RESUMO
Malvaceae comprise some 4,225 species in 243 genera and nine subfamilies and include economically important species, such as cacao, cotton, durian, and jute, with cotton an important model system for studying the domestication of polyploids. Here, we use chromosome-level genome assemblies from representatives of five or six subfamilies (depending on the placement of Ochroma) to differentiate coexisting subgenomes and their evolution during the family's deep history. The results reveal that the allohexaploid Helicteroideae partially derive from an allotetraploid Sterculioideae and also form a component of the allodecaploid Bombacoideae and Malvoideae. The ancestral Malvaceae karyotype consists of 11 protochromosomes. Four subfamilies share a unique reciprocal chromosome translocation, and two other subfamilies share a chromosome fusion. DNA alignments of single-copy nuclear genes do not yield the same relationships as inferred from chromosome structural traits, probably because of genes originating from different ancestral subgenomes. These results illustrate how chromosome-structural data can unravel the evolutionary history of groups with ancient hybrid genomes.
Assuntos
Genoma de Planta , Gossypium , Genoma de Planta/genética , Gossypium/genética , Genômica/métodos , Poliploidia , Cariótipo , Evolução MolecularRESUMO
A substantial body of literature, including our own, points to a connection between hepatitis B virus (HBV) infection and the development of drug resistance in hepatocellular carcinoma (HCC), particularly against sorafenib. However, the influence of HBV on resistance to regorafenib, another therapeutic agent, has been less studied. In this study, we used the GEO database (GSE87630) and clinical samples to demonstrate that C-C motif chemokine receptor 9 (CCR9) was highly expressed in HBV-related HCC and predicted poor overall survival. Its overexpression correlated with HBsAg-positive HCC patients. Both univariate and multivariable Cox regression analysis elucidated CCR9 was an independent risk factor for poor overall survival in HCC patients. Our in vitro findings further revealed that HBV structural proteins, small HBV surface antigen (SHBs), triggered an upregulation of CCR9. Functional assays showed that SHBs enhanced HCC cell proliferation, migration, and invasion, increased ABCB1 and ABCC1 expression, and promoted regorafenib resistance via CCR9. Intriguingly, overexpression of HBV plasmid and an AAV-HBV mouse model both exhibited a significant elevation in global N6-methyladenosine (m6A) levels. Further investigations revealed that SHBs elevated these m6A levels, upregulated CCR9 and stabilized CCR9 mRNA through KIAA1429-mediated m6A modification, with sites 1373 and 1496 on CCR9 mRNA being critical for modification. In conclusion, SHBs promoted HCC progression and regorafenib resistance via KIAA1429-mediated m6A modification of CCR9. Our findings suggested that CCR9 could be a potential prognostic biomarker and a valuable molecular therapeutic target of regorafenib resistance in HBV-related HCC.
Assuntos
Carcinoma Hepatocelular , Resistencia a Medicamentos Antineoplásicos , Antígenos de Superfície da Hepatite B , Neoplasias Hepáticas , Compostos de Fenilureia , Piridinas , Carcinoma Hepatocelular/virologia , Carcinoma Hepatocelular/tratamento farmacológico , Humanos , Neoplasias Hepáticas/virologia , Neoplasias Hepáticas/tratamento farmacológico , Resistencia a Medicamentos Antineoplásicos/genética , Animais , Antígenos de Superfície da Hepatite B/genética , Antígenos de Superfície da Hepatite B/metabolismo , Piridinas/farmacologia , Piridinas/uso terapêutico , Compostos de Fenilureia/farmacologia , Compostos de Fenilureia/uso terapêutico , Camundongos , Masculino , Feminino , Receptores CCR/genética , Receptores CCR/metabolismo , Linhagem Celular Tumoral , Vírus da Hepatite B/genética , Vírus da Hepatite B/efeitos dos fármacos , Pessoa de Meia-Idade , Hepatite B/virologia , Hepatite B/complicações , Hepatite B/tratamento farmacológico , Antineoplásicos/farmacologia , Antineoplásicos/uso terapêutico , Proliferação de Células/efeitos dos fármacos , Adenosina/análogos & derivadosRESUMO
In pursuit of enhancing the anti-resistance efficacy and solubility of our previously identified NNRTI 1, a series of biphenyl-quinazoline derivatives were synthesized employing a structure-based drug design strategy. Noteworthy advancements in anti-resistance efficacy were discerned among some of these analogs, prominently exemplified by compound 7ag, which exhibited a remarkable 1.37 to 602.41-fold increase in potency against mutant strains (Y181C, L100I, Y188L, F227L + V106A, and K103N + Y181C) in comparison to compound 1. Compound 7ag also demonstrated comparable anti-HIV activity against both WT HIV and K103N, albeit with a marginal reduction in activity against E138K. Of significance, this analog showed augmented selectivity index (SI > 5368) relative to compound 1 (SI > 37764), Nevirapine (SI > 158), Efavirenz (SI > 269), and Etravirine (SI > 1519). Moreover, it displayed a significant enhancement in water solubility, surpassing that of compound 1, Etravirine, and Rilpivirine. To elucidate the underlying molecular mechanisms, molecular docking studies were undertaken to probe the critical interactions between 7ag and both WT and mutant strains of HIV-1 RT. These findings furnish invaluable insights driving further advancements in the development of DAPYs for HIV therapy.
