bims-axbals 2024-09-01 papers

bims-axbals

Biomed News

on Axonal biology and ALS

Issue of 2024–09–01
eightteen papers selected by
TJ Krzystek, ALS Therapy Development Institute

Disruption of nuclear speckle integrity dysregulates RNA splicing in C9ORF72-FTD/ALS.
KIF1A, R1457Q, and P1688L Mutations Induce Protein Abnormal Aggregation and Autophagy Impairment in iPSC-Derived Motor Neurons.
Single acetylation-mimetic mutation in TDP-43 Nuclear Localization Signal disrupts importin α1/β signaling.
The RNA-binding protein EIF4A3 promotes axon development by direct control of the cytoskeleton.
Dysregulation of muscle cholesterol transport in amyotrophic lateral sclerosis.
HiPSC-derived 3D neural models reveal neurodevelopmental pathomechanisms of the Cockayne Syndrome B.
Pathological characteristics of axons and alterations of proteomic and lipidomic profiles in midbrain dopaminergic neurodegeneration induced by WDR45-deficiency.
Inflammatory signature in amyotrophic lateral sclerosis predicting disease progression.
Neurofilaments in neurologic disease.
Regulation of TIR-1/SARM-1 by miR-71 Protects Dopaminergic Neurons in a C. elegans Model of LRRK2-Induced Parkinson's Disease.
RILP Induces Cholesterol Accumulation in Lysosomes by Inhibiting Endoplasmic Reticulum-Endolysosome Interactions.
Botulinum toxin intoxication requires retrograde transport and membrane translocation at the ER in RenVM neurons.
A neural network for religious fundamentalism derived from patients with brain lesions.
Experimental study on small molecule combinations inducing reprogramming of rat fibroblasts into functional neurons.
Machine Learning-Enhanced Estimation of Cellular Protein Levels from Bright-Field Images.
Axonal mitochondria regulate gentle touch response through control of axonal actin dynamics.
Stem cells for organoids.
Neuritin controls axonal branching in serotonin neurons: A possible mediator involved in the regulation of depressive and anxiety behaviors via FGF signaling.

Neuron. 2024 Aug 17. pii: S0896-6273(24)00570-1. [Epub ahead of print]

Disruption of nuclear speckle integrity dysregulates RNA splicing in C9ORF72-FTD/ALS.

Rong Wu, Yingzhi Ye, Daoyuan Dong, Zhe Zhang, Shaopeng Wang, Yini Li, Noelle Wright, Javier Redding-Ochoa, Koping Chang, Shaohai Xu, Xueting Tu, Chengzhang Zhu, Lyle W Ostrow, Xavier Roca, Juan C Troncoso, Bin Wu, Shuying Sun.

  Expansion of an intronic (GGGGCC)n repeat within the C9ORF72 gene is the most common genetic cause of both frontotemporal dementia (FTD) and amyotrophic lateral sclerosis (ALS) (C9-FTD/ALS), characterized with aberrant repeat RNA foci and noncanonical translation-produced dipeptide repeat (DPR) protein inclusions. Here, we elucidate that the (GGGGCC)n repeat RNA co-localizes with nuclear speckles and alters their phase separation properties and granule dynamics. Moreover, the essential nuclear speckle scaffold protein SRRM2 is sequestered into the poly-GR cytoplasmic inclusions in the C9-FTD/ALS mouse model and patient postmortem tissues, exacerbating the nuclear speckle dysfunction. Impaired nuclear speckle integrity induces global exon skipping and intron retention in human iPSC-derived neurons and causes neuronal toxicity. Similar alternative splicing changes can be found in C9-FTD/ALS patient postmortem tissues. This work identified novel molecular mechanisms of global RNA splicing defects caused by impaired nuclear speckle function in C9-FTD/ALS and revealed novel potential biomarkers or therapeutic targets.

Keywords:  ALS; C9ORF72; FTD; RNA splicing; SRRM2; neurodegeneration; nuclear speckle; repeat expansion

DOI:  https://doi.org/10.1016/j.neuron.2024.07.025
Biomedicines. 2024 Jul 30. pii: 1693. [Epub ahead of print]12(8):

KIF1A, R1457Q, and P1688L Mutations Induce Protein Abnormal Aggregation and Autophagy Impairment in iPSC-Derived Motor Neurons.

Mingri Zhao, Junling Wang, Miao Liu, Yaoyao Xu, Jiali Huang, Yiti Zhang, Jianfeng He, Ao Gu, Mujun Liu, Xionghao Liu.

