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1.
Photochem Photobiol ; 2024 Jun 23.
Artigo em Inglês | MEDLINE | ID: mdl-38922889

RESUMO

Glioblastoma stem cells (GSCs) are potent tumor initiators resistant to radiochemotherapy, and this subpopulation is hypothesized to re-populate the tumor milieu due to selection following conventional therapies. Here, we show that 5-aminolevulinic acid (ALA) treatment-a pro-fluorophore used for fluorescence-guided cancer surgery-leads to elevated levels of fluorophore conversion in patient-derived GSC cultures, and subsequent red light-activation induces apoptosis in both intrinsically temozolomide chemotherapy-sensitive and -resistant GSC phenotypes. Red light irradiation of ALA-treated cultures also exhibits the ability to target mesenchymal GSCs (Mes-GSCs) with induced temozolomide resistance. Furthermore, sub-lethal light doses restore Mes-GSC sensitivity to temozolomide, abrogating GSC-acquired chemoresistance. These results suggest that ALA is not only useful for fluorescence-guided glioblastoma tumor resection, but that it also facilitates a GSC drug-resistance agnostic, red light-activated modality to mop up the surgical margins and prime subsequent chemotherapy.

2.
Sci Rep ; 13(1): 19740, 2023 11 13.
Artigo em Inglês | MEDLINE | ID: mdl-37957190

RESUMO

Yes-associated protein (YAP) is a transcriptional coactivator that is essential for the malignancy of various cancers. We have previously shown that YAP activity is positively regulated by phosphatidylserine (PS) in recycling endosomes (REs). However, the mechanism by which YAP is activated by PS in REs remains unknown. In the present study, we examined a group of protein phosphatases (11 phosphatases) that we had identified previously as PS-proximity protein candidates. Knockdown experiments of these phosphatases suggested that PPP1R12A, a regulatory subunit of the myosin phosphatase complex, was essential for YAP-dependent proliferation of triple-negative breast cancer MDA-MB-231 cells. Knockdown of PPP1R12A increased the level of phosphorylated YAP, reduced that of YAP in the nucleus, and suppressed the transcription of CTGF (a YAP-regulated gene), reinforcing the role of PPP1R12A in YAP activation. ATP8A1 is a PS-flippase that concentrates PS in the cytosolic leaflet of the RE membrane and positively regulates YAP signalling. In subcellular fractionation experiments using cell lysates, PPP1R12A in control cells was recovered exclusively in the microsomal fraction. In contrast, a fraction of PPP1R12A in ATP8A1-depleted cells was recovered in the cytosolic fraction. Cohort data available from the Cancer Genome Atlas showed that high expression of PPP1R12A, PP1B encoding the catalytic subunit of the myosin phosphatase complex, or ATP8A1 correlated with poor prognosis in breast cancer patients. These results suggest that the "ATP8A1-PS-YAP phosphatase" axis in REs facilitates YAP activation and thus cell proliferation.


Assuntos
Monoéster Fosfórico Hidrolases , Transdução de Sinais , Humanos , Monoéster Fosfórico Hidrolases/metabolismo , Fosfatase de Miosina-de-Cadeia-Leve/genética , Fosfatase de Miosina-de-Cadeia-Leve/metabolismo , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo , Endossomos/metabolismo , Proliferação de Células , Adenosina Trifosfatases/metabolismo , Proteínas de Transferência de Fosfolipídeos/metabolismo
3.
Commun Biol ; 6(1): 665, 2023 06 23.
Artigo em Inglês | MEDLINE | ID: mdl-37353538

RESUMO

Cellular senescence, a state of irreversible cell-cycle arrest caused by a variety of cellular stresses, is critically involved in age-related tissue dysfunction in various organs. However, the features of cells in the central nervous system that undergo senescence and their role in neural impairment are not well understood as yet. Here, through comprehensive investigations utilising single-cell transcriptome analysis and various mouse models, we show that microglia, particularly in the white matter, undergo cellular senescence in the brain and spinal cord during ageing and in disease models involving demyelination. Microglial senescence is predominantly detected in disease-associated microglia, which appear in ageing and neurodegenerative diseases. We also find that commensal bacteria promote the accumulation of senescent microglia and disease-associated microglia during ageing. Furthermore, knockout of p16INK4a, a key senescence inducer, ameliorates the neuroinflammatory phenotype in damaged spinal cords in mice. These results advance our understanding of the role of cellular senescence in the central nervous system and open up possibilities for the treatment of age-related neural disorders.