Assuntos
Fármacos Anti-HIV , Compostos de Bifenilo , Desenho de Fármacos , Transcriptase Reversa do HIV , HIV-1 , Quinazolinas , Inibidores da Transcriptase Reversa , Solubilidade , Humanos , Fármacos Anti-HIV/farmacologia , Fármacos Anti-HIV/química , Fármacos Anti-HIV/síntese química , Compostos de Bifenilo/antagonistas & inibidores , Compostos de Bifenilo/farmacologia , Compostos de Bifenilo/química , Relação Dose-Resposta a Droga , Farmacorresistência Viral/efeitos dos fármacos , Transcriptase Reversa do HIV/antagonistas & inibidores , Transcriptase Reversa do HIV/metabolismo , HIV-1/efeitos dos fármacos , HIV-1/enzimologia , Testes de Sensibilidade Microbiana , Simulação de Acoplamento Molecular , Estrutura Molecular , Quinazolinas/farmacologia , Quinazolinas/química , Quinazolinas/síntese química , Inibidores da Transcriptase Reversa/farmacologia , Inibidores da Transcriptase Reversa/química , Inibidores da Transcriptase Reversa/síntese química , Relação Estrutura-AtividadeRESUMO
Targeting Ribonuclease H (RNase H) has been considered a viable strategy for HIV therapy. In this study, a series of novel thiazolo[3, 2-a]pyrimidine derivatives were firstly designed and synthesized as potential inhibitors of HIV-1 RNase H. Among these compounds, A28 exhibited the most potent inhibition against HIV-1 RNase H with an IC50 value of 4.14 µM, which was about 5-fold increase in potency than the hit compound A1 (IC50 = 21.49 µM). To gain deeper insights into the structure-activity relationship (SAR), a CoMFA model was constructed to yield reasonable statistical results (q2 = 0.658 and R2 = 0.969). Results from magnesium ion chelation experiments and molecular docking studies revealed that these thiazolopyrimidine inhibitors may exert their inhibitory activity by binding to an allosteric site on RNase H at the interface between subunits p51 and p66. Furthermore, this analog demonstrated favorable physicochemical properties. Our findings provide valuable groundwork for further development of allosteric inhibitors targeting HIV-1 RNase H.
Assuntos
Desenho de Fármacos , HIV-1 , Simulação de Acoplamento Molecular , Pirimidinas , Relação Estrutura-Atividade , Pirimidinas/química , Pirimidinas/farmacologia , Pirimidinas/síntese química , HIV-1/efeitos dos fármacos , HIV-1/enzimologia , Humanos , Tiazóis/química , Tiazóis/farmacologia , Tiazóis/síntese química , Estrutura Molecular , Fármacos Anti-HIV/farmacologia , Fármacos Anti-HIV/síntese química , Fármacos Anti-HIV/química , Ribonuclease H/antagonistas & inibidores , Ribonuclease H/metabolismo , Relação Dose-Resposta a Droga , Inibidores Enzimáticos/farmacologia , Inibidores Enzimáticos/síntese química , Inibidores Enzimáticos/química , Ribonuclease H do Vírus da Imunodeficiência Humana/antagonistas & inibidores , Ribonuclease H do Vírus da Imunodeficiência Humana/metabolismoRESUMO
Acetaminophen (N-acetyl-p-aminophenol; APAP) is one of the most widely used analgesics. To examine the toxicity of APAP, we used zebrafish embryos as model animals to detect the effect of APAP on the thyroid system of zebrafish embryos. The zebrafish embryos were exposed to APAP from 4 h post fertilization (4 hpf) until observation. The experimental results showed that APAP caused pericardial edema and decreased pigmentation in the zebrafish embryos or larvae. The APAP treatment caused a decrease in the expression of tpo and thrß in the zebrafish at 36 and 72 hpf. The transcriptomic analysis found that APAP affected retinol metabolism, the metabolism of xenobiotics by cytochrome P450, and the tyrosine metabolism pathway. The harmful effect of APAP on zebrafish embryos might be due to its disrupting effect on the functional regulation of the thyroid hormone system.
Assuntos
Acetaminofen , Perciformes , Animais , Acetaminofen/toxicidade , Peixe-Zebra , Tiroxina , Pigmentação , Glândula TireoideRESUMO
BACKGROUND: Eutrema salsugineum (2n = 14), a halophyte in the family Brassicaceae, is an attractive model to study abiotic stress tolerance in plants. Two versions of E. salsugineum genomes that previously reported were based on relatively short reads; thus, the repetitive regions were difficult to characterize. RESULTS: We report the sequencing and assembly of the E. salsugineum (Shandong accession) genome using long-read sequencing and chromosome conformation capture data. We generated Oxford Nanopore long reads at high depth (> 60X) of genome coverage with additional short reads for error correction. The new assembly has a total size of 295.5 Mb with 52.8% repetitive sequences, and the karyotype of E. salsugineum is consistent with the ancestral translocation Proto-Calepineae Karyotype structure in both order and orientation. Compared with previous assemblies, this assembly has higher contiguity, especially in the centromere region. Based on this new assembly, we predicted 25,399 protein-coding genes and identified the positively selected genes associated with salt and drought stress responses. CONCLUSION: The new genome assembly will provide a valuable resource for future genomic studies and facilitate comparative genomic analysis with other plants.
Assuntos
Brassicaceae , Extremófilos , Brassicaceae/genética , Genômica , Estresse Fisiológico , CromossomosRESUMO
In this Letter, a polymer optical fiber (POF) detector with a convex spherical aperture microstructure probe is designed for low-energy and low-dose rate gamma-ray detection. Simulation and experimental results demonstrate that this structure has a higher optical coupling efficiency and that the angular coherence of the detector depends strongly on the depth of the probe micro-aperture. By modeling the relationship between angular coherence and micro-aperture depth, the optimal depth of the micro-aperture is determined. The sensitivity of the fabricated POF detector is 701 cps at 59.5-keV gamma-ray of 2.78 µSv/h and the maximum percentage error of the average count rate at different angles is 5.16%.
RESUMO
The spectacular nature of non-wetting drops mainly arises from their extreme mobility, and quick-silver, for instance, was named after this property. There are two ways to make water non-wetting, and they both rely on texture: either we can roughen a hydrophobic solid, which makes drops looking like pearls, or we can texture the liquid with a hydrophobic powder that "isolates" the resulting marble from its substrate. We observe, here, races between pearls and marbles, and report two effects: (1) the static adhesion of the two objects is different in nature, which we interpret as a consequence of the way they meet their substrates; (2) when they move, pearls are generally quicker than marbles, which might arise from the dissimilarity of the liquid/air interface between these two kinds of globules.
RESUMO
Idiopathic intracranial hypertension (IIH) is a disorder characterized by elevated intracranial pressure (ICP) that predominantly affects young obese women. IIH is a diagnosis of exclusion. That is, if increased ICP is suspected, magnetic resonance imaging and magnetic resonance venography of the brain are recommended to exclude secondary causes. Imaging findings, such as empty sella, orbital findings, meningocele, and encephalocele, are not diagnostic of ICP, nor does their absence exclude ICP either. Therefore, venous manometry is recommended as the gold standard for evaluation, regardless of previous anatomic imaging results. Venous manometry is an invasive examination that is frequently applied to derive physiologic information concerning the nature of the pressure gradient. However, the pathogenesis of IIH has not been fully elucidated. The presence of venous sinus stenosis in a subset of patients has provided some support for the potential mechanisms underlying this condition. Hence, this review provides an up-to-date discussion on the potential pathogenic mechanisms of IIH with a special focus on venous sinus stenosis. LEVEL OF EVIDENCE: 1 TECHNICAL EFFICACY STAGE: 2.
Assuntos
Hipertensão Intracraniana , Pseudotumor Cerebral , Doenças Vasculares , Constrição Patológica/diagnóstico por imagem , Cavidades Cranianas/diagnóstico por imagem , Feminino , Humanos , Hipertensão Intracraniana/diagnóstico por imagem , Flebografia , Pseudotumor Cerebral/complicações , Pseudotumor Cerebral/diagnóstico por imagem , StentsRESUMO
Compatible solutes are key for the ability of halophilic bacteria to resist high osmotic stress. They have received wide attention from researchers for their excellent osmotic protection properties. Hydroxyectoine is a particularly important compatible solute, but its production by microbes faces several challenges, including low titer/yield, the presence of the byproduct ectoine, and the requirement of high salinity. Here, we aimed to metabolically engineer Escherichia coli to efficiently produce hydroxyectoine in the absence of osmotic stress without accumulating the byproduct ectoine. First, combinatorial optimization of the expression strength of key genes in the ectoine synthesis module and hydroxyectoine synthesis module was conducted. After optimization of the expression of these genes, 12.12 g/L hydroxyectoine and 0.24 g/L ectoine were obtained at 36 h in shake-flask fermentation with the addition of the co-substrate α-ketoglutarate. Further optimization of the addition of α-ketoglutarate achieved the sole production of hydroxyectoine (i.e., no ectoine accumulation), indicating that the supply of α-ketoglutarate is critically important for sole hydroxyectoine production. Finally, quorum sensing-based auto-regulation of intracellular α-ketoglutarate pool was implemented as an alternative to α-ketoglutarate addition by coupling the expression of sucA with the esaI/esaR circuit, which led to 14.93 g/L hydroxyectoine with a unit cell yield of 1.678 g/g and no ectoine accumulation in the absence of osmotic stress. This is the highest reported titer of sole hydroxyectoine production under salinity-free fermentation to date.
Assuntos
Diamino Aminoácidos/metabolismo , Escherichia coli , Engenharia Metabólica/métodos , Escherichia coli/genética , Escherichia coli/metabolismo , Ácidos Cetoglutáricos/metabolismo , Pressão Osmótica , Percepção de QuorumRESUMO
In vitro fermentation was used to evaluate the possible effects of intervention with Lactobacillus paracasei N1115 (LP N1115) on gut microbiota and metabolite shortchain fatty acids (SCFAs) in pregnant women with constipation and diarrhea. Feces were collected from pregnant women and fermented by YCFA medium to profile the changes in the gut microbiota before and after intervention with LP N1115 using 16SrRNA sequencing. At the same time, the changes in several specific bacteria were detected using quantitative real-time PCR (qPCR) and the SCFAs in fermentation were detected using gas chromatography (GC) for each subject to determine the effect of the intervention. In vitro intervention with LP N1115 significantly increased the relative abundances of Lactobacillus, Faecalibacterium prausnitzii, and Bifidobacterium in constipated pregnant women and reduced the contents of acetic acid, propanoic acid. Moreover, 16S rRNA gene analysis showed that LP N1115 also reduced the relative abundance of Clostridium_XI. The results of this study suggest that LP N1115 might increase the content of beneficial bacteria and reduce the relative abundance of pathogenic bacteria, which might be beneficial to gut health in pregnant women.
Assuntos
Lacticaseibacillus paracasei , Microbiota , Bactérias , Constipação Intestinal , Diarreia , Fezes/microbiologia , Feminino , Humanos , Lacticaseibacillus paracasei/genética , Gravidez , Gestantes , RNA Ribossômico 16S/genética , RNA Ribossômico 16S/metabolismoRESUMO
Through urban traffic patrols, problems such as traffic congestion and accidents can be found and dealt with in time to maintain the stability of the urban traffic system. The most common way to patrol is using ground vehicles, which may be inflexible and inefficient. The vehicle-drone coordination maximizes utilizing the flexibility of drones and addresses their limited battery capacity issue. This paper studied a vehicle-drone arc routing problem (VD-ARP), consisting of one vehicle and multiple drones. Considering the coordination mode and constraints of the vehicle-drone system, a mathematical model of VD-ARP that minimized the total patrol time was constructed. To solve this problem, an improved, adaptive, large neighborhood search algorithm (IALNS) was proposed. First, the initial route planning scheme was generated by the heuristic rule of "Drone-First, Vehicle-Then". Then, several problem-based neighborhood search strategies were embedded into the improved, adaptive, large neighborhood search framework to improve the quality of the solution. The superiority of IALNS is verified by numerical experiments on instances with different scales. Several critical factors were tested to determine the effects of coordinated traffic patrol; an example based on a real road network verifies the feasibility and applicability of the algorithm.
Assuntos
Algoritmos , Dispositivos Aéreos não Tripulados , Fontes de Energia Elétrica , Heurística , Modelos TeóricosRESUMO
Aquaponics is a green and efficient agricultural production model that combines aquaculture and vegetable cultivation. It is worth looking into optimizing the proportion of fish and plants to improve the quality and yield. However, there is little non-destructive monitoring of plant growth in aquaponics monitoring systems currently. In this paper, based on the Internet of Things technologies, a monitoring system is designed with miniaturization, modularization, and low-cost features for cultivation-breeding ratio research. The system can realize remote monitoring and intelligent control of parameters needed to keep fish and plants under optimal conditions. First, a 32-bit chip is used as the Microcontroller Unit to develop the intelligent sensing unit, which can realize 16 different data acquisitions as stand-alone extensible modules. Second, to achieve plant data acquisition and upload, the Raspberry Pi embedded with image processing algorithms is introduced to realize edge-computing. Finally, all the collected data is stored in the Ali-cloud through Wi-Fi and a WeChat Mini Program is designed to display data and control devices. The results show that there is no packet loss within 90 m for wireless transmission, and the error rate of environment parameters is limited to 5%. It was proven that the system is intelligent, flexible, low-cost, and stable which is suitable for small-scale aquaponics well.
Assuntos
Eletrocardiografia , Melhