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1.
Cytometry A ; 105(8): 607-620, 2024 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-38943226

RESUMO

Autofluorescence lifetime imaging microscopy (FLIM) is sensitive to metabolic changes in single cells based on changes in the protein-binding activities of the metabolic co-enzymes NAD(P)H. However, FLIM typically relies on time-correlated single-photon counting (TCSPC) detection electronics on laser-scanning microscopes, which are expensive, low-throughput, and require substantial post-processing time for cell segmentation and analysis. Here, we present a fluorescence lifetime-sensitive flow cytometer that offers the same TCSPC temporal resolution in a flow geometry, with low-cost single-photon excitation sources, a throughput of tens of cells per second, and real-time single-cell analysis. The system uses a 375 nm picosecond-pulsed diode laser operating at 50 MHz, alkali photomultiplier tubes, an FPGA-based time tagger, and can provide real-time phasor-based classification (i.e., gating) of flowing cells. A CMOS camera produces simultaneous brightfield images using far-red illumination. A second PMT provides two-color analysis. Cells are injected into the microfluidic channel using a syringe pump at 2-5 mm/s with nearly 5 ms integration time per cell, resulting in a light dose of 2.65 J/cm2 that is well below damage thresholds (25 J/cm2 at 375 nm). Our results show that cells remain viable after measurement, and the system is sensitive to autofluorescence lifetime changes in Jurkat T cells with metabolic perturbation (sodium cyanide), quiescent versus activated (CD3/CD28/CD2) primary human T cells, and quiescent versus activated primary adult mouse neural stem cells, consistent with prior studies using multiphoton FLIM. This TCSPC-based autofluorescence lifetime flow cytometer provides a valuable label-free method for real-time analysis of single-cell function and metabolism with higher throughput than laser-scanning microscopy systems.


Assuntos
Citometria de Fluxo , Fótons , Citometria de Fluxo/métodos , Humanos , Animais , Camundongos , Análise de Célula Única/métodos , Imagem Óptica/métodos , Células Jurkat , Fluorescência
2.
J Neurosci ; 37(40): 9632-9644, 2017 10 04.
Artigo em Inglês | MEDLINE | ID: mdl-28871032

RESUMO

Neurons in the adult mammalian CNS decrease in intrinsic axon growth capacity during development in concert with changes in Krüppel-like transcription factors (KLFs). KLFs regulate axon growth in CNS neurons including retinal ganglion cells (RGCs). Here, we found that knock-down of KLF9, an axon growth suppressor that is normally upregulated 250-fold in RGC development, promotes long-distance optic nerve regeneration in adult rats of both sexes. We identified a novel binding partner, MAPK10/JNK3 kinase, and found that JNK3 (c-Jun N-terminal kinase 3) is critical for KLF9's axon-growth-suppressive activity. Interfering with a JNK3-binding domain or mutating two newly discovered serine phosphorylation acceptor sites, Ser106 and Ser110, effectively abolished KLF9's neurite growth suppression in vitro and promoted axon regeneration in vivo These findings demonstrate a novel, physiologic role for the interaction of KLF9 and JNK3 in regenerative failure in the optic nerve and suggest new therapeutic strategies to promote axon regeneration in the adult CNS.SIGNIFICANCE STATEMENT Injured CNS nerves fail to regenerate spontaneously. Promoting intrinsic axon growth capacity has been a major challenge in the field. Here, we demonstrate that knocking down Krüppel-like transcription factor 9 (KLF9) via shRNA promotes long-distance axon regeneration after optic nerve injury and uncover a novel and important KLF9-JNK3 interaction that contributes to axon growth suppression in vitro and regenerative failure in vivo These studies suggest potential therapeutic approaches to promote axon regeneration in injury and other degenerative diseases in the adult CNS.


Assuntos
Axônios/fisiologia , Encéfalo/fisiologia , Fatores de Transcrição Kruppel-Like/metabolismo , Proteína Quinase 10 Ativada por Mitógeno/metabolismo , Regeneração Nervosa/fisiologia , Fatores Etários , Animais , Sequência de Bases , Células Cultivadas , Sistema Nervoso Central/fisiologia , Feminino , Fatores de Transcrição Kruppel-Like/deficiência , Fatores de Transcrição Kruppel-Like/genética , Masculino , Camundongos , Proteína Quinase 10 Ativada por Mitógeno/genética , Traumatismos do Nervo Óptico/genética , Traumatismos do Nervo Óptico/metabolismo , Técnicas de Cultura de Órgãos , Ligação Proteica/fisiologia , Ratos , Células Ganglionares da Retina/fisiologia
3.
J Vis Exp ; (206)2024 Apr 12.
Artigo em Inglês | MEDLINE | ID: mdl-38682901

RESUMO

Neural stem cells (NSCs) divide and produce newborn neurons in the adult brain through a process called adult neurogenesis. Adult NSCs are primarily quiescent, a reversible cell state where they have exited the cell cycle (G0) yet remain responsive to the environment. In the first step of adult neurogenesis, quiescent NSCs (qNSCs) receive a signal and activate, exiting quiescence and re-entering the cell cycle. Thus, understanding the regulators of NSC quiescence and quiescence exit is critical for future strategies targeting adult neurogenesis. However, our understanding of NSC quiescence is limited by technical constraints in identifying quiescent NSCs (qNSCs) and activated NSCs (aNSCs). This protocol describes a new approach to identify and enrich qNSCs and aNSCs generated in in vitro cultures by imaging NSC autofluorescence. First, this protocol describes how to use a confocal microscope to identify autofluorescent markers of qNSCs and aNSCs to classify NSC activation state using autofluorescence intensity. Second, this protocol describes how to use a fluorescent activated cell sorter (FACS) to classify NSC activation state and enrich samples for qNSCs or aNSCs using autofluorescence intensity. Third, this protocol describes how to use a multiphoton microscope to perform fluorescence lifetime imaging (FLIM) at single-cell resolution, classify NSC activation state, and track the dynamics of quiescent exit using both autofluorescence intensities and fluorescence lifetimes. Thus, this protocol provides a live-cell, label-free, single-cell resolution toolkit for studying NSC quiescence and quiescence exit.


Assuntos
Células-Tronco Neurais , Células-Tronco Neurais/citologia , Animais , Camundongos , Microscopia Confocal/métodos , Citometria de Fluxo/métodos , Imagem Óptica/métodos , Neurogênese/fisiologia
4.
bioRxiv ; 2024 May 18.
Artigo em Inglês | MEDLINE | ID: mdl-38798331

RESUMO

Autofluorescence lifetime imaging microscopy (FLIM) is sensitive to metabolic changes in single cells based on changes in the protein-binding activities of the metabolic co-enzymes NAD(P)H. However, FLIM typically relies on time-correlated single-photon counting (TCSPC) detection electronics on laser-scanning microscopes, which are expensive, low-throughput, and require substantial post-processing time for cell segmentation and analysis. Here, we present a fluorescence lifetime-sensitive flow cytometer that offers the same TCSPC temporal resolution in a flow geometry, with low-cost single-photon excitation sources, a throughput of tens of cells per second, and real-time single-cell analysis. The system uses a 375nm picosecond-pulsed diode laser operating at 50MHz, alkali photomultiplier tubes, an FPGA-based time tagger, and can provide real-time phasor-based classification ( i.e ., gating) of flowing cells. A CMOS camera produces simultaneous brightfield images using far-red illumination. A second PMT provides two-color analysis. Cells are injected into the microfluidic channel using a syringe pump at 2-5 mm/s with nearly 5ms integration time per cell, resulting in a light dose of 2.65 J/cm 2 that is well below damage thresholds (25 J/cm 2 at 375 nm). Our results show that cells remain viable after measurement, and the system is sensitive to autofluorescence lifetime changes in Jurkat T cells with metabolic perturbation (sodium cyanide), quiescent vs. activated (CD3/CD28/CD2) primary human T cells, and quiescent vs. activated primary adult mouse neural stem cells, consistent with prior studies using multiphoton FLIM. This TCSPC-based autofluorescence lifetime flow cytometer provides a valuable label-free method for real-time analysis of single-cell function and metabolism with higher throughput than laser-scanning microscopy systems.