Assuntos
Microglia , Substância Branca , Camundongos , Animais , Envelhecimento/fisiologia , Senescência Celular/fisiologia , Fenótipo
4.
Nat Cell Biol ; 25(6): 865-876, 2023 06.
Artigo em Inglês | MEDLINE | ID: mdl-37169880

RESUMO

The elucidation of the mechanisms of ageing and the identification of methods to control it have long been anticipated. Recently, two factors associated with ageing-the accumulation of senescent cells and the change in the composition of gut microbiota-have been shown to play key roles in ageing. However, little is known about how these phenomena occur and are related during ageing. Here we show that the persistent presence of commensal bacteria gradually induces cellular senescence in gut germinal centre B cells. Importantly, this reduces both the production and diversity of immunoglobulin A (IgA) antibodies that target gut bacteria, thereby changing the composition of gut microbiota in aged mice. These results have revealed the existence of IgA-mediated crosstalk between the gut microbiota and cellular senescence and thus extend our understanding of the mechanism of gut microbiota changes with age, opening up possibilities for their control.


Assuntos
Microbioma Gastrointestinal , Animais , Camundongos , Bactérias , Imunoglobulina A , Senescência Celular , Linfócitos B
5.
Immunol Lett ; 245: 51-60, 2022 05.
Artigo em Inglês | MEDLINE | ID: mdl-35413354

RESUMO

Recent advances in genetic analysis and intensive research have revealed that microglial cells respond to external stimuli in various ways and adopt diverse states. Herein, we summarize the current knowledge on the possible microglial states during development, homeostasis, and perturbation. These states are characterized either by irreversible cellular changes, such as cell death and cellular senescence, or reversible alterations, triggered by temporary changes in the microenvironment. Subsequently, we discuss the modalities of the transcriptional and functional recovery of microglia from a molecular perspective. An understanding of (ir)reversible changes in their cellular fates or transcriptional states could provide novel options for the treatment of age-related and neurodegenerative disorders.


Assuntos
Senescência Celular , Microglia , Morte Celular , Senescência Celular/fisiologia , Homeostase , Microglia/metabolismo
6.
Nat Aging ; 2(2): 115-124, 2022 02.
Artigo em Inglês | MEDLINE | ID: mdl-37117754

RESUMO

Reports of post-acute COVID-19 syndrome, in which the inflammatory response persists even after SARS-CoV-2 has disappeared, are increasing1, but the underlying mechanisms of post-acute COVID-19 syndrome remain unknown. Here, we show that SARS-CoV-2-infected cells trigger senescence-like cell-cycle arrest2,3 in neighboring uninfected cells in a paracrine manner via virus-induced cytokine production. In cultured human cells or bronchial organoids, these SASR-CoV-2 infection-induced senescent cells express high levels of a series of inflammatory factors known as senescence-associated secretory phenotypes (SASPs)4 in a sustained manner, even after SARS-CoV-2 is no longer detectable. We also show that the expression of the senescence marker CDKN2A (refs. 5,6) and various SASP factor4 genes is increased in the pulmonary cells of patients with severe post-acute COVID-19 syndrome. Furthermore, we find that mice exposed to a mouse-adapted strain of SARS-CoV-2 exhibit prolonged signs of cellular senescence and SASP in the lung at 14 days after infection when the virus was undetectable, which could be substantially reduced by the administration of senolytic drugs7. The sustained infection-induced paracrine senescence described here may be involved in the long-term inflammation caused by SARS-CoV-2 infection.