  Mutations in the C-terminal of KIF1A (Kinesin family member 1A) may lead to amyotrophic lateral sclerosis (ALS) through unknown mechanisms that are not yet understood. Using iPSC reprogramming technology and motor neuron differentiation techniques, we generated iPSCs from a healthy donor and two ALS patients with KIF1A mutations (R1457Q and P1688L) and differentiated them into spinal motor neurons (iPSC-MN) to investigate KIF1A-related ALS pathology. Our in vitro iPSC-iMN model faithfully recapitulated specific aspects of the disease, such as neurite fragmentation. Through this model, we observed that these mutations led to KIF1A aggregation at the proximal axon of motor neurons and abnormal accumulation of its transport cargo, LAMP1, resulting in autophagy dysfunction and cell death. RNAseq analysis also indicated that the functions of the extracellular matrix, structure, and cell adhesion were significantly disturbed. Notably, using rapamycin during motor neuron differentiation can effectively prevent motor neuron death.

Keywords:  ALS; KIF1A; iPSC; motor neuron

DOI:  https://doi.org/10.3390/biomedicines12081693
J Mol Biol. 2024 Aug 22. pii: S0022-2836(24)00360-7. [Epub ahead of print] 168751

Single acetylation-mimetic mutation in TDP-43 Nuclear Localization Signal disrupts importin α1/β signaling.

Ying-Hui Ko, Ravi K Lokareddy, Steven G Doll, Daniel P Yeggoni, Amandeep Girdhar, Ian Mawn, Joseph R Klim, Noreen F Rizvi, Rachel Meyers, Richard E Gillilan, Lin Guo, Gino Cingolani.

  Cytoplasmic aggregation of the TAR-DNA binding protein of 43 kDa (TDP-43) is the hallmark of sporadic amyotrophic lateral sclerosis (ALS). Most ALS patients with TDP-43 aggregates in neurons and glia do not have mutations in the TDP-43 gene but contain aberrantly post-translationally modified TDP-43. Here, we found that a single acetylation-mimetic mutation (K82Q) near the TDP-43 minor Nuclear Localization Signal (NLS) box, which mimics a post-translational modification identified in an ALS patient, can lead to TDP-43 mislocalization to the cytoplasm and irreversible aggregation. We demonstrate that the acetylation mimetic disrupts binding to importins, halting nuclear import and preventing importin α 1/ β anti-aggregation activity. We propose that perturbations near the NLS are an additional mechanism by which a cellular insult other than a genetically inherited mutation leads to TDP-43 aggregation and loss of function. Our findings are relevant to deciphering the molecular etiology of sporadic ALS.

Keywords:  ALS; TDP-43; acetylation; importin α 1; importin β; nuclear import; nuclear localization signal

DOI:  https://doi.org/10.1016/j.jmb.2024.168751
Cell Rep. 2024 Aug 23. pii: S2211-1247(24)01017-9. [Epub ahead of print]43(9): 114666

The RNA-binding protein EIF4A3 promotes axon development by direct control of the cytoskeleton.

Fernando C Alsina, Bianca M Lupan, Lydia J Lin, Camila M Musso, Federica Mosti, Carly R Newman, Lisa M Wood, Aussie Suzuki, Mark Agostino, Jeffrey K Moore, Debra L Silver.

  The exon junction complex (EJC), nucleated by EIF4A3, is indispensable for mRNA fate and function throughout eukaryotes. We discover that EIF4A3 directly controls microtubules, independent of RNA, which is critical for neural wiring. While neuronal survival in the developing mouse cerebral cortex depends upon an intact EJC, axonal tract development requires only Eif4a3. Using human cortical organoids, we show that EIF4A3 disease mutations also impair neuronal growth, highlighting conserved functions relevant for neurodevelopmental pathology. Live imaging of growing neurons shows that EIF4A3 is essential for microtubule dynamics. Employing biochemistry and competition experiments, we demonstrate that EIF4A3 directly binds to microtubules, mutually exclusive of the EJC. Finally, in vitro reconstitution assays and rescue experiments demonstrate that EIF4A3 is sufficient to promote microtubule polymerization and that EIF4A3-microtubule association is a major contributor to axon growth. This reveals a fundamental mechanism by which neurons re-utilize core gene expression machinery to directly control the cytoskeleton.

Keywords:  CP: Cell biology; CP: Neuroscience; EIF4A3; Eif4a3; RNA; axon growth; axonal tracts; cortical development; cortical organoids; exon junction complex; microtubules; neural development; neuronal maturation

DOI:  https://doi.org/10.1016/j.celrep.2024.114666
Brain. 2024 Aug 28. pii: awae270. [Epub ahead of print]

Dysregulation of muscle cholesterol transport in amyotrophic lateral sclerosis.

