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17 pages, 4502 KiB

Open AccessArticle

Molecular Profiling of Mouse Models of Loss or Gain of Function of the KCNT1 (Slack) Potassium Channel and Antisense Oligonucleotide Treatment

by Fangxu Sun, Huafeng Wang, Jing Wu, Imran H. Quraishi, Yalan Zhang, Maysam Pedram, Benbo Gao, Elizabeth A. Jonas, Viet Nguyen, Sijia Wu, Omar S. Mabrouk, Paymaan Jafar-nejad and Leonard K. Kaczmarek

Biomolecules 2024, 14(11), 1397; https://doi.org/10.3390/biom14111397 - 2 Nov 2024

Viewed by 1149

Abstract

The potassium sodium-activated channel subtype T member 1 (KCNT1) gene encodes the Slack channel K_Na1.1, which is expressed in neurons throughout the brain. Gain-of-function variants in KCNT1 are associated with a spectrum of epilepsy syndromes, and mice carrying those [...] Read more.

The potassium sodium-activated channel subtype T member 1 (KCNT1) gene encodes the Slack channel K_Na1.1, which is expressed in neurons throughout the brain. Gain-of-function variants in KCNT1 are associated with a spectrum of epilepsy syndromes, and mice carrying those variants exhibit a robust phenotype similar to that observed in patients. Kcnt1 knockout (KO) mice, however, have a normal lifespan without any epileptic phenotype. To understand the molecular differences between these two models, we conducted a comprehensive proteomic analysis of the cerebral cortices of Kcnt1 KO and Kcnt1^R455H/+ mice, an animal model bearing a cytoplasmic C-terminal mutation homologous to a human R474H variant that results in EIMFS. The greatest change observed in Kcnt1 KO mice compared to the wild-type mice was the increased expression of multiple proteins of the inner mitochondrial membrane. Electron microscopy studies of cortical mitochondria from Kcnt1 KO mice further confirmed a significant increase in the density of mitochondrial cristae compared to that in wild-type mice. Kcnt1 reduction by a murine-specific Kcnt1 antisense oligonucleotide (ASO) in Kcnt1^R455H/+ mice partially corrected the proteomic dysregulations in the disease model. The results support the hypothesis that ASO-mediated KCNT1 reduction could be therapeutically useful in the treatment of KCNT1 epilepsies. Full article

(This article belongs to the Special Issue Advances in Neuroproteomics)

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17 pages, 2828 KiB

Open AccessArticle

Proteomic Profile of Circulating Extracellular Vesicles in the Brain after Δ9-Tetrahydrocannabinol Inhalation

by Valeria Lallai, TuKiet T. Lam, Rolando Garcia-Milian, Yen-Chu Chen, James P. Fowler, Letizia Manca, Daniele Piomelli, Kenneth Williams, Angus C. Nairn and Christie D. Fowler

Biomolecules 2024, 14(9), 1143; https://doi.org/10.3390/biom14091143 - 10 Sep 2024

Viewed by 1116

Abstract

Given the increasing use of cannabis in the US, there is an urgent need to better understand the drug’s effects on central signaling mechanisms. Extracellular vesicles (EVs) have been identified as intercellular signaling mediators that contain a variety of cargo, including proteins. Here, [...] Read more.

Given the increasing use of cannabis in the US, there is an urgent need to better understand the drug’s effects on central signaling mechanisms. Extracellular vesicles (EVs) have been identified as intercellular signaling mediators that contain a variety of cargo, including proteins. Here, we examined whether the main psychoactive component in cannabis, Δ9-tetrahydrocannabinol (THC), alters EV protein signaling dynamics in the brain. We first conducted in vitro studies, which found that THC activates signaling in choroid plexus epithelial cells, resulting in transcriptional upregulation of the cannabinoid 1 receptor and immediate early gene c-fos, in addition to the release of EVs containing RNA cargo. Next, male and female rats were examined for the effects of either acute or chronic exposure to aerosolized (‘vaped’) THC on circulating brain EVs. Cerebrospinal fluid was extracted from the brain, and EVs were isolated and processed with label-free quantitative proteomic analyses via high-resolution tandem mass spectrometry. Interestingly, circulating EV-localized proteins were differentially expressed based on acute or chronic THC exposure in a sex-specific manner. Taken together, these findings reveal that THC acts in the brain to modulate circulating EV signaling, thereby providing a novel understanding of how exogenous factors can regulate intercellular communication in the brain. Full article

(This article belongs to the Special Issue Advances in Neuroproteomics)

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17 pages, 2727 KiB

Open AccessEditor’s ChoiceArticle

Concept of Normativity in Multi-Omics Analysis of Axon Regeneration

by Isabella Moceri, Sean Meehan, Emily Gonzalez, Kevin K. Park, Abigail Hackam, Richard K. Lee and Sanjoy Bhattacharya

Biomolecules 2024, 14(7), 735; https://doi.org/10.3390/biom14070735 - 21 Jun 2024

Viewed by 1225

Abstract

Transcriptomes and proteomes can be normalized with a handful of RNAs or proteins (or their peptides), such as GAPDH, β-actin, RPBMS, and/or GAP43. Even with hundreds of standards, normalization cannot be achieved across different molecular mass ranges for small molecules, such as lipids [...] Read more.

Transcriptomes and proteomes can be normalized with a handful of RNAs or proteins (or their peptides), such as GAPDH, β-actin, RPBMS, and/or GAP43. Even with hundreds of standards, normalization cannot be achieved across different molecular mass ranges for small molecules, such as lipids and metabolites, due to the non-linearity of mass by charge ratio for even the smallest part of the spectrum. We define the amount (or range of amounts) of metabolites and/or lipids per a defined amount of a protein, consistently identified in all samples of a multiple-model organism comparison, as the normative level of that metabolite or lipid. The defined protein amount (or range) is a normalized value for one cohort of complete samples for which intrasample relative protein quantification is available. For example, the amount of citrate (a metabolite) per µg of aconitate hydratase (normalized protein amount) identified in the proteome is the normative level of citrate with aconitase. We define normativity as the amount of metabolites (or amount range) detected when compared to normalized protein levels. We use axon regeneration as an example to illustrate the need for advanced approaches to the normalization of proteins. Comparison across different pharmacologically induced axon regeneration mouse models entails the comparison of axon regeneration, studied at different time points in several models designed using different agents. For the normalization of the proteins across different pharmacologically induced models, we perform peptide doping (fixed amounts of known peptides) in each sample to normalize the proteome across the samples. We develop Regen V peptides, divided into Regen III (SEB, LLO, CFP) and II (HH4B, A1315), for pre- and post-extraction comparisons, performed with the addition of defined, digested peptides (bovine serum albumin tryptic digest) for protein abundance normalization beyond commercial labeled relative quantification (for example, 18-plex tandem mass tags). We also illustrate the concept of normativity by using this normalization technique on regenerative metabolome/lipidome profiles. As normalized protein amounts are different in different biological states (control versus axon regeneration), normative metabolite or lipid amounts are expected to be different for specific biological states. These concepts and standardization approaches are important for the integration of different datasets across different models of axon regeneration. Full article

(This article belongs to the Special Issue Advances in Neuroproteomics)

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22 pages, 11609 KiB

Open AccessArticle

Involvement of Glucosamine 6 Phosphate Isomerase 2 (GNPDA2) Overproduction in β-Amyloid- and Tau P301L-Driven Pathomechanisms

by Mercedes Lachén-Montes, Paz Cartas-Cejudo, Adriana Cortés, Elena Anaya-Cubero, Erika Peral, Karina Ausín, Ramón Díaz-Peña, Joaquín Fernández-Irigoyen and Enrique Santamaría

Biomolecules 2024, 14(4), 394; https://doi.org/10.3390/biom14040394 - 25 Mar 2024

Viewed by 1896

Abstract

Alzheimer’s disease (AD) is a neurodegenerative olfactory disorder affecting millions of people worldwide. Alterations in the hexosamine- or glucose-related pathways have been described through AD progression. Specifically, an alteration in glucosamine 6 phosphate isomerase 2 (GNPDA2) protein levels has been observed in olfactory [...] Read more.

Alzheimer’s disease (AD) is a neurodegenerative olfactory disorder affecting millions of people worldwide. Alterations in the hexosamine- or glucose-related pathways have been described through AD progression. Specifically, an alteration in glucosamine 6 phosphate isomerase 2 (GNPDA2) protein levels has been observed in olfactory areas of AD subjects. However, the biological role of GNPDA2 in neurodegeneration remains unknown. Using mass spectrometry, multiple GNPDA2 interactors were identified in human nasal epithelial cells (NECs) mainly involved in intraciliary transport. Moreover, GNPDA2 overexpression induced an increment in NEC proliferation rates, accompanied by transcriptomic alterations in Type II interferon signaling or cellular stress responses. In contrast, the presence of beta-amyloid or mutated Tau-P301L in GNPDA2-overexpressing NECs induced a slowdown in the proliferative capacity in parallel with a disruption in protein processing. The proteomic characterization of Tau-P301L transgenic zebrafish embryos demonstrated that GNPDA2 overexpression interfered with collagen biosynthesis and RNA/protein processing, without inducing additional changes in axonal outgrowth defects or neuronal cell death. In humans, a significant increase in serum GNPDA2 levels was observed across multiple neurological proteinopathies (AD, Lewy body dementia, progressive supranuclear palsy, mixed dementia and amyotrophic lateral sclerosis) (n = 215). These data shed new light on GNPDA2-dependent mechanisms associated with the neurodegenerative process beyond the hexosamine route. Full article