Assuntos
COVID-19 , Humanos , Camundongos , Animais , SARS-CoV-2 , Senescência Celular/genética , Pulmão , Inflamação
7.
Nat Commun ; 12(1): 5674, 2021 09 28.
Artigo em Inglês | MEDLINE | ID: mdl-34584098

RESUMO

Emerging evidence is revealing that alterations in gut microbiota are associated with colorectal cancer (CRC). However, very little is currently known about whether and how gut microbiota alterations are causally associated with CRC development. Here we show that 12 faecal bacterial taxa are enriched in CRC patients in two independent cohort studies. Among them, 2 Porphyromonas species are capable of inducing cellular senescence, an oncogenic stress response, through the secretion of the bacterial metabolite, butyrate. Notably, the invasion of these bacteria is observed in the CRC tissues, coinciding with the elevation of butyrate levels and signs of senescence-associated inflammatory phenotypes. Moreover, although the administration of these bacteria into ApcΔ14/+ mice accelerate the onset of colorectal tumours, this is not the case when bacterial butyrate-synthesis genes are disrupted. These results suggest a causal relationship between Porphyromonas species overgrowth and colorectal tumourigenesis which may be due to butyrate-induced senescence.


Assuntos
Bactérias/metabolismo , Butiratos/metabolismo , Carcinogênese/patologia , Neoplasias Colorretais/patologia , Microbioma Gastrointestinal , Bactérias/classificação , Bactérias/genética , Senescência Celular/fisiologia , Neoplasias Colorretais/microbiologia , Células Epiteliais/microbiologia , Células Epiteliais/fisiologia , Fezes/microbiologia , Humanos , Intestinos/citologia , Intestinos/microbiologia , Intestinos/fisiologia , Porphyromonas/genética , Porphyromonas/metabolismo , RNA Ribossômico 16S/genética
8.
Front Cell Dev Biol ; 9: 783857, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-35004683

RESUMO

Cells internalize proteins and lipids in the plasma membrane (PM) and solutes in the extracellular space by endocytosis. The removal of PM by endocytosis is constantly balanced by the replenishment of proteins and lipids to PM through recycling pathway. Recycling endosomes (REs) are specific subsets of endosomes. Besides the established role of REs in recycling pathway, recent studies have revealed unanticipated roles of REs in membrane traffic and cell signalling. In this review, we highlight these emerging issues, with a particular focus on phosphatidylserine (PS), a phospholipid that is highly enriched in the cytosolic leaflet of RE membranes. We also discuss the pathogenesis of Hermansky Pudlak syndrome type 2 (HPS2) that arises from mutations in the AP3B1 gene, from the point of view of dysregulated RE functions.

9.
Nat Commun ; 11(1): 1935, 2020 04 22.
Artigo em Inglês | MEDLINE | ID: mdl-32321921

RESUMO

Although cellular senescence acts primarily as a tumour suppression mechanism, the accumulation of senescent cells in vivo eventually exerts deleterious side effects through inflammatory/tumour-promoting factor secretion. Thus, the development of new drugs that cause the specific elimination of senescent cells, termed senolysis, is anticipated. Here, by an unbiased high-throughput screening of chemical compounds and a bio-functional analysis, we identify BET family protein degrader (BETd) as a promising senolytic drug. BETd provokes senolysis through two independent but integrated pathways; the attenuation of non-homologous end joining (NHEJ), and the up-regulation of autophagic gene expression. BETd treatment eliminates senescent hepatic stellate cells in obese mouse livers, accompanied by the reduction of liver cancer development. Furthermore, the elimination of chemotherapy-induced senescent cells by BETd increases the efficacy of chemotherapy against xenograft tumours in immunocompromised mice. These results reveal the vulnerability of senescent cells and open up possibilities for its control.