PULSE study group

  Amyotrophic lateral sclerosis (ALS) is a progressive neurodegenerative disorder affecting motor neurons, with a typical lifespan of 3-5 years. Altered metabolism is a key feature of ALS that strongly influences prognosis, with an increase in whole-body energy expenditure and changes in skeletal muscle metabolism, including greater reliance on fat oxidation. Dyslipidemia has been described in ALS as part of the metabolic dysregulation, but its role in the pathophysiology of the disease remains controversial. Among the lipids, cholesterol is of particular interest as a vital component of cell membranes, playing a key role in signal transduction and mitochondrial function in muscle. The aim of this study was to investigate whether motor dysfunction in ALS might be associated with dysregulation of muscle cholesterol metabolism. We determined cholesterol content and analyzed the expression of key determinants of the cholesterol metabolism pathway in muscle biopsies from thirteen ALS patients and ten asymptomatic ALS-mutation gene carriers compared to sixteen controls. Using human control primary myotubes, we further investigated the potential contribution of cholesterol dyshomeostasis to reliance on mitochondrial fatty acid. We found that cholesterol accumulates in the skeletal muscle of ALS patients and that cholesterol overload significantly correlates with disease severity evaluated by the Revised ALS Functional Rating Scale. These defects are associated with overexpression of the genes of the lysosomal cholesterol transporters Niemann-Pick type C1 (NPC1) and 2 (NPC2), which are required for cholesterol transfer from late endosomes/lysosomes to cellular membranes. Most notably, a significant increase in NPC2 mRNA levels could be detected in muscle samples from asymptomatic ALS-mutation carriers, long before disease onset. We found that filipin-stained unesterified cholesterol accumulated in the lysosomal compartment in ALS muscle samples, suggesting dysfunction of the NPC1/2 system. Accordingly, we report here that experimental NPC1 inhibition or lysosomal pH alteration in human primary myotubes was sufficient to induce the overexpression of NPC1 and NPC2 mRNA. Finally, acute NPC1 inhibition in human control myotubes induced a shift towards a preferential use of fatty acids, thus reproducing the metabolic defect characteristic of ALS muscle. We conclude that cholesterol homeostasis is dysregulated in ALS muscle from the presymptomatic stage. Targeting NPC1/2 dysfunction may be a new therapeutic strategy for ALS to restore muscle energy metabolism and slow motor symptom progression.

Keywords:  disease progression; metabolism; presymptomatic stage

DOI:  https://doi.org/10.1093/brain/awae270
Cell Mol Life Sci. 2024 Aug 23. 81(1): 368

HiPSC-derived 3D neural models reveal neurodevelopmental pathomechanisms of the Cockayne Syndrome B.

Julia Kapr, Ilka Scharkin, Haribaskar Ramachandran, Philipp Westhoff, Marius Pollet, Selina Dangeleit, Gabriele Brockerhoff, Andrea Rossi, Katharina Koch, Jean Krutmann, Ellen Fritsche.

  Cockayne Syndrome B (CSB) is a hereditary multiorgan syndrome which-through largely unknown mechanisms-can affect the brain where it clinically presents with microcephaly, intellectual disability and demyelination. Using human induced pluripotent stem cell (hiPSC)-derived neural 3D models generated from CSB patient-derived and isogenic control lines, we here provide explanations for these three major neuropathological phenotypes. In our models, CSB deficiency is associated with (i) impaired cellular migration due to defective autophagy as an explanation for clinical microcephaly; (ii) altered neuronal network functionality and neurotransmitter GABA levels, which is suggestive of a disturbed GABA switch that likely impairs brain circuit formation and ultimately causes intellectual disability; and (iii) impaired oligodendrocyte maturation as a possible cause of the demyelination observed in children with CSB. Of note, the impaired migration and oligodendrocyte maturation could both be partially rescued by pharmacological HDAC inhibition.

Keywords:  Autophagy; Brain development; Disease modeling; GABA; HDAC; In vitro; MEA; Migration; Oligodendrocytes; Personalized

DOI:  https://doi.org/10.1007/s00018-024-05406-w
Mol Neurodegener. 2024 Aug 26. 19(1): 62

Pathological characteristics of axons and alterations of proteomic and lipidomic profiles in midbrain dopaminergic neurodegeneration induced by WDR45-deficiency.

Panpan Wang, Yaping Shao, Murad Al-Nusaif, Jun Zhang, Huijia Yang, Yuting Yang, Kunhyok Kim, Song Li, Cong Liu, Huaibin Cai, Weidong Le.