(This article belongs to the Special Issue Advances in Neuroproteomics)

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17 pages, 3368 KiB

Open AccessArticle

Identification of Central Nervous System Oncologic Disease Biomarkers in EVs from Cerebrospinal Fluid (CSF) of Pediatric Patients: A Pilot Neuro-Proteomic Study

by Xhuliana Kajana, Sonia Spinelli, Andrea Garbarino, Ganna Balagura, Martina Bartolucci, Andrea Petretto, Marco Pavanello, Giovanni Candiano, Isabella Panfoli and Maurizio Bruschi

Biomolecules 2023, 13(12), 1730; https://doi.org/10.3390/biom13121730 - 30 Nov 2023

Cited by 1 | Viewed by 1788

Abstract

Cerebrospinal fluid (CSF) is a biochemical–clinical window into the brain. Unfortunately, its wide dynamic range, low protein concentration, and small sample quantity significantly limit the possibility of using it routinely. Extraventricular drainage (EVD) of CSF allows us to solve quantitative problems and to [...] Read more.

Cerebrospinal fluid (CSF) is a biochemical–clinical window into the brain. Unfortunately, its wide dynamic range, low protein concentration, and small sample quantity significantly limit the possibility of using it routinely. Extraventricular drainage (EVD) of CSF allows us to solve quantitative problems and to study the biological role of extracellular vesicles (EVs). In this study, we implemented bioinformatic analysis of our previous data of EVD of CSF and its EVs obtained from congenital hydrocephalus with the aim of identifying a comprehensive list of potential tumor and non-tumor biomarkers of central nervous system diseases. Among all proteins identified, those enriched in EVs are associated with synapses, synaptosomes, and nervous system diseases including gliomas, embryonal tumors, and epilepsy. Among these EV-enriched proteins, given the broad consensus present in the recent scientific literature, we validated syntaxin-binding protein 1 (STXBP1) as a marker of malignancy in EVD of CSF and its EVs from patients with pilocytic astrocytoma and medulloblastoma. Our results show that STXBP1 is negatively enriched in EVs compared to non-tumor diseases and its downregulation correlates with adverse outcomes. Further experiments are needed to validate this and other EV markers in the blood of pediatric patients for translational medicine applications. Full article

(This article belongs to the Special Issue Advances in Neuroproteomics)

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19 pages, 3324 KiB

Open AccessArticle

Neuroproteomic Analysis after SARS-CoV-2 Infection Reveals Overrepresented Neurodegeneration Pathways and Disrupted Metabolic Pathways

by Indranil Basak, Rhodri Harfoot, Jennifer E. Palmer, Abhishek Kumar, Miguel E. Quiñones-Mateu, Lucia Schweitzer and Stephanie M. Hughes

Biomolecules 2023, 13(11), 1597; https://doi.org/10.3390/biom13111597 - 30 Oct 2023

Cited by 1 | Viewed by 2819

Abstract

Besides respiratory illness, SARS-CoV-2, the causative agent of COVID-19, leads to neurological symptoms. The molecular mechanisms leading to neuropathology after SARS-CoV-2 infection are sparsely explored. SARS-CoV-2 enters human cells via different receptors, including ACE-2, TMPRSS2, and TMEM106B. In this study, we used a [...] Read more.