Assuntos
Antineoplásicos/administração & dosagem , Autofagia/efeitos dos fármacos , Senescência Celular/efeitos dos fármacos , Reparo do DNA por Junção de Extremidades/efeitos dos fármacos , Neoplasias/fisiopatologia , Animais , Proteínas de Ciclo Celular/antagonistas & inibidores , Proteínas de Ciclo Celular/genética , Proteínas de Ciclo Celular/metabolismo , Linhagem Celular Tumoral , Humanos , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Nus , Neoplasias/tratamento farmacológico , Neoplasias/genética , Neoplasias/metabolismo , Fatores de Transcrição/antagonistas & inibidores , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo
10.
Nat Commun ; 8(1): 1246, 2017 11 01.
Artigo em Inglês | MEDLINE | ID: mdl-29093443

RESUMO

Yes-associated protein (YAP) is a recently discovered growth-promoting transcription coactivator that has been shown to regulate the malignancy of various cancers. How YAP is regulated is not fully understood. Here, we show that one of the factors regulating YAP is phosphatidylserine (PS) in recycling endosomes (REs). We use proximity biotinylation to find proteins proximal to PS. Among these proteins are YAP and multiple proteins related to YAP signalling. Knockdown of ATP8A1 (an RE PS-flippase) or evectin-2 (an RE-resident protein) and masking of PS in the cytoplasmic leaflet of membranes, all suppress nuclear localization of YAP and YAP-dependent transcription. ATP8A1 knockdown increases the phosphorylated (activated) form of Lats1 that phosphorylates and inactivates YAP, whereas evectin-2 knockdown reduces the ubiquitination and increased the level of Lats1. The proliferation of YAP-dependent metastatic cancer cells is suppressed by knockdown of ATP8A1 or evectin-2. These results suggest a link between a membrane phospholipid and cell proliferation.


Assuntos
Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Adenosina Trifosfatases/genética , Proteínas de Membrana/genética , Fosfatidilserinas/metabolismo , Proteínas de Transferência de Fosfolipídeos/genética , Fosfoproteínas/metabolismo , Proteínas Serina-Treonina Quinases/metabolismo , Animais , Biotinilação , Células COS , Núcleo Celular/metabolismo , Proliferação de Células , Chlorocebus aethiops , Endossomos/metabolismo , Técnicas de Silenciamento de Genes , Células HEK293 , Humanos , Fosforilação , Transporte Proteico/genética , Transdução de Sinais , Fatores de Transcrição , Ubiquitinação , Proteínas de Sinalização YAP
11.
J Cell Sci ; 128(16): 3131-42, 2015 Aug 15.
Artigo em Inglês | MEDLINE | ID: mdl-26136365

RESUMO

The retrograde pathway is defined by the transport of proteins and lipids from the plasma membrane through endosomes to the Golgi complex, and is essential for a variety of cellular activities. Recycling endosomes are important sorting stations for some retrograde cargo. SMAP2, a GTPase-activating protein (GAP) for Arf1 with a putative clathrin-binding domain, has previously been shown to participate in the retrograde transport of the cholera toxin B-subunit (CTxB) from recycling endosomes. Here, we found that clathrin, a vesicle coat protein, and clathrin adaptor protein complex 1 (AP-1) were present at recycling endosomes and were needed for the retrograde transport of CTxB from recycling endosomes to the Golgi, but not from the plasma membrane to recycling endosomes. SMAP2 immunoprecipitated clathrin and AP-1 through a putative clathrin-binding domain and a CALM-binding domain, and SMAP2 mutants that did not interact with clathrin or AP-1 could not localize to recycling endosomes. Moreover, knockdown of Arf1 suppressed the retrograde transport of CTxB from recycling endosomes to the Golgi. These findings suggest that retrograde transport is mediated by clathrin-coated vesicles from recycling endosomes and that the role of the coat proteins is in the recruitment of Arf GAP to transport vesicles.