   BACKGROUND: Although WD repeat domain 45 (WDR45) mutations have been linked to β -propeller protein-associated neurodegeneration (BPAN), the precise molecular and cellular mechanisms behind this disease remain elusive. This study aims to shed light on the impacts of WDR45-deficiency on neurodegeneration, specifically axonal degeneration, within the midbrain dopaminergic (DAergic) system. We hope to better understand the disease process by examining pathological and molecular alterations, especially within the DAergic system.
METHODS: To investigate the impacts of WDR45 dysfunction on mouse behaviors and DAergic neurons, we developed a mouse model in which WDR45 was conditionally knocked out in the midbrain DAergic neurons (WDR45cKO). Through a longitudinal study, we assessed alterations in the mouse behaviors using open field, rotarod, Y-maze, and 3-chamber social approach tests. We utilized a combination of immunofluorescence staining and transmission electron microscopy to examine the pathological changes in DAergic neuron soma and axons. Additionally, we performed proteomic and lipidomic analyses of the striatum from young and aged mice to identify the molecules and processes potentially involved in the striatal pathology during aging. Further more, primary midbrain neuronal culture was employed to explore the molecular mechanisms leading to axonal degeneration.
RESULTS: Our study of WDR45cKO mice revealed a range of deficits, including impaired motor function, emotional instability, and memory loss, coinciding with the profound reduction of midbrain DAergic neurons. The neuronal loss, we observed massive axonal enlargements in the dorsal and ventral striatum. These enlargements were characterized by the accumulation of extensively fragmented tubular endoplasmic reticulum (ER), a hallmark of axonal degeneration. Proteomic analysis of the striatum showed that the differentially expressed proteins were enriched in metabolic processes. The carbohydrate metabolic and protein catabolic processes appeared earlier, and amino acid, lipid, and tricarboxylic acid metabolisms were increased during aging. Of note, we observed a tremendous increase in the expression of lysophosphatidylcholine acyltransferase 1 (Lpcat1) that regulates phospholipid metabolism, specifically in the conversion of lysophosphatidylcholine (LPC) to phosphatidylcholine (PC) in the presence of acyl-CoA. The lipidomic results consistently suggested that differential lipids were concentrated on PC and LPC. Axonal degeneration was effectively ameliorated by interfering Lpcat1 expression in primary cultured WDR45-deficient DAergic neurons, proving that Lpcat1 and its regulated lipid metabolism, especially PC and LPC metabolism, participate in controlling the axonal degeneration induced by WDR45 deficits.
CONCLUSIONS: In this study, we uncovered the molecular mechanisms underlying the contribution of WDR45 deficiency to axonal degeneration, which involves complex relationships between phospholipid metabolism, autophagy, and tubular ER. These findings greatly advance our understanding of the fundamental molecular mechanisms driving axonal degeneration and may provide a foundation for developing novel mechanistically based therapeutic interventions for BPAN and other neurodegenerative diseases.

Keywords:  Autophagy; Axonal degeneration; Lpcat1; Phospholipid metabolism; Tubular ER; WDR45

DOI:  https://doi.org/10.1186/s13024-024-00746-4
Sci Rep. 2024 08 27. 14(1): 19796

Inflammatory signature in amyotrophic lateral sclerosis predicting disease progression.

Cinzia Femiano, Antonio Bruno, Luana Gilio, Fabio Buttari, Ettore Dolcetti, Giovanni Galifi, Federica Azzolini, Angela Borrelli, Roberto Furlan, Annamaria Finardi, Alessandra Musella, Georgia Mandolesi, Marianna Storto, Diego Centonze, Mario Stampanoni Bassi.

  Experimental studies identified a role of neuroinflammation in the pathogenesis of neurodegenerative diseases, including amyotrophic lateral sclerosis (ALS). However, the role of inflammatory molecules as diagnostic and prognostic biomarkers in patients with ALS is unclear. In this cross-sectional study, the cerebrospinal fluid (CSF) levels of a set of inflammatory cytokines and chemokines were analyzed in 56 newly diagnosed ALS patients and in 47 age- and sex-matched control patients without inflammatory or degenerative neurological disorders. The molecules analyzed included: interleukin (IL)-1β, IL-2, IL-4, IL-5, IL-6, IL-7, IL-8, IL-9, IL-10, IL-12, IL-13, IL-17, granulocyte colony stimulating factor (GCSF), macrophage inflammatory protein (MIP)-1a, MIP-1b, tumor necrosis factors (TNF), eotaxin. Principal component analysis (PCA) was used to explore possible associations between CSF molecules and ALS diagnosis. In addition, we analyzed the association between CSF cytokine profiles and clinical characteristics, including the disease progression rate score, and peripheral inflammation assessed using the Neutrophil-to-lymphocyte ratio (NLR). PCA identified six principal components (PCs) explaining 70.67% of the total variance in the CSF cytokine set. The principal component (PC1) explained 26.8% of variance and showed a positive load with CSF levels of IL-9, IL-4, GCSF, IL-7, IL-17, IL-13, IL-6, IL-1β, TNF, and IL-2. Logistic regression showed a significant association between PC1 and ALS diagnosis. In addition, in ALS patients, the same component was significantly associated with higher disease progression rate score and positively correlated with NLR. CSF inflammatory activation in present in ALS at the time of diagnosis and may characterize patients at higher risk for disease progression.

