adaptation particularly beneficial for multiple motor-driven long-distance axonal transport. These findings reveal that KIF5A has evolved specific adaptations to facilitate efficient axonal transport in large vertebrates, highlighting the evolutionary plasticity of kinesin motors.

DOI: https://doi.org/10.1101/2025.05.04.652084
bioRxiv. 2025 Jun 12. pii: 2025.06.09.658689. [Epub ahead of print]

Treatment of age-related decreases in GTP levels restores endocytosis and autophagy.

R A Santana, J M McWhirt, G J Brewer.

Age-related declines in neuronal bioenergetic levels may limit vesicular trafficking and autophagic clearance of damaged organelles and proteins. Age-related ATP depletion would impact cognition dependent on ionic homeostasis, but limits on proteostasis powered by GTP are less clear. We used neurons isolated from aged 3xTg-AD Alzheimer's model mice and a novel genetically encoded fluorescent GTP sensor (GEVAL) to evaluate live GTP levels in situ. We report an age-dependent reduction in ratiometric measurements of free/bound GTP levels in living hippocampal neurons. Free-GTP co-localized in the mitochondria decreased with age accompanied by the accumulation of free-GTP labeled vesicular structures. The energy dependence of autophagy was demonstrated by depletion of GTP with rapamycin stimulation, while bafilomycin inhibition of autophagy raised GTP levels. 24 hr. supplementation of aged neurons with the NAD precursor nicotinamide and the Nrf2 redox modulator EGCG restored GTP levels to youthful levels and mobilized endocytosis and lysosomal consumption for autophagy via the respective GTPases Rab7 and Arl8b. This vesicular mobilization promoted the clearance of intraneuronal Aβ aggregates and lowered protein oxidative nitration in AD model neurons. Our results reveal age- and AD-related neuronal GTP energy deficits that impair autophagy and endocytosis. GTP deficits were remediated by an external NAD precursor together with a Nrf2 redox modulator which suggests a translational path.

DOI: https://doi.org/10.1101/2025.06.09.658689
bioRxiv. 2025 Jun 25. pii: 2025.06.23.661190. [Epub ahead of print]

Tau endo-lysosomal processing in human iPSC-derived microglia is impacted by tau aggregation state, but not by microglial activation status.

Andrew Shultz, Anna Vincze, Todd T Yau, Emma L Poirier, Jennifer N Rauch.

Microglia are the tissue resident macrophages of the brain and their contribution to tau pathology progression remains to be fully understood. In this study, we developed a quantitative platform to elucidate the endo- lysosomal regulation of tau within human induced pluripotent stem cell (iPSC)-derived microglia. We show that iPSC-derived microglia internalize monomeric and fibrillar tau through different cellular mechanisms and with different degradation kinetics. Acute inflammatory activation of microglia alters tau endocytosis, but surprisingly does not impact lysosomal clearance. These results highlight the importance of the microglial endo-lysosome system as a regulator of tau pathology that is decoupled from acute microglial activation.
Highlights: Human iPSC-derived microglia endocytose tau using different cellular mechanismsNanoBiT system can measure tau endocytosis/degradation in iPSC-derived microgliaAggregation of tau impacts the rate of lysosomal degradation after endocytosisAcute inflammation affects total endocytosed tau, but not degradation in microglia.

DOI: https://doi.org/10.1101/2025.06.23.661190
Cell Commun Signal. 2025 Jul 16. 23(1): 341

Mitochondria from huntington´s disease striatal astrocytes are hypermetabolic and compromise neuronal branching.

Laura López-Molina, Alba Pereda-Velarde, Nadia di Franco, Imme Aerts, Elisa Sebastià, Laura Valls-Roca, Mariona Guitart-Mampel, Gloria Garrabou, Silvia Gines.

   BACKGROUND: Deficits in mitochondrial bioenergetics and dynamics are strongly implicated in the selective vulnerability of striatal neurons in Huntington´s disease. Beyond these neuron-intrinsic factor, increasing evidence suggest that non-neuronal mechanisms, particularly astrocytic dysfunction involving disrupted homeostasis and metabolic support also contribute to disease progression. These findings underscore the critical role of metabolic crosstalk between neurons and astrocytes in maintaining striatal integrity. However, it remains unclear whether this impaired communication affects the transfer of mitochondria from astrocytes to striatal neurons, a potential metabolic support mechanism that may be compromised in Huntington´s Disease.
METHODS: Primary striatal astrocytes were obtained from wild-type and R6/1 mice to investigate mitochondrial dynamics. Expression levels of key mitochondrial fusion and fission proteins were quantified by Western blotting and RT-PCR. Mitochondria morphology, oxidative stress and membrane potential were assessed using confocal microscopy following staining with mitochondria-specific dyes. Mitochondrial respiration was measured using the Oxygraph-2k respirometer system (Oroboros Instruments). Transmitophagy was evaluated by confocal imaging after labeling astrocytic mitochondria with Mitotracker dyes. To assess the functional impact of mitochondrial transfer on neurons, Sholl analysis, neuronal death and oxidative stress levels were quantified using specific fluorogenic probes.
RESULTS: Striatal astrocytes from HD mice exhibited a significant increase in mitochondrial fission, and mitochondrial oxidative stress, mirroring alterations previously reported in striatal neurons. Analysis of mitochondrial oxygen consumption rate (OCR) revealed elevated respiration activity and enhanced ATP-linked respiration, indicative of a hypermetabolic state. Concurrently, increased lactate production suggested a shift toward dysregulated astrocytic energy metabolism. These mitochondrial alterations were functionally detrimental: astrocytic mitochondria derived from HD mice when taken up by striatal neurons via transmitophagy, led to reduced neuronal branching and disrupted oxidative homeostasis.
CONCLUSIONS: Our findings demonstrate that striatal astrocytes from HD mice exhibit a hypermetabolic phenotype, characterized by increased mitochondrial respiration, disrupted mitochondrial dynamics, and elevated mitochondrial oxidative stress. Importantly, we identify a novel mechanism of astrocyte-neuron interaction involving the transfer of dysfunctional mitochondria from astrocytes to neurons. The uptake of these compromised mitochondria by striatal neurons results in reduced neuronal branching and increased reactive oxygen species (ROS) production. Collectively, these results highlight the pathological relevance of impaired astrocyte-to-neuron mitochondrial transfer and emphasize the contributory role of astrocytic dysfunction in Huntington´s disease progression.

Keywords:  Astrocytes; Huntingtin; Mitochondria transfer; Neuroglial communication; R6/1 mice; Striatum

DOI:  https://doi.org/10.1186/s12964-025-02341-6