Assuntos
Toxina da Cólera/metabolismo , Clatrina/genética , Proteínas Ativadoras de GTPase/genética , Proteínas de Membrana/genética , Fator de Transcrição AP-1/genética , Fator 1 de Ribosilação do ADP/genética , Animais , Células COS , Membrana Celular/metabolismo , Chlorocebus aethiops , Clatrina/metabolismo , Endossomos/metabolismo , Proteínas Ativadoras de GTPase/metabolismo , Técnicas de Silenciamento de Genes , Complexo de Golgi/genética , Complexo de Golgi/metabolismo , Humanos , Proteínas de Membrana/metabolismo , Fator de Transcrição AP-1/metabolismo
12.
EMBO J ; 34(5): 669-88, 2015 Mar 04.
Artigo em Inglês | MEDLINE | ID: mdl-25595798

RESUMO

P4-ATPases translocate aminophospholipids, such as phosphatidylserine (PS), to the cytosolic leaflet of membranes. PS is highly enriched in recycling endosomes (REs) and is essential for endosomal membrane traffic. Here, we show that PS flipping by an RE-localized P4-ATPase is required for the recruitment of the membrane fission protein EHD1. Depletion of ATP8A1 impaired the asymmetric transbilayer distribution of PS in REs, dissociated EHD1 from REs, and generated aberrant endosomal tubules that appear resistant to fission. EHD1 did not show membrane localization in cells defective in PS synthesis. ATP8A2, a tissue-specific ATP8A1 paralogue, is associated with a neurodegenerative disease (CAMRQ). ATP8A2, but not the disease-causative ATP8A2 mutant, rescued the endosomal defects in ATP8A1-depleted cells. Primary neurons from Atp8a2-/- mice showed a reduced level of transferrin receptors at the cell surface compared to Atp8a2+/+ mice. These findings demonstrate the role of P4-ATPase in membrane fission and give insight into the molecular basis of CAMRQ.


Assuntos
Adenosina Trifosfatases/metabolismo , Endossomos/metabolismo , Modelos Biológicos , Proteínas de Transferência de Fosfolipídeos/metabolismo , Proteínas de Transporte Vesicular/metabolismo , Adenosina Trifosfatases/genética , Análise de Variância , Animais , Proteínas de Bactérias , Transporte Biológico/fisiologia , Western Blotting , Células COS , Chlorocebus aethiops , Primers do DNA/genética , DNA Complementar/genética , Células HeLa , Humanos , Imuno-Histoquímica , Camundongos , Camundongos Knockout , Microscopia Confocal , Fosfatidilserinas/metabolismo , Proteínas de Transferência de Fosfolipídeos/genética , Reação em Cadeia da Polimerase , Interferência de RNA , Estreptolisinas
13.
PLoS One ; 8(7): e69145, 2013.
Artigo em Inglês | MEDLINE | ID: mdl-23861959

RESUMO

Retrograde transport is where proteins and lipids are transported back from the plasma membrane (PM) and endosomes to the Golgi, and crucial for a diverse range of cellular functions. Recycling endosomes (REs) serve as a sorting station for the retrograde transport and we recently identified evection-2, an RE protein with a pleckstrin homology (PH) domain, as an essential factor of this pathway. How evection-2 regulates retrograde transport from REs to the Golgi is not well understood. Here, we report that evection-2 binds to SMAP2, an Arf GTPase-activating protein. Endogenous SMAP2 localized mostly in REs and to a lesser extent, the trans-Golgi network (TGN). SMAP2 binds evection-2, and the RE localization of SMAP2 was abolished in cells depleted of evection-2. Knockdown of SMAP2, like that of evection-2, impaired the retrograde transport of cholera toxin B subunit (CTxB) from REs. These findings suggest that evection-2 recruits SMAP2 to REs, thereby regulating the retrograde transport of CTxB from REs to the Golgi.


Assuntos
Endossomos/metabolismo , Complexo de Golgi/metabolismo , Proteínas de Membrana/metabolismo , Animais , Células COS , Chlorocebus aethiops , Toxina da Cólera/metabolismo , Técnicas de Silenciamento de Genes , Ligação Proteica , Transporte Proteico , Rede trans-Golgi/metabolismo
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