Keywords:  Amyotrophic lateral sclerosis (ALS); Cerebrospinal fluid (CSF); Cytokines; Disease progression; Neuroinflammation; Neutrophil-to-lymphocytes ratio (NLR)

DOI:  https://doi.org/10.1038/s41598-024-67165-9
Adv Clin Chem. 2024 ;pii: S0065-2423(24)00103-3. [Epub ahead of print]123 65-128

Neurofilaments in neurologic disease.

Christina Mousele, David Holden, Sharmilee Gnanapavan.

  Neurofilaments (NFs), major cytoskeletal constituents of neurons, have emerged as universal biomarkers of neuronal injury. Neuroaxonal damage underlies permanent disability in various neurological conditions. It is crucial to accurately quantify and longitudinally monitor this damage to evaluate disease progression, evaluate treatment effectiveness, contribute to novel treatment development, and offer prognostic insights. Neurofilaments show promise for this purpose, as their levels increase with neuroaxonal damage in both cerebrospinal fluid and blood, independent of specific causal pathways. New assays with high sensitivity allow reliable measurement of neurofilaments in body fluids and open avenues to investigate their role in neurological disorders. This book chapter will delve into the evolving landscape of neurofilaments, starting with their structure and cellular functions within neurons. It will then provide a comprehensive overview of their broad clinical value as biomarkers in diseases affecting the central or peripheral nervous system.

Keywords:  Alzheimer’s disease; Amyotrophic lateral sclerosis; Biomarkers; Charcot-Marie-Tooth; Cytoskeletal; Frontotemporal dementia; Huntington’s disease; Inflammatory polyneuropathies; Intermediate filaments; Multiple sclerosis; Neurofilaments; Neuronal injury; Parkinson’s disease

DOI:  https://doi.org/10.1016/bs.acc.2024.06.010
Int J Mol Sci. 2024 Aug 13. pii: 8795. [Epub ahead of print]25(16):

Regulation of TIR-1/SARM-1 by miR-71 Protects Dopaminergic Neurons in a C. elegans Model of LRRK2-Induced Parkinson's Disease.

Devin Naidoo, Alexandre de Lencastre.

  Parkinson's disease (PD) is a common neurodegenerative disorder characterized by symptoms such as bradykinesia, resting tremor, and rigidity, primarily driven by the degradation of dopaminergic (DA) neurons in the substantia nigra. A significant contributor to familial autosomal dominant PD cases is mutations in the LRRK2 gene, making it a primary therapeutic target. This study explores the role of microRNAs (miRNAs) in regulating the proteomic stress responses associated with neurodegeneration in PD using C. elegans models. Our focus is on miR-71, a miRNA known to affect stress resistance and act as a pro-longevity factor in C. elegans. We investigated miR-71's function in C. elegans models of PD, where mutant LRRK2 expression correlates with dopaminergic neuronal death. Our findings reveal that miR-71 overexpression rescues motility defects and slows dopaminergic neurodegeneration in these models, suggesting its critical role in mitigating the proteotoxic effects of mutant LRRK2. Conversely, miR-71 knockout exacerbates neuronal death caused by mutant LRRK2. Additionally, our data indicate that miR-71's neuroprotective effect involves downregulating the toll receptor domain protein tir-1, implicating miR-71 repression of tir-1 as vital in the response to LRRK2-induced proteotoxicity. These insights into miR-71's role in C. elegans models of PD not only enhance our understanding of molecular mechanisms in neurodegeneration but also pave the way for potential research into human neurodegenerative diseases, leveraging the conservation of miRNAs and their targets across species.

Keywords:  Parkinson; aging; elegans; miRNA; neurodegeneration

DOI:  https://doi.org/10.3390/ijms25168795
Cells. 2024 Aug 06. pii: 1313. [Epub ahead of print]13(16):

RILP Induces Cholesterol Accumulation in Lysosomes by Inhibiting Endoplasmic Reticulum-Endolysosome Interactions.