Besides respiratory illness, SARS-CoV-2, the causative agent of COVID-19, leads to neurological symptoms. The molecular mechanisms leading to neuropathology after SARS-CoV-2 infection are sparsely explored. SARS-CoV-2 enters human cells via different receptors, including ACE-2, TMPRSS2, and TMEM106B. In this study, we used a human-induced pluripotent stem cell-derived neuronal model, which expresses ACE-2, TMPRSS2, TMEM106B, and other possible SARS-CoV-2 receptors, to evaluate its susceptibility to SARS-CoV-2 infection. The neurons were exposed to SARS-CoV-2, followed by RT-qPCR, immunocytochemistry, and proteomic analyses of the infected neurons. Our findings showed that SARS-CoV-2 infects neurons at a lower rate than other human cells; however, the virus could not replicate or produce infectious virions in this neuronal model. Despite the aborted SARS-CoV-2 replication, the infected neuronal nuclei showed irregular morphology compared to other human cells. Since cytokine storm is a significant effect of SARS-CoV-2 infection in COVID-19 patients, in addition to the direct neuronal infection, the neurons were treated with pre-conditioned media from SARS-CoV-2-infected lung cells, and the neuroproteomic changes were investigated. The limited SARS-CoV-2 infection in the neurons and the neurons treated with the pre-conditioned media showed changes in the neuroproteomic profile, particularly affecting mitochondrial proteins and apoptotic and metabolic pathways, which may lead to the development of neurological complications. The findings from our study uncover a possible mechanism behind SARS-CoV-2-mediated neuropathology that might contribute to the lingering effects of the virus on the human brain. Full article

(This article belongs to the Special Issue Advances in Neuroproteomics)

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Review

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21 pages, 2613 KiB

Open AccessReview

Mitochondrial Proteomes in Neural Cells: A Systematic Review

by Aya Nusir, Patricia Sinclair and Nadine Kabbani

Biomolecules 2023, 13(11), 1638; https://doi.org/10.3390/biom13111638 - 11 Nov 2023

Cited by 2 | Viewed by 2574

Abstract

Mitochondria are ancient endosymbiotic double membrane organelles that support a wide range of eukaryotic cell functions through energy, metabolism, and cellular control. There are over 1000 known proteins that either reside within the mitochondria or are transiently associated with it. These mitochondrial proteins [...] Read more.

Mitochondria are ancient endosymbiotic double membrane organelles that support a wide range of eukaryotic cell functions through energy, metabolism, and cellular control. There are over 1000 known proteins that either reside within the mitochondria or are transiently associated with it. These mitochondrial proteins represent a functional subcellular protein network (mtProteome) that is encoded by mitochondrial and nuclear genomes and significantly varies between cell types and conditions. In neurons, the high metabolic demand and differential energy requirements at the synapses are met by specific modifications to the mtProteome, resulting in alterations in the expression and functional properties of the proteins involved in energy production and quality control, including fission and fusion. The composition of mtProteomes also impacts the localization of mitochondria in axons and dendrites with a growing number of neurodegenerative diseases associated with changes in mitochondrial proteins. This review summarizes the findings on the composition and properties of mtProteomes important for mitochondrial energy production, calcium and lipid signaling, and quality control in neural cells. We highlight strategies in mass spectrometry (MS) proteomic analysis of mtProteomes from cultured cells and tissue. The research into mtProteome composition and function provides opportunities in biomarker discovery and drug development for the treatment of metabolic and neurodegenerative disease. Full article

(This article belongs to the Special Issue Advances in Neuroproteomics)

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23 pages, 2364 KiB

Open AccessReview

Cell-Type-Specific Neuroproteomics of Synapses

by Yun Young Yim and Eric J. Nestler

Biomolecules 2023, 13(6), 998; https://doi.org/10.3390/biom13060998 - 16 Jun 2023

Cited by 1 | Viewed by 3632

Abstract

In the last two decades, our knowledge of synaptic proteomes and their relationship to normal brain function and neuropsychiatric disorders has been expanding rapidly through the use of more powerful neuroproteomic approaches. However, mass spectrometry (MS)-based neuroproteomic studies of synapses still require cell-type, [...] Read more.

In the last two decades, our knowledge of synaptic proteomes and their relationship to normal brain function and neuropsychiatric disorders has been expanding rapidly through the use of more powerful neuroproteomic approaches. However, mass spectrometry (MS)-based neuroproteomic studies of synapses still require cell-type, spatial, and temporal proteome information. With the advancement of sample preparation and MS techniques, we have just begun to identify and understand proteomes within a given cell type, subcellular compartment, and cell-type-specific synapse. Here, we review the progress and limitations of MS-based neuroproteomics of synapses in the mammalian CNS and highlight the recent applications of these approaches in studying neuropsychiatric disorders such as major depressive disorder and substance use disorders. Combining neuroproteomic findings with other omics studies can generate an in-depth, comprehensive map of synaptic proteomes and possibly identify new therapeutic targets and biomarkers for several central nervous system disorders. Full article

(This article belongs to the Special Issue Advances in Neuroproteomics)

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