bims-axbals Biomed News
on Axonal biology and ALS
Issue of 2026–03–15
eighteen papers selected by
TJ Krzystek
  1. ALS and Huntington Disease: Unraveling the Connections between TDP-43 and Huntingtin.
  2. Cofilin hyperphosphorylation triggers TDP-43 pathology in sporadic amyotrophic lateral sclerosis.
  3. cGAS inhibition delays TDP-43-driven ALS Pathogenesis.
  4. Ubiquitin-specific peptidase-19 links TDP-43 aggregation to ER stress.
  5. APP Deficiency Ameliorates FAD Presenilin 1 F105C and A246E Mutations-induced Mitochondrial Dysfunction in Human Cortical Neurons.
  6. Robust activity-dependent mitochondrial calcium dynamics at the AIS is dispensable for action potential generation.
  7. Multi-modal dissection of cell-type specific TDP-43 pathology in the motor cortex.
  8. Decoding the functions of nuclear speckles in neurodegeneration.
  9. Fructose-2,6-bisphosphate restores TDP-43 pathology-driven genome repair deficiency in motor neuron diseases.
  10. Spastin Is Required to Prevent SPAST-Related Demyelination.
  11. SGK1 inhibition restores excitability of cortical neurons derived from Alzheimer's disease patients.
  12. Effect of G4C2repeat expansions on the motion of lysosomes inside neurites.
  13. A Novel Model System to Identify Cellular and Molecular Defects Underlying Rare Genetic Disorders.
  14. Modeling diseases of aging in larval zebrafish, a paradoxical yet powerful strategy.
  15. Quantifying Lysosomal Degradation of Extracellular Proteins With a Fluorescent Protein-Based Internalization Assay.
  16. Live imaging of neuronal dynamics in transparent mouse brains.
  17. Impaired α-Synuclein aggregate clearance in neuronal cells drive their spread to microglia through tunneling nanotubes.
  18. NeuronID: An automatic toolkit for identifying neurons in two-photon calcium imaging data.
  1. J Neurosci. 2026 03 11. pii: e0263252026. [Epub ahead of print]46(10):
    ALS and Huntington Disease: Unraveling the Connections between TDP-43 and Huntingtin.
    Cailyn M Perry, Dale D O Martin.
      Amyotrophic lateral sclerosis (ALS) and Huntington disease (HD) are lethal neurodegenerative diseases affecting motor function. Though their etiology and pathology are distinct, recent evidence suggests commonalities between TAR DNA-binding protein (TDP-43), which is associated with 97% of ALS cases, and huntingtin (HTT), the causative protein of HD. ALS is a heterogeneous, lethal neurodegenerative disease characterized by the progressive loss of upper and lower motor neurons, as well as brainstem and spinal cord degeneration. The causes of ALS are complex, variable, and, in some cases, unknown, but most cases involve mislocalization of the protein TDP-43. In contrast, HD is a monogenic, autosomal dominant, lethal neurodegenerative disease caused by polyglutamine expansion in HTT protein and characterized by the progressive loss of neurons in the brain, particularly in the striatum, which results in motor, cognitive, and behavioral changes. Although HD is not typically associated with motor neuron loss, recent evidence suggests a link between HTT and TDP-43 within the context of both ALS and HD, as well as links to related neurodegenerative diseases, such as frontotemporal dementia (FTD) and spinocerebellar ataxia type 2 (SCA2). Herein, we discuss confirmed cases of concurrent ALS and HD and the overlap of underlying disease mechanisms that potentially contribute to the onset and progression of these two devastating neurodegenerative diseases, with a focus on commonalities between TDP-43 and HTT. We propose that elucidating these commonalities will aid in the identification of broad-spectrum disease risk factors and potential overlapping treatment targets.
    DOI:  https://doi.org/10.1523/JNEUROSCI.0263-25.2026
  2. Brain. 2026 Mar 10. pii: awag096. [Epub ahead of print]
    Cofilin hyperphosphorylation triggers TDP-43 pathology in sporadic amyotrophic lateral sclerosis.
    Cyril Jones Jagaraj, Sayanthooran Saravanabavan, Sonam Parakh, Mrithulabashini Jayakumar, Sara Assar Kashani, Sina Shadfar, Prachi Mehta, Shashi Gautam, Fabiha Farzana, Alexandra K Suchowerska, Kuok Yap, Marta Vidal, Audrey M G Ragagnin, Md Shafi Jamali, Shu Yang, Thomas Fath, David Craik, Julie D Atkin.