Yang Han, Xiaoqing Liu, Liju Xu, Ziheng Wei, Yueting Gu, Yandan Ren, Wenyi Hua, Yongtao Zhang, Xiaoxi Liu, Cong Jiang, Ruijuan Zhuang, Wanjin Hong, Tuanlao Wang.

  Endoplasmic reticulum (ER)-endolysosome interactions regulate cholesterol exchange between the ER and the endolysosome. ER-endolysosome membrane contact sites mediate the ER-endolysosome interaction. VAP-ORP1L (vesicle-associated membrane protein-associated protein- OSBP-related protein 1L) interaction forms the major contact site between the ER and the lysosome, which is regulated by Rab7. RILP (Rab7-interacting lysosomal protein) is the downstream effector of Rab7, but its role in the organelle interaction between the ER and the lysosome is not clear. In this study, we found RILP interacts with ORP1L to competitively inhibit the formation of the VAP-ORP1L contact site. Immunofluorescence microscopy revealed that RILP induces late endosome/lysosome clustering, which reduces the contact of endolysosomes with the ER, interfering with the ER-endolysosome interaction. Further examination demonstrated that over-expression of RILP results in the accumulation of cholesterol in the clustered endolysosomes, which triggers cellular autophagy depending on RILP. Our results suggest that RILP interferes with the ER-endolysosome interaction to inhibit cholesterol flow from the endolysosome to the ER, which feedbacks to trigger autophagy.

Keywords:  RILP; Rab7; autophagy; cholesterol transport; organelle interaction

DOI:  https://doi.org/10.3390/cells13161313
Elife. 2024 Aug 28. pii: RP92806. [Epub ahead of print]12

Botulinum toxin intoxication requires retrograde transport and membrane translocation at the ER in RenVM neurons.

Jeremy C Yeo, Felicia P Tay, Rebecca Bennion, Omar Loss, Jacquie Maignel, Laurent Pons, Keith Foster, Matthew Beard, Frederic Bard.

  Botulinum neurotoxin A (BoNT/A) is a highly potent proteolytic toxin specific for neurons with numerous clinical and cosmetic uses. After uptake at the synapse, the protein is proposed to translocate from synaptic vesicles to the cytosol through a self-formed channel. Surprisingly, we found that after intoxication proteolysis of a fluorescent reporter occurs in the neuron soma first and then centrifugally in neurites. To investigate the molecular mechanisms at play, we use a genome-wide siRNA screen in genetically engineered neurons and identify over three hundred genes. An organelle-specific split-mNG complementation indicates BoNT/A traffic from the synapse to the soma-localized Golgi in a retromer-dependent fashion. The toxin then moves to the ER and appears to require the Sec61 complex for retro-translocation to the cytosol. Our study identifies genes and trafficking processes hijacked by the toxin, revealing a new pathway mediating BoNT/A cellular toxicity.

Keywords:  botulinum toxin; cell biology; endoplasmic reticulum; human; intoxication; neuron; translocation

DOI:  https://doi.org/10.7554/eLife.92806
Proc Natl Acad Sci U S A. 2024 Sep 03. 121(36): e2322399121

A neural network for religious fundamentalism derived from patients with brain lesions.

Michael A Ferguson, Erik W Asp, Isaiah Kletenik, Daniel Tranel, Aaron D Boes, Jenae M Nelson, Frederic L W V J Schaper, Shan Siddiqi, Joseph I Turner, J Seth Anderson, Jared A Nielsen, James R Bateman, Jordan Grafman, Michael D Fox.

  Religious fundamentalism, characterized by rigid adherence to a set of beliefs putatively revealing inerrant truths, is ubiquitous across cultures and has a global impact on society. Understanding the psychological and neurobiological processes producing religious fundamentalism may inform a variety of scientific, sociological, and cultural questions. Research indicates that brain damage can alter religious fundamentalism. However, the precise brain regions involved with these changes remain unknown. Here, we analyzed brain lesions associated with varying levels of religious fundamentalism in two large datasets from independent laboratories. Lesions associated with greater fundamentalism were connected to a specific brain network with nodes in the right orbitofrontal, dorsolateral prefrontal, and inferior parietal lobe. This fundamentalism network was strongly right hemisphere lateralized and highly reproducible across the independent datasets (r = 0.82) with cross-validations between datasets. To explore the relationship of this network to lesions previously studied by our group, we tested for similarities to twenty-one lesion-associated conditions. Lesions associated with confabulation and criminal behavior showed a similar connectivity pattern as lesions associated with greater fundamentalism. Moreover, lesions associated with poststroke pain showed a similar connectivity pattern as lesions associated with lower fundamentalism. These findings are consistent with the current understanding of hemispheric specializations for reasoning and lend insight into previously observed epidemiological associations with fundamentalism, such as cognitive rigidity and outgroup hostility.