5.
Cell Stem Cell ; 31(4): 570-581.e7, 2024 Apr 04.
Artigo em Inglês | MEDLINE | ID: mdl-38521057

RESUMO

Neural stem cells (NSCs) must exit quiescence to produce neurons; however, our understanding of this process remains constrained by the technical limitations of current technologies. Fluorescence lifetime imaging (FLIM) of autofluorescent metabolic cofactors has been used in other cell types to study shifts in cell states driven by metabolic remodeling that change the optical properties of these endogenous fluorophores. Using this non-destructive, live-cell, and label-free strategy, we found that quiescent NSCs (qNSCs) and activated NSCs (aNSCs) have unique autofluorescence profiles. Specifically, qNSCs display an enrichment of autofluorescence localizing to a subset of lysosomes, which can be used as a graded marker of NSC quiescence to predict cell behavior at single-cell resolution. Coupling autofluorescence imaging with single-cell RNA sequencing, we provide resources revealing transcriptional features linked to deep quiescence and rapid NSC activation. Together, we describe an approach for tracking mouse NSC activation state and expand our understanding of adult neurogenesis.


Assuntos
Células-Tronco Neurais , Camundongos , Animais , Células-Tronco Neurais/metabolismo , Neurogênese/fisiologia , Neurônios , Biomarcadores/metabolismo
6.
Nat Commun ; 15(1): 8676, 2024 Oct 07.
Artigo em Inglês | MEDLINE | ID: mdl-39375347

RESUMO

Aging is characterized by the accumulation of proteins that display amyloid-like behavior. However, the molecular mechanisms by which these proteins arise remain unclear. Here, we demonstrate that amyloid-like proteins are produced in a variety of human cell types, including stem cells, brain organoids and fully differentiated neurons by mistakes that occur in messenger RNA molecules. Some of these mistakes generate mutant proteins already known to cause disease, while others generate proteins that have not been observed before. Moreover, we show that these mistakes increase when cells are exposed to DNA damage, a major hallmark of human aging. When taken together, these experiments suggest a mechanistic link between the normal aging process and age-related diseases.


Assuntos
Dano ao DNA , Neurônios , RNA Mensageiro , Humanos , Neurônios/metabolismo , RNA Mensageiro/metabolismo , RNA Mensageiro/genética , Proteínas Amiloidogênicas/metabolismo , Proteínas Amiloidogênicas/genética , Envelhecimento/metabolismo , Envelhecimento/genética , Organoides/metabolismo , Encéfalo/metabolismo , Amiloide/metabolismo , Mutação
8.
Front Neurosci ; 17: 1198041, 2023.
Artigo em Inglês | MEDLINE | ID: mdl-37425013

RESUMO

Axon regeneration is limited in the adult mammalian central nervous system (CNS) due to both intrinsic and extrinsic factors. Rodent studies have shown that developmental age can drive differences in intrinsic axon growth ability, such that embryonic rodent CNS neurons extend long axons while postnatal and adult CNS neurons do not. In recent decades, scientists have identified several intrinsic developmental regulators in rodents that modulate growth. However, whether this developmentally programmed decline in CNS axon growth is conserved in humans is not yet known. Until recently, there have been limited human neuronal model systems, and even fewer age-specific human models. Human in vitro models range from pluripotent stem cell-derived neurons to directly reprogrammed (transdifferentiated) neurons derived from human somatic cells. In this review, we discuss the advantages and disadvantages of each system, and how studying axon growth in human neurons can provide species-specific knowledge in the field of CNS axon regeneration with the goal of bridging basic science studies to clinical trials. Additionally, with the increased availability and quality of 'omics datasets of human cortical tissue across development and lifespan, scientists can mine these datasets for developmentally regulated pathways and genes. As there has been little research performed in human neurons to study modulators of axon growth, here we provide a summary of approaches to begin to shift the field of CNS axon growth and regeneration into human model systems to uncover novel drivers of axon growth.