      Pathological forms of TAR-binding protein 43 (TDP-43), involving its aberrant mislocalization to the cytoplasm, inclusion formation, hyperphosphorylation and fragmentation, are present in ∼45-50% frontotemporal dementia (FTD) and Alzheimer's disease individuals, and most (97%) amyotrophic lateral sclerosis (ALS) cases. Hence, identifying mechanisms that induce TDP-43 pathology are central to neurodegeneration and developing new therapeutic targets in these conditions. Cofilin is a multi-functional protein with a crucial role in regulating the actin cytoskeleton. Actin has important neuronal-specific activities in dendritic spines, axonal growth cones and synapses and it is in constant equilibrium between two forms: monomeric globular actin (G-actin) and polymeric filamentous actin (F-actin). Cofilin controls actin dynamics by depolymerising and severing actin filaments. When cofilin is phosphorylated (at Serine-3) by LIM kinase1 (LIMK1), it becomes inactive, leading to production of more F-actin. Defects in cofilin are well described in other neurodegenerative disorders, unlike in ALS. We examined phosphorylation of cofilin and actin dynamics in post-mortem spinal cord tissue from sporadic ALS (SALS) patients, the TDP-43 rNLS8 transgenic mouse model, and NSC34 motor neuronal cells expressing cytoplasmic TDP-43. F-actin was pharmacologically stabilized to mimic cofilin hyperphosphorylation, and TDP-43 pathology was assessed. Neuronal cells were treated with a non-phosphorylatable cofilin S3A peptide (MAAGVAVSDGVIKVFN), and TDP-43 pathology and apoptosis were evaluated. Here, we show that cofilin is hyper-phosphorylated in human ALS and disease models compared to controls. This was detected in spinal motor neurons from sporadic ALS (SALS) patients and a TDP-43 mouse model (rNLS8) displaying key ALS phenotypes, and in motor neuronal NSC34-cells expressing cytoplasmic TDP-43. Supporting this observation, more F-actin relative to G-actin was present in cortical/spinal cord lysates from SALS patients and TDP-43 rNLS8 mice, and NSC34-cells expressing TDP-43. We also show that mimicking cofilin hyperphosphorylation by pharmacological stabilization of F-actin induced TDP-43 pathology: cytoplasmic mislocalization, inclusion formation, hyperphosphorylation, and fragmentation, and promoted its recruitment into stress granules (SGs). Furthermore, we detected increased levels of LIMK1 phosphorylation and tropomyosin isoforms 4.1 and 4.2 in SALS patients. These findings reveal aberrant cofilin hyperphosphorylation disrupts actin dynamics, triggering TDP-43 pathology and SG recruitment in SALS. They imply that preventing cofilin phosphorylation is a novel therapeutic strategy applicable to most ALS cases. Treatment of neuronal cells with the S3A peptide prevented features of TDP-43 pathology and apoptosis compared to control peptides. These findings thus describe a novel pathogenic mechanism producing TDP-43 pathology, applicable to most ALS cases and other neurodegenerative diseases.
    Keywords:  ALS/MND; LIMK1 dysregulation; TDP-43 pathology; actin dysregulation; cofilin hyperphosphorylation and dysregulation
    DOI:  https://doi.org/10.1093/brain/awag096
  3. bioRxiv. 2026 Feb 26. pii: 2026.02.24.707791. [Epub ahead of print]
    cGAS inhibition delays TDP-43-driven ALS Pathogenesis.
    Yajing Liu, Weixi Feng, Abulimiti Aikedan, Se-In Lee, Maitreyee Bhagwat, Ravi Kumar Nagiri, Man Ying Wong, Sadaf Amin, Wenhui Qu, Jingjie Zhu, Si-Yu Wang, Pearly Ye, Kendra Norman, Guillermo Coronas-Samano, Marta Olah, Hagen U Tilgner, Li Fan, Subhash C Sinha, Li Gan.
      Amyotrophic lateral sclerosis (ALS) is a fatal neurodegenerative disorder marked by motor neuron loss and cytoplasmic mislocalization of TAR DNA-binding protein 43 (TDP-43), a key regulator of RNA splicing. However, the upstream modulators of this process remain poorly defined. Here we identify cyclic GMP-AMP synthase (cGAS) as a central mediator of TDP-43 pathology and associated mis-splicing. cGAS expression was elevated in ALS patient brains and enriched across activated microglia. In human iPSC-derived microglia-motor neuron co-cultures, neuronal TDP-43 pathology triggered microglial cGAS activation, whereas pharmacological inhibition with a potent human cGAS inhibitor reduced phosphorylated TDP-43, restored lysosomal and phagocytic programs, normalized microglial reactivity, and reversed TDP-43-associated RNA splicing defects. In vivo, cGAS inhibition in TDP-43 Q331K mice reversed widespread RNA splicing abnormalities across neurons and oligodendrocyte lineage cells, attenuated neurodegenerative pathology, and preserved motor function. Together, these findings identify cGAS as a druggable upstream regulator linking innate immune signaling to TDP-43-dependent RNA mis-splicing and neurodegeneration, and establish cGAS inhibition as a promising therapeutic strategy for ALS.
    DOI:  https://doi.org/10.64898/2026.02.24.707791
  4. Proc Natl Acad Sci U S A. 2026 Mar 17. 123(11): e2514355123
    Ubiquitin-specific peptidase-19 links TDP-43 aggregation to ER stress.
    Yan Yan, Xinming Wang, Hanna Jeon, Teresa R Kee, Khoi D Tran, Hyunkyoung Lee, Xingyu Zhao, Tian Liu, Simon S Wing, Jung-A A Woo, David E Kang.
      Aggregation and deposition of TAR DNA-binding protein 43 (TDP-43) is a salient pathological signature of amyotrophic lateral sclerosis (ALS) and frontotemporal lobar degeneration-TDP (FTLD-TDP). TDP-43 proteostasis and aggregation are controlled by several posttranslational modifications, including ubiquitination. While multiple E3 ubiquitin ligases are known to facilitate TDP-43 clearance, little is known about the role of deubiquitinases (DUBs) in controlling TDP-43 proteostasis. Through an unbiased discovery screen of DUBs, here we identify and demonstrate using in vitro and in vivo models, as well as human brain tissue, that ubiquitin-specific peptidase-19 (USP19) acts as a TDP-43-directed DUB that removes K48- and K63-linked ubiquitin conjugates from TDP-43 and preferentially promotes cytoplasmic aggregation of TDP-43 C-terminal fragments (TDP-CTFs) through its catalytic activity. Specifically, the endoplasmic reticulum (ER)-anchored USP19 isoform (USP19-ER) exhibits superior activity in deubiquitinating TDP-CTFs, enhancing its phase separation and aggregation, compared to its cytosolic isoform (USP19-Cyto). Furthermore, as TDP-CTFs are generated at the ER, USP19 acts to couple the aggregation of TDP-CTFs to ER stress (ATF6, ATF4, IRE1, & CHOP). In humans, USP19 protein levels increase in FTLD-TDP brains, which extensively colocalize with cytoplasmic phospho-TDP-43 (pTDP-43) pathology. Importantly, we demonstrate in vivo that genetic reduction of usp19 mitigates pTDP-43 pathology, astrogliosis, and ER stress while reversing long-term potentiation (LTP) and motor deficits in a mouse model of TDP-43 pathogenesis (TAR4 mice). These findings establish a critical role of USP19 at the nexus of TDP-43 proteostasis and ER stress, implicating its pathogenic role in FTLD-TDP and ALS.
    Keywords:  ALS; ER stress; FTD; TDP-43; USP19
    DOI:  https://doi.org/10.1073/pnas.2514355123
  5. Int J Biol Sci. 2026 ;22(5): 2720-2735
    APP Deficiency Ameliorates FAD Presenilin 1 F105C and A246E Mutations-induced Mitochondrial Dysfunction in Human Cortical Neurons.
    Yu-Hsin Yen, Fang Yuan, Daijiao Tang, Jing-Fang Luo, Chen Ming, Phil-Jun Kang, Huanxing Su, Cheong-Meng Chong, Su-Chun Zhang.
       Background: Mitochondrial dysfunction is widely regarded as a central and early feature of Alzheimer's disease (AD) pathology. Prior studies suggest that the accumulation of amyloid precursor protein (APP) within mitochondria contributes to this dysfunction. Mutations in presenilin-1 (PS1), which account for most cases of early-onset familial AD (FAD), have also been shown to impair mitochondrial function. In this study, we investigated how APP influences PS1 mutation-induced mitochondrial dysfunction in human cortical neurons derived from patient induced pluripotent stem cells (iPSCs).
    Methods: We analyzed transcriptomic and proteomic datasets from postmortem sporadic AD cortex to identify key dysregulated pathways. To functionally interrogate selected mechanisms, we established a panel of CRISPR/Cas9-engineered human iPSC lines, including PS1 mutant lines (PS1+/F105C and PS1+/A246E), an APP knockout derivative (APP-/-_PS1+/F105C), and their isogenic wild-type controls. These iPSCs were differentiated into cortical neurons for functional studies. Following directed differentiation into cortical neurons, biochemical analyses and super-resolution imaging were conducted to evaluate mitochondrial and neuronal phenotypes.