Keywords:  brain lesions; human behavior; neurology; neuroscience; religion

DOI:  https://doi.org/10.1073/pnas.2322399121
Zhejiang Da Xue Xue Bao Yi Xue Ban. 2024 Aug 25. 53(4): 498-508

Experimental study on small molecule combinations inducing reprogramming of rat fibroblasts into functional neurons.

Qunwei Gao, Zhenjia Dai, Xinkang Yang, Changqing Liu, Gaofeng Liu.

   OBJECTIVES: To establish a methodological system for reprogramming rat embryonic fibroblasts (REF) into chemically induced neurons (ciNCs) via small molecule compounds to provide safe and effective donor cells for treatment of neurodegenerative diseases.
METHODS: Based on the method established by PEI Gang's research group to directly reprogram human fibroblasts into neurons, the induction medium and maturation medium was optimized by replacing the coating solution, mitigating oxidative stress injury, adding neurogenic protective factors, adjusting the concentration of trichothecenes, performing small-molecule removal experiments, and carrying out immunofluorescence and Western blotting on cells at different stages of induction to validate the effect of induction.
RESULTS: When the original protocol was used for induction, the cell survival rate was (34.24±2.77)%. After replacing the coating solution gelatin with matrigel, the cell survival rate increased to (45.41±4.27)%; after adding melatonin, the cell survival rate increased to (67.95±5.61)% and (23.43±1.42)% were transformed into neural-like cells; after adding the small molecule P7C3-A20, the cell survival rate was further increased to (76.27±1.41)%, and (39.72±4.75)% of the cells were transformed into neural-like cells. When the concentration of trichothecene was increased to 30 μmol/L, the proportion of neural-like cells reached (55.79±1.90)%; after the removal of SP600125, (86.96±2.15)% of the cells survived, and the rate of neural-like cell production increased to (63.43±1.60)%. With the optimized protocol, REF could be successfully induced into ciNC through the neural precursor cell stage, in which the neural precursor cells were able to highly express the neural precursor cell markers SRY-related HMG-box gene 2 (Sox2) and paired box 6 (Pax6) as well as neuron-specific marker tubulin 1 (Tuj1), while the expression of fiber-associated protein vimentin was reduced. After two weeks of induction of neural precursor cells in a maturation medium, most cells displayed neuronal-like cell morphology. The induced ciNCs were able to highly express the mature neuronal surface markers Tuj1 and microtubule-associated protein 2 (MAP2), while the expression of vimentin was reduced.
CONCLUSIONS: The small molecule combinations optimized in this study can reprogram REF to ciNCs under normoxic conditions.

Keywords:  Chemically induced neurons; Embryo fibroblast; Rats; Small molecule compound; Somatic reprogramming

DOI:  https://doi.org/10.3724/zdxbyxb-2024-0007
Bioengineering (Basel). 2024 Jul 31. pii: 774. [Epub ahead of print]11(8):

Machine Learning-Enhanced Estimation of Cellular Protein Levels from Bright-Field Images.

Takeshi Tohgasaki, Arisa Touyama, Shohei Kousai, Kaita Imai.

  In this study, we aimed to develop a novel method for non-invasively determining intracellular protein levels, which is essential for understanding cellular phenomena. This understanding hinges on insights into gene expression, cell morphology, dynamics, and intercellular interactions. Traditional cell analysis techniques, such as immunostaining, live imaging, next-generation sequencing, and single-cell analysis, despite rapid advancements, face challenges in comprehensively integrating gene and protein expression data with spatiotemporal information. Leveraging advances in machine learning for image analysis, we designed a new model to estimate cellular biomarker protein levels using a blend of phase-contrast and fluorescent immunostaining images of epidermal keratinocytes. By iterating this process across various proteins, our model can estimate multiple protein levels from a single phase-contrast image. Additionally, we developed a system for analyzing multiple protein expression levels alongside spatiotemporal data through live imaging and phase-contrast methods. Our study offers valuable tools for cell-based research and presents a new avenue for addressing molecular biological challenges.

Keywords:  artificial intelligence; biomarker; keratinocyte; live cell imaging; machine learning

DOI:  https://doi.org/10.3390/bioengineering11080774
bioRxiv. 2024 Aug 13. pii: 2024.08.13.607780. [Epub ahead of print]

Axonal mitochondria regulate gentle touch response through control of axonal actin dynamics.

Sneha Hegde, Souvik Modi, Ennis W Deihl, Oliver Vinzenz Glomb, Shaul Yogev, Frederic J Hoerndli, Sandhya P Koushika.

Actin in neuronal processes is both stable and dynamic. The origin & functional roles of the different pools of actin is not well understood. We find that mutants that lack mitochondria, ric-7 and mtx-2; miro-1 , in neuronal processes also lack dynamic actin. Mitochondria can regulate actin dynamics upto a distance ∼80 μm along the neuronal process. Absence of axonal mitochondria and dynamic actin does not markedly alter the Spectrin Membrane Periodic Skeleton (MPS) in touch receptor neurons (TRNs). Restoring mitochondria inTRNs cell autonomously restores dynamic actin in a sod-2 dependent manner. We find that dynamic actin is necessary and sufficient for the localization of gap junction proteins in the TRNs and for the C. elegans gentle touch response. We identify an in vivo mechanism by which axonal mitochondria locally facilitate actin dynamics through reactive oxygen species that we show is necessary for electrical synapses & behaviour.

DOI: https://doi.org/10.1101/2024.08.13.607780
Smart Med. 2022 Dec;1(1): e20220007

Stem cells for organoids.

Shutong Qian, Jiayi Mao, Zhimo Liu, Binfan Zhao, Qiuyu Zhao, Bolun Lu, Liucheng Zhang, Xiyuan Mao, Liying Cheng, Wenguo Cui, Yuguang Zhang, Xiaoming Sun.

  Organoids are three-dimensional (3D) cell culture systems that simulate the structures and functions of organs, involving applications in disease modeling, drug screening, and cellular developmental biology. The material matrix in organoids can provide a 3D environment for stem cells to differentiate into different cell types and continuously self-renew, thereby realizing the in vitro culture of organs, which has received extensive attention in recent years. However, some challenges still exist in organoids, including low maturity, high heterogeneity, and lack of spatiotemporal regulation. Therefore, in this review, we summarized the culturing protocols and various applications of stem cell-derived organoids and proposed insightful thoughts for engineering stem cells into organoids in view of the current shortcomings, to achieve the further application and clinical translation of stem cells and engineered stem cells in organoid research.

Keywords:  cell surface engineering; gene editing; organoid; stem cell

DOI:  https://doi.org/10.1002/SMMD.20220007
J Neurosci. 2024 Aug 28. pii: e0129232024. [Epub ahead of print]

Neuritin controls axonal branching in serotonin neurons: A possible mediator involved in the regulation of depressive and anxiety behaviors via FGF signaling.

Tadayuki Shimada, Kuniko Kohyama, Tomoyuki Yoshida, Kanato Yamagata.

Abnormal neuronal morphological features, such as dendrite branching, axonal branching, and spine density, is thought to contribute to the symptoms of depression and anxiety. However, the role and molecular mechanisms of aberrant neuronal morphology in the regulation of mood disorders remain poorly characterized. Here, we show that Neuritin, an activity-dependent protein, regulates the axonal morphology of serotonin neurons. Male neuritin knockout mice harbored impaired axonal branches of serotonin neurons in the medial prefrontal cortex and basolateral region of the amygdala (BLA), and male neuritin knockout mice exhibited depressive and anxiety-like behaviors. We also observed that the expression of Neuritin was decreased by unpredictable chronic stress (UCS) in the male mouse brain and that decreased expression of neuritin was associated with reduced axonal branching of serotonin neurons in the brain and with depressive and anxiety behaviors in mice. Furthermore, the stress-mediated impairments in axonal branching and depressive behaviors were reversed by the overexpression of Neuritin in the BLA. The ability of neuritin to increase axonal branching in serotonin neurons involves FGF signaling, and neuritin contributes to FGF-2-mediated axonal branching regulation in vitro Finally, the oral administration of an FGF inhibitor reduced the axonal branching of serotonin neurons in the brain and caused depressive and anxiety behaviors in male mice. Our results support the involvement of neuritin in models of stress-induced depression and suggest that neuronal morphological plasticity may play a role in controlling animal behavior.Significance statement Axonal atrophy of serotonin neurons is one of the representative neuroanatomical features of depression. We found that the secreted/membrane-anchored neurotrophic factor Neuritin regulated axonal branch formation, which is involved in the development of depression and anxiety. In addition, Neuritin and the secreted signaling protein fibroblast growth factor 2 (FGF-2) cooperate to promote axonal branching in serotonin neurons. Furthermore, the inhibition of FGF signaling promoted axonal branching impairments and depressive behavior in mice. Taken together, these findings suggest that Neuritin regulates axonal branching in serotonin neurons and that the loss of neuritin is related to the development of depression. FGF signaling is involved in the neuritin-mediated axonal branching of serotonin neurons.

DOI: https://doi.org/10.1523/JNEUROSCI.0129-23.2024