9.
Trends Endocrinol Metab ; 34(8): 446-461, 2023 08.
Artigo em Inglês | MEDLINE | ID: mdl-37380501

RESUMO

Metabolism has emerged as a key regulator of stem cell behavior. Mitochondria are crucial metabolic organelles that are important for differentiated cells, yet considered less so for stem cells. However, recent studies have shown that mitochondria influence stem cell maintenance and fate decisions, inviting a revised look at this topic. In this review, we cover the current literature addressing the role of mitochondrial metabolism in mouse and human neural stem cells (NSCs) in the embryonic and adult brain. We summarize how mitochondria are implicated in fate regulation and how substrate oxidation affects NSC quiescence. We further explore single-cell RNA sequencing (scRNA-seq) data for metabolic signatures of adult NSCs, highlight emerging technologies reporting on metabolic signatures, and discuss mitochondrial metabolism in other stem cells.


Assuntos
Células-Tronco Adultas , Células-Tronco Neurais , Humanos , Camundongos , Animais , Células-Tronco Neurais/metabolismo , Diferenciação Celular/fisiologia , Mitocôndrias/metabolismo , Células-Tronco Adultas/metabolismo , Oxirredução
10.
bioRxiv ; 2023 Sep 29.
Artigo em Inglês | MEDLINE | ID: mdl-37808866

RESUMO

The brain is a high energy tissue, and the cell types of which it is comprised are distinct in function and in metabolic requirements. The transcriptional co-activator PGC-1a is a master regulator of mitochondrial function and is highly expressed in the brain; however, its cell-type specific role in regulating metabolism has not been well established. Here, we show that PGC-1a is responsive to aging and that expression of the neuron specific PGC-1a isoform allows for specialization in metabolic adaptation. Transcriptional profiles of the cortex from male mice show an impact of age on immune, inflammatory, and neuronal functional pathways and a highly integrated metabolic response that is associated with decreased expression of PGC-1a. Proteomic analysis confirms age-related changes in metabolism and further shows changes in ribosomal and RNA splicing pathways. We show that neurons express a specialized PGC-1a isoform that becomes active during differentiation from stem cells and is further induced during the maturation of isolated neurons. Neuronal but not astrocyte PGC-1a responds robustly to inhibition of the growth sensitive kinase GSK3b, where the brain specific promoter driven dominant isoform is repressed. The GSK3b inhibitor lithium broadly reprograms metabolism and growth signaling, including significantly lower expression of mitochondrial and ribosomal pathway genes and suppression of growth signaling, which are linked to changes in mitochondrial function and neuronal outgrowth. In vivo, lithium treatment significantly changes the expression of genes involved in cortical growth, endocrine, and circadian pathways. These data place the GSK3b/PGC-1a axis centrally in a growth and metabolism network that is directly relevant to brain aging.

11.
bioRxiv ; 2023 May 24.
Artigo em Inglês | MEDLINE | ID: mdl-37292613

RESUMO

Injury to adult mammalian central nervous system (CNS) axons results in limited regeneration. Rodent studies have revealed a developmental switch in CNS axon regenerative ability, yet whether this is conserved in humans is unknown. Using human fibroblasts from 8 gestational-weeks to 72 years-old, we performed direct reprogramming to transdifferentiate fibroblasts into induced neurons (Fib-iNs), avoiding pluripotency which restores cells to an embryonic state. We found that early gestational Fib-iNs grew longer neurites than all other ages, mirroring the developmental switch in regenerative ability in rodents. RNA-sequencing and screening revealed ARID1A as a developmentally-regulated modifier of neurite growth in human neurons. These data suggest that age-specific epigenetic changes may drive the intrinsic loss of neurite growth ability in human CNS neurons during development. One-Sentence Summary: Directly-reprogrammed human neurons demonstrate a developmental decrease in neurite growth ability.

12.
Mol Cell Neurosci ; 47(4): 233-43, 2011 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-21635952

RESUMO

The Krüppel-like family of transcription factors (KLFs) have been widely studied in proliferating cells, though very little is known about their role in post-mitotic cells, such as neurons. We have recently found that the KLFs play a role in regulating intrinsic axon growth ability in retinal ganglion cells (RGCs), a type of central nervous system (CNS) neuron. Previous KLF studies in other cell types suggest that there may be cell-type specific KLF expression patterns, and that their relative expression allows them to compete for binding sites, or to act redundantly to compensate for another's function. With at least 15 of 17 KLF family members expressed in neurons, it will be important for us to determine how this complex family functions to regulate the intricate gene programs of axon growth and regeneration. By further characterizing the mechanisms of the KLF family in the nervous system, we may better understand how they regulate neurite growth and axon regeneration.


Assuntos
Axônios/fisiologia , Fatores de Transcrição Kruppel-Like/metabolismo , Regeneração Nervosa/fisiologia , Sistema Nervoso/metabolismo , Neuritos/fisiologia , Humanos , Fatores de Transcrição Kruppel-Like/classificação , Fatores de Transcrição Kruppel-Like/genética , Isoformas de Proteínas/classificação , Isoformas de Proteínas/genética , Isoformas de Proteínas/metabolismo , Processamento de Proteína Pós-Traducional
13.
Sci Rep ; 12(1): 15001, 2022 Sep 02.
Artigo em Inglês | MEDLINE | ID: mdl-36056070

RESUMO

The aggresome is a protein turnover system in which proteins are trafficked along microtubules to the centrosome for degradation. Despite extensive focus on aggresomes in immortalized cell lines, it remains unclear if the aggresome is conserved in all primary cells and all cell-states. Here we examined the aggresome in primary adult mouse dermal fibroblasts shifted into four distinct cell-states. We found that in response to proteasome inhibition, quiescent and immortalized fibroblasts formed aggresomes, whereas proliferating and senescent fibroblasts did not. Using this model, we generated a resource to provide a characterization of the proteostasis networks in which the aggresome is used and transcriptomic features associated with the presence or absence of aggresome formation. Using this resource, we validate a previously reported role for p38 MAPK signaling in aggresome formation and identify TAK1 as a novel driver of aggresome formation upstream of p38 MAPKs. Together, our data demonstrate that the aggresome is a non-universal protein degradation system which can be used cell-state specifically and provide a resource for studying aggresome formation and function.


Assuntos
Corpos de Inclusão , Microtúbulos , Animais , Centrossomo/metabolismo , Fibroblastos/metabolismo , Corpos de Inclusão/metabolismo , Camundongos , Microtúbulos/metabolismo , Complexo de Endopeptidases do Proteassoma/metabolismo , Proteínas/metabolismo
14.
Mol Cell Neurosci ; 44(1): 43-54, 2010 May.
Artigo em Inglês | MEDLINE | ID: mdl-20159039

RESUMO

Neurons in the central nervous system lose their intrinsic capacity for axon regeneration as they mature, and it is widely hypothesized that changes in gene expression are responsible. Testing this hypothesis and identifying the relevant genes has been challenging because hundreds to thousands of genes are developmentally regulated in CNS neurons, but only a small subset are likely relevant to axon growth. Here we used automated high content analysis (HCA) methods to functionally test 743 plasmids encoding developmentally regulated genes in neurite outgrowth assays using postnatal cortical neurons. We identified both growth inhibitors (Ephexin, Aldolase A, Solute Carrier 2A3, and Chimerin), and growth enhancers (Doublecortin, Doublecortin-like, Kruppel-like Factor 6, and CaM-Kinase II gamma), some of which regulate established growth mechanisms like microtubule dynamics and small GTPase signaling. Interestingly, with only one exception the growth-suppressing genes were developmentally upregulated, and the growth-enhancing genes downregulated. These data provide important support for the hypothesis that developmental changes in gene expression control neurite outgrowth, and identify potential new gene targets to promote neurite outgrowth.


Assuntos
Diferenciação Celular/fisiologia , Córtex Cerebral/crescimento & desenvolvimento , Córtex Cerebral/metabolismo , Cones de Crescimento/metabolismo , Fatores de Crescimento Neural/metabolismo , Regeneração Nervosa/fisiologia , Animais , Bioensaio , Células Cultivadas , Córtex Cerebral/citologia , Proteínas do Domínio Duplacortina , Proteína Duplacortina , Frutose-Bifosfato Aldolase/análise , Frutose-Bifosfato Aldolase/genética , Frutose-Bifosfato Aldolase/metabolismo , Regulação da Expressão Gênica no Desenvolvimento/genética , Cones de Crescimento/ultraestrutura , Inibidores do Crescimento/análise , Inibidores do Crescimento/genética , Inibidores do Crescimento/metabolismo , Fatores de Troca do Nucleotídeo Guanina/análise , Fatores de Troca do Nucleotídeo Guanina/genética , Fatores de Troca do Nucleotídeo Guanina/metabolismo , Fator 6 Semelhante a Kruppel , Fatores de Transcrição Kruppel-Like/análise , Fatores de Transcrição Kruppel-Like/genética , Fatores de Transcrição Kruppel-Like/metabolismo , Proteínas Associadas aos Microtúbulos/análise , Proteínas Associadas aos Microtúbulos/genética , Proteínas Associadas aos Microtúbulos/metabolismo , Fatores de Crescimento Neural/análise , Fatores de Crescimento Neural/genética , Neuritos/metabolismo , Neuritos/ultraestrutura , Neurogênese/fisiologia , Plasticidade Neuronal/fisiologia , Neuropeptídeos/análise , Neuropeptídeos/genética , Neuropeptídeos/metabolismo , Proteínas Proto-Oncogênicas/análise , Proteínas Proto-Oncogênicas/genética , Proteínas Proto-Oncogênicas/metabolismo , Tratos Piramidais/citologia , Tratos Piramidais/crescimento & desenvolvimento , Tratos Piramidais/metabolismo , Ratos
15.
STAR Protoc ; 2(3): 100744, 2021 09 17.
Artigo em Inglês | MEDLINE | ID: mdl-34430917

RESUMO

Although exogenous overexpression of a protein fused to a fluorescent tag can provide insight for the protein's function, it also can produce artifacts attributed to its upregulation and may not fully report the endogenous regulation of the protein of interest. To circumvent these issues, we adapted a protocol to label endogenous proteins with fluorescent tags in primary adult mouse neural stem cells in vitro. Here, we describe reagent construction, reagent delivery, and a screening strategy to isolate edited cells. For complete details on the use and execution of this protocol, please refer to Morrow et al. (2020).


Assuntos
Imunofluorescência/métodos , Células-Tronco Neurais/metabolismo , Engenharia de Proteínas/métodos , Animais , Sistemas CRISPR-Cas/genética , Linhagem Celular , Edição de Genes/métodos , Camundongos
16.
Cell Stem Cell ; 28(5): 967-977.e8, 2021 05 06.
Artigo em Inglês | MEDLINE | ID: mdl-33631115

RESUMO

Neural stem cells (NSCs) generate neurons throughout life in the hippocampal dentate gyrus. With advancing age, levels of neurogenesis sharply drop, which has been associated with a decline in hippocampal memory function. However, cell-intrinsic mechanisms mediating age-related changes in NSC activity remain largely unknown. Here, we show that the nuclear lamina protein lamin B1 (LB1) is downregulated with age in mouse hippocampal NSCs, whereas protein levels of SUN-domain containing protein 1 (SUN1), previously implicated in Hutchinson-Gilford progeria syndrome (HGPS), increase. Balancing the levels of LB1 and SUN1 in aged NSCs restores the strength of the endoplasmic reticulum diffusion barrier that is associated with segregation of aging factors in proliferating NSCs. Virus-based restoration of LB1 expression in aged NSCs enhances stem cell activity in vitro and increases progenitor cell proliferation and neurogenesis in vivo. Thus, we here identify a mechanism that mediates age-related decline of neurogenesis in the mammalian hippocampus.


Assuntos
Envelhecimento , Lamina Tipo B , Células-Tronco Neurais , Progéria , Animais , Hipocampo/citologia , Camundongos , Neurogênese
18.
J Neuroophthalmol ; 30(4): 347-60, 2010 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-21107123

RESUMO

The failure of the optic nerve to regenerate after injury or in neurodegenerative disease remains a major clinical and scientific problem. Retinal ganglion cell (RGC) axons course through the optic nerve and carry all the visual information to the brain, but after injury, they fail to regrow through the optic nerve and RGC cell bodies typically die, leading to permanent loss of vision. There are at least 4 hurdles to overcome in preserving RGCs and regenerating their axons: 1) increase RGC survival, 2) overcome the inhibitory environment of the optic nerve, 3) enhance RGC intrinsic axon growth potential, and 4) optimize the mapping of RGC connections back into their targets in the brain.


Assuntos
Sobrevivência Celular/fisiologia , Regeneração Nervosa/fisiologia , Doenças do Nervo Óptico/patologia , Doenças do Nervo Óptico/terapia , Nervo Óptico/fisiologia , Células Ganglionares da Retina/fisiologia , Animais , Humanos , Nervo Óptico/patologia , Doenças do Nervo Óptico/fisiopatologia , Células Ganglionares da Retina/patologia
19.
Cytoskeleton (Hoboken) ; 77(11): 515-523, 2020 11.
Artigo em Inglês | MEDLINE | ID: mdl-33190414

RESUMO

Intermediate filaments (IFs) perform a diverse set of well-known functions including providing structural support for the cell and resistance to mechanical stress, yet recent evidence has revealed unexpected roles for IFs as stress response proteins. Previously, it was shown that the type III IF protein vimentin forms cage-like structures around centrosome-associated proteins destined for degradation, structures referred to as aggresomes, suggesting a role for vimentin in protein turnover. However, vimentin's function at the aggresome has remained largely understudied. In a recent report, vimentin was shown to be dispensable for aggresome formation, but played a critical role in protein turnover at the aggresome through localizing proteostasis-related machineries, such as proteasomes, to the aggresome. Here, we review evidence for vimentin's function in proteostasis and highlight the organismal implications of these findings.


Assuntos
Filamentos Intermediários/metabolismo , Proteostase/fisiologia , Vimentina/metabolismo , Animais , Humanos , Mamíferos
20.
Cell Stem Cell ; 26(4): 558-568.e9, 2020 04 02.
Artigo em Inglês | MEDLINE | ID: mdl-32109376

RESUMO

Maintaining a healthy proteome throughout life is critical for proper somatic stem cell function, but the complexities of the stem cell response to increases in damaged or aggregated proteins remain unclear. Here we demonstrate that adult neural stem cells (NSCs) utilize aggresomes to recover from disrupted proteostasis and describe a novel function for the intermediate filament vimentin in proteostasis as a spatial coordinator of proteasomes to the aggresome. In the absence of vimentin, NSCs have a reduced capacity to exit quiescence, a time when NSCs are required to clear a wave of aggregated proteins, and demonstrate an early age-dependent decline in proliferation and neurogenesis. Taken together, these data reveal a significant role of vimentin and aggresomes in the regulation of proteostasis during quiescent NSC activation.


Assuntos
Células-Tronco Adultas , Células-Tronco Neurais , Vimentina , Humanos , Filamentos Intermediários , Neurogênese
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