    Results: Analyses of sporadic AD cortical transcriptomes and proteomes identified mitochondrial dysfunction as a prominently altered pathway. In agreement, cortical neurons differentiated from FAD PS1 mutant (F105C and A246E) iPSCs displayed mitochondrial defects and AD-related phenotypes, both of which were mitigated by APP knockout.
    Conclusions: These findings provide critical insights into the bridging role of APP in FAD PS1 mutant-mediated mitochondrial dysfunction, advancing our understanding of the cellular mechanisms underlying AD.
    Keywords:  Alzheimer's disease; CRISPR; amyloid precursor protein; iPSCs; mitochondrial dysfunction; presenilin 1
    DOI:  https://doi.org/10.7150/ijbs.120062
  6. J Physiol. 2026 Mar 10.
    Robust activity-dependent mitochondrial calcium dynamics at the AIS is dispensable for action potential generation.
    Koen Kole, Maarten H P Kole.
      Mitochondria are diverse and multifaceted intracellular organelles regulating oxidative energy supply, lipid metabolism and calcium (Ca2+) signalling. In neurons the spatial sequestration of cytoplasmic Ca2+ by mitochondria plays a critical role in determining activity-dependent spine plasticity, shaping the presynaptic transmitter release characteristics and contributing to sustained action potential firing. Here, we tested the hypothesis that mitochondria at the axon initial segment (AIS) affect the microdomain cytoplasmic Ca2+ transients, thereby regulating Ca2+-dependent voltage-gated ion channels at the plasma membrane and initiation of action potentials. Using 3D electron microscopy reconstructions and virally injecting genetically encoded fluorescence indicators we visualized the ultrastructure and distribution of mitochondria selectively in thick-tufted layer 5 pyramidal neurons. We found that most mitochondria were stably clustered to the proximal AIS, while few were observed at distal sites. Simultaneous two-photon imaging of action potential-dependent cytoplasmic and mitochondrial Ca2+, combined with electrophysiological recordings showed that AIS mitochondria exhibit powerful activity-dependent cytosolic Ca2+ uptake. However, while intracellular application of the mitochondrial Ca2+ uniporter inhibitor Ru360 fully blocked mitochondrial Ca2+ import and increased the slow afterhyperpolarization duration, it did not affect action potential input-output function, action potential dynamics nor the ability to produce high-frequency burst output. Together, the results indicate that AIS mitochondria are dispensable for temporal and rate encoding, suggesting that mt-Ca2+ buffering at the AIS may be involved in non-electrical roles. KEY POINTS: Mitochondrial Ca2+ buffering controls multiple Ca2+-dependent intracellular processes and their subcellular location of the organelles defines local physiological properties in neurons. Recent studies implicate mitochondrial Ca2+ uptake in the slow afterhyperpolarization and maintenance of action potential firing. Using electron microscopy and virally delivered genetically encoded tools we examined mitochondria in the layer 5 pyramidal neuron axon initial segment (AIS), the site where action potentials initiate, and found that cytoplasmic Ca2+ influx is powerfully buffered by proximally clustered mitochondria. Electrophysiological recordings during the block of the mitochondrial calcium uniporter reveal a role in the slow afterhyperpolarization, while AIS action potential initiation and action potential waveforms are independent from mitochondria. These findings indicate AIS mitochondria under physiological conditions exert non-electrical roles.
    Keywords:  action potential; axon initial segment; calcium buffer; mitochondria; pyramidal neuron
    DOI:  https://doi.org/10.1113/JP289290
  7. Nat Commun. 2026 Mar 09. pii: 2406. [Epub ahead of print]17(1):
    Multi-modal dissection of cell-type specific TDP-43 pathology in the motor cortex.
    Wolfgang P Ruf, Julia K Kühlwein, Laura Meier, Sarah J Brockmann, Jaehyun LeeBae, Ghazaleh Sadri-Vakili, Deniz Yilmazer-Hanke, Susanne Petri, Dietmar R Thal, Veselin Grozdanov, Karin M Danzer.
      Cytoplasmic TDP-43 pathology is a pathological sign of ALS/ALS-FTD and a converging disease event across different genotypes, phenotypes and CNS areas. To understand this process and target it therapeutically, we need to define which cell types are affected and which cell-type specific effects make them particularly vulnerable. We coupled flow-cytometry nuclear sorting and sequencing with single-nucleus multi-omic ATAC-seq and RNA-seq and spatial transcriptomics to define the transcriptional cell type of affected neurons in the post-mortem ALS/ALS-FTD motor cortex (30 ALS, 20 ALS-FTD & 32 control samples). Here, we show that mainly excitatory cortical neurons are affected by TDP-43 pathology and define the cell types that are affected the most: intratelencephalic L2-L3-LINC00507-FREM3, L3-L5-RORB-LNX2, L3-L5-RORB-ADGRL4 & L6-THEMIS-LINC00343 neurons and extratelencephalic L5-FEZF2-NTNG1 neurons. Transcriptional aberrations by TDP-43 pathology, like cryptic exon inclusion, are cell-type specific and affect distinct gene sets in each cell type, highlighting the need to address TDP-43 pathology in a cell-type specific manner.
    DOI:  https://doi.org/10.1038/s41467-026-69944-6
  8. Trends Neurosci. 2026 Mar 06. pii: S0166-2236(26)00013-5. [Epub ahead of print]
    Decoding the functions of nuclear speckles in neurodegeneration.
    Xu Chen, Bokai Zhu.
      Nuclear speckles, traditionally considered mainly as reservoirs of splicing factors, are increasingly recognized as dynamic biomolecular condensates essential for RNA metabolism, transcriptional regulation, and chromatin organization. Recent advances reveal their phase separation properties, compositional complexity, and stress-responsive remodeling, positioning nuclear speckles as key regulators of proteostasis and stress adaptation. Here, we synthesize emerging evidence linking nuclear speckle dysfunction to neurodegenerative proteinopathies, particularly amyotrophic lateral sclerosis (ALS)/frontotemporal dementia (FTD) and tauopathies. We highlight how disease-associated repeat RNAs, dipeptide repeat proteins, and hyperphosphorylated tau disrupt nuclear speckle integrity, driving transcriptional and splicing defects. Finally, we discuss therapeutic strategies to rejuvenate nuclear speckles, emphasizing their potential as novel targets for restoring proteostasis and mitigating neurodegeneration. This review underscores nuclear speckles as critical yet underexplored regulators of neuronal resilience.
    Keywords:  ALS/FTD; RNA metabolism; cellular stress response; nuclear condensates; proteinopathy; tauopathy
    DOI:  https://doi.org/10.1016/j.tins.2026.01.010
  9. Commun Biol. 2026 Mar 10.
    Fructose-2,6-bisphosphate restores TDP-43 pathology-driven genome repair deficiency in motor neuron diseases.
    Anirban Chakraborty, Joy Mitra, Vikas H Malojirao, Manohar Kodavati, Santi M Mandal, Satkarjeet K Gill, Sravan Gopalkrishnashetty Sreenivasmurthy, Velmarini Vasquez, Mikita Mankevich, Ludo Van Den Bosch, Ralph M Garruto, Ian Robey, Balaji Krishnan, Gourisankar Ghosh, Muralidhar L Hegde, Tapas Hazra.
      TDP-43 proteinopathy is central to amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD). TDP-43 plays a key role in DNA double-strand break repair (DSBR), though the underlying mechanisms remain unclear. Here, we demonstrate that ALS patients' brains exhibit persistent DNA damage within transcribed genes. Mechanistically, activity of polynucleotide kinase 3'-phosphatase (PNKP), an essential DNA end-processing enzyme required for DSBR in transcribed genes, is impaired in ALS brains and TDP-43-depleted cells. Such defect stems from reduced levels of PNKP-interacting enzyme phosphofructo-2- kinase/fructose-2,6-bisphosphatase 3 (PFKFB3) and its metabolic product fructose-2,6- bisphosphate (F2,6BP), an essential cofactor of PNKP. F2,6BP supplementation reduces cytosolic aggregation of phosphorylated and polyubiquitinated TDP-43 in patient-derived induced neurons, rescues PNKP activity in ALS/FTD brain extracts, and improves motor deficits in Drosophila TDP-43 model. Together, these findings reveal a critical link between metabolic dysregulation and genomic instability in TDP-43 pathology-associated motor neuron diseases, and underscore therapeutic potential of F2,6BP.
    DOI:  https://doi.org/10.1038/s42003-026-09787-5
  10. J Neurochem. 2026 Mar;170(3): e70407
    Spastin Is Required to Prevent SPAST-Related Demyelination.
    Şeyma Akarsu, Didem Müge Orhan, Timuçin Avşar, Arzu Karabay.
      Mutations in the SPAST gene, encoding the microtubule-severing protein Spastin, cause the most common type of hereditary spastic paraplegia (HSP): SPG4, a disorder primarily characterized by length-dependent axonal degeneration. Clinically, most SPG4 patients present with a pure phenotype marked by progressive spasticity in the lower extremities. It has also been reported that complex cases exhibit demyelination and cognitive deficits. Additionally, some SPAST variants have been determined in patients with multiple sclerosis (MS), indicating potential shared pathological mechanisms. Spastin is known to promote axonal regeneration by remodeling microtubules, whereas mutant Spastin disrupts microtubule dynamics and causes axonal transport defects in SPG4. However, whether Spastin dysfunction impairs regenerative processes such as myelination remains unknown. In this study, we investigated whether the SPG4-associated SPAST mutations affect axonal myelination. Using an in vitro cortical neuron-oligodendrocyte co-culture model, we found that pathogenic SPAST mutations result in a significant reduction in the myelination index. Furthermore, a cuprizone-induced demyelination mouse model revealed a decrease in Spastin protein levels in demyelinated white matter. Given Spastin's role in axonal regeneration, we hypothesized that Spastin may also protect against demyelination. Supporting this, wild-type Spastin expression protected neurons from demyelination in a cuprizone-induced cell culture demyelination model. Together, these results suggest a role for Spastin in axonal myelination, and its dysfunction may compromise myelin stability. Our findings highlight that impaired myelin stability may represent a secondary pathological feature of SPG4, contributing to disease complexity. This dual role of Spastin in axonal maintenance and myelin stability suggests its potential relevance for contributing to complex forms of SPG4.
    Keywords:  Spastin; cuprizone; hereditary spastic paraplegia 4; multiple sclerosis; myelin
    DOI:  https://doi.org/10.1111/jnc.70407
  11. Neurobiol Dis. 2026 Mar 05. pii: S0969-9961(26)00089-6. [Epub ahead of print]221 107345
    SGK1 inhibition restores excitability of cortical neurons derived from Alzheimer's disease patients.
    Zhongjiao Jiang, Komal Saleem, Emily Fisher, Li Li, Zhen Yan, Jian Feng.
      The vast majority of Alzheimer's disease (AD) cases are sporadic, without a clear etiology. We have previously found increased expression of Serum and Glucocorticoid-regulated Kinase 1 (SGK1) in mouse models of dementia, postmortem cortical tissues and induced pluripotent stem cells (iPSCs)-derived cortical neurons from patients with sporadic AD (sAD). SGK1 is induced by a variety of cellular stress. The physiological consequences of elevated SGK1 in sAD is unclear. Here, we differentiated iPSCs from four sAD patients and four age- and sex-matched healthy controls into electrophysiologically mature cortical neurons with prolonged culture for more than 100 days. The sAD cortical neurons exhibited significant reductions in voltage-gated Na+ currents, amplitudes of evoked action potentials, and frequencies of spontaneous excitatory postsynaptic currents and spontaneous action potentials. Application of a selective inhibitor of SGK1 reversed all these phenotypes in sAD neurons without affecting control neurons. The SGK1-dependent hypoexcitability suggests that a convergent and inborn mechanism attenuates neuronal communications despite different genetic background of the sAD patients. Their iPSC-derived cortical neurons have captured the defective neurotransmission, which underlies cognitive and memory symptoms of AD, many decades before clinical manifestations. The study offers a new pathway to restore synaptic transmission in AD.
    Keywords:  Alzheimer's disease; Cortical neurons; Induced pluripotent stem cells; Neuronal excitability; SGK1; Synaptic transmission
    DOI:  https://doi.org/10.1016/j.nbd.2026.107345
  12. Phys Biol. 2026 Mar 12.
    Effect of G4C2repeat expansions on the motion of lysosomes inside neurites.
    Maria Mytiliniou, Joeri A J Wondergem, Marleen Feliksik, Thomas Schmidt, Thomas Fuhs, Doris Heinrich.
      The G4C2hexanucleotide repeat expansion in the c9orf72 locus is a mutation associated with amyotrophic lateral sclerosis. Recent evidence suggests a link with disrupted axonal trafficking in neurons. Here, using a neuronal-like cell line without or transfected with G4C2repeats, we characterize the motion of lysosomes inside neurites. The neurites grew either aligned to patterned lines, or oriented freely on a 2D-substrate. Implementing time-resolved (local) mean squared displacement analysis lysosome trajectories were split into sub-diffusive, diffusive, and super-diffusive parts. Our results suggest that in the presence of the G4C2repeats, lysosome trafficking is hampered, exhibiting overall decreased mean squared displacement and speed, more prominently inside aligned neurites. Moreover, a prominent effect in the super-diffusive drift velocity and diffusive motion diffusion coefficient was evident when the motion occurred inside aligned neurites. Trajectories which included super-diffusive motion, exhibited a varied ratio of anterograde/retrograde/neutral for both neurite geometries in the presence of G4C2repeats but a similar velocity decrease for both directions in each neurite geometry. Our findings support the hypothesis that impaired axonal trafficking emerges in the presence of the G4C2hexanucleotide repeat expansion, and demonstrate that this effect is more prominent when the neurites are aligned.
    Keywords:  G4C2 hexanucleotide repeat expansion; Lysosome; PC12 Neurite; amyotrophic lateral sclerosis; axonal trafficking; c9orf72 locus
    DOI:  https://doi.org/10.1088/1478-3975/ae5143
  13. Exp Dermatol. 2026 Mar;35(3): e70238
    A Novel Model System to Identify Cellular and Molecular Defects Underlying Rare Genetic Disorders.
    Maddison N Salois, Saiphone Webb, Isaiah A Proctor, Peter J Koch, Maranke I Koster.
      Ankyloblepharon-ectodermal defects-cleft lip/palate (AEC) is a disorder caused by autosomal-dominant mutations in the TP63 gene. AEC is characterised by the presence of severe and painful skin erosions that can take years to heal. Current treatment options for these devastating lesions are limited, highlighting the need for new therapeutic strategies. We previously generated keratinocytes from patient-derived induced pluripotent stem cells (iPSC-K) and identified defects in several cell adhesion complexes, including desmosomes, hemidesmosomes and focal adhesions. In the present study, we developed a complementary in vitro model using NTERT keratinocytes transduced with lentiviral constructs expressing AEC-related TP63 mutations (N-AEC). This model allows for the large-scale production of disease-relevant material, overcoming the limitations of iPSC-derived keratinocytes, which have the characteristics of primary keratinocytes, including limited cell doublings and lifespan. We demonstrate that N-AEC keratinocytes exhibit key defects observed in AEC iPSC-K and AEC patient skin, including downregulation of cell adhesion proteins. In addition, 3D epidermal equivalents generated from these cells replicate pathological features seen in AEC patient skin, such as intra-epidermal cysts, reduced desmosomal protein expression and altered expression of differentiation markers. Our N-AEC model provides a valuable tool for investigating the mechanisms underlying skin fragility in AEC and other genetic skin disorders and advances the potential for novel therapeutic development.
    DOI:  https://doi.org/10.1111/exd.70238
  14. Genetics. 2026 Mar 10. pii: iyag027. [Epub ahead of print]
    Modeling diseases of aging in larval zebrafish, a paradoxical yet powerful strategy.
    Güliz Gürel Özcan, Jason Rihel.
      Neurodegenerative diseases are a set of devastating medical conditions in which neuronal loss associated with the aggregation of toxic proteins leads to progressive cognitive impairment. These diseases are usually modeled in animals by mimicking late disease stages through genetic modifications that aggressively accumulate proteins that damage the brain. However, these diseases typically unfold over decades, and disease-associated genes are known to have important, but understudied, biological functions in early life stages. To address this research gap, we suggest that the larval zebrafish, which has conserved orthologs of most neurodegeneration-linked genes, is an excellent model to examine early mechanisms that set the stage for disease progression, such as altered neuronal function, synaptic re-wiring, and proteostasis. We propose a systematic genetic modeling and phenotyping pipeline in zebrafish that integrates CRISPR editing, high-throughput behavioral assays, brain-wide activity mapping, and pharmacological screens to capture neurodegenerative disease-related changes that occur well before clinical disease emerges. Studying diseases of aging in larval zebrafish may sound paradoxical; however, by uncovering cellular dysfunction at the earliest stages of disease in a living vertebrate brain, this approach could identify critical therapeutic targets at timepoints before degeneration becomes irreversible.
    Keywords:  ALS; Alzheimer's disease; Parkinson's disease; behavioral fingerprinting; behavioral pharmacology; brain-wide activity mapping; genetic disease modeling; neurodegeneration; zebrafish
    DOI:  https://doi.org/10.1093/genetics/iyag027
  15. Bio Protoc. 2026 Mar 05. 16(5): e5619
    Quantifying Lysosomal Degradation of Extracellular Proteins With a Fluorescent Protein-Based Internalization Assay.
    Sayana Bun, Kanna Kamikawa, Akira Matsuura, Eisuke Itakura.
      Endocytosis is an essential membrane transport mechanism that is indispensable for the maintenance of life. It is responsible for the selective internalization and subsequent degradation or recycling of specific extracellular proteins and nutrients, thereby facilitating cellular nutrient supply, modulation of receptor signaling, and clearance of foreign substances. However, methods for the quantitative analysis of lysosomal degradation of extracellular proteins via endocytosis remain limited. This protocol describes a method for purifying the protein-of-interest (POI)-red fluorescent protein (RFP)-green fluorescent protein (GFP) fusion protein, which is modified with specific mammalian cell glycans or other modifications, from the conditioned medium of mammalian cell cultures. Subsequently, the protocol details a quantitative approach for evaluating its internalization and lysosomal degradation within cells using the RFP-GFP tandem fluorescent reporter. Following the addition of POI-RFP-GFP to the medium, cells can be subjected to cell biological assays, such as flow cytometry, as well as biochemical analyses, such as immunoblotting. This protocol is broadly applicable to studies of the internalization of extracellular proteins. Key features • Purification of secreted GFP-RFP-fused POI from mammalian cell culture supernatant. • Quantification of POI-RFP-GFP internalization through measurement of GFP and RFP signals using flow cytometry. • Confirmation of lysosomal degradation of POI-RFP-GFP by immunoblotting.
    Keywords:  Endocytosis; Extracellular protein; Internalization; Lysosome; Protein degradation; Proteostasis; Secreted protein
    DOI:  https://doi.org/10.21769/BioProtoc.5619
  16. Nat Methods. 2026 Mar 12.
    Live imaging of neuronal dynamics in transparent mouse brains.
      
    DOI:  https://doi.org/10.1038/s41592-026-03022-z
  17. Nat Commun. 2026 Mar 12.
    Impaired α-Synuclein aggregate clearance in neuronal cells drive their spread to microglia through tunneling nanotubes.
    Ranabir Chakraborty, Francesca Palese, Philippa Samella, Veronica Testa, Jara Montero-Muñoz, Sylvie Syan, Takashi Nonaka, Masato Hasegawa, Antonella Consiglio, Chiara Zurzolo.
      Tunneling nanotubes (TNTs) play a crucial role in intercellular communication, enabling transfer of molecular cargoes over long distances between connected cells. Previous studies have demonstrated efficient, directional transfer of α-Synuclein (α-Syn) aggregates from neurons to microglia, with endosomal trafficking and lysosomal processing identified as the primary events following α-Syn internalization. Using human neuronal and microglial cell lines, we show that microglia exhibit higher lysosomal turnover, particularly through lysophagy, whereas neuronal lysosomes display compromised degradative capacity and impaired autophagic flux upon α-Syn exposure, resulting in compromised aggregate clearance. Such a response to α-Syn aggregates is also conserved in human iPSC-derived neurons and microglia. Moreover, perturbing aggregate clearance via autophagy inhibition enhances TNT-mediated transfer of α-Syn from neuronal cells to microglia. Microglia co-cultured with α-Syn-containing neurons upregulate autophagy flux, enabling efficient degradation of the transferred aggregates. These results highlight dysfunctional autophagy in neurons as a key driver outsourcing α-Syn aggregates to microglia.
    DOI:  https://doi.org/10.1038/s41467-026-69930-y
  18. PLoS One. 2026 ;21(3): e0343516
    NeuronID: An automatic toolkit for identifying neurons in two-photon calcium imaging data.
    Jikan Peng, Tian Xu.
      Two-photon calcium imaging has emerged as a powerful technique for monitoring neuronal activity in neuroscience; however, its data processing remains challenging. Here, we introduce NeuronID, an automatic toolkit designed to process two-photon calcium imaging data. The NeuronID toolkit features a modular architecture that includes motion correction, noise reduction, segmentation of neuronal components, and extraction of neuronal signals. Notably, the NeuronID toolkit offers an optimized strategy for segmenting neuronal components, which systematically integrates morphological boundary identification, cross-correlation analysis between pixels, and evaluation of neuronal signal quality. Compared to existing tools or manual annotation by experts, the NeuronID toolkit reduces the likelihood of over-segmentation while achieving near-human accuracy. Overall, this study provides an effective solution to the segmentation of neuronal components, offering a standardized analytical tool for processing two-photon calcium imaging data.
    DOI:  https://doi.org/10.1371/journal.pone.0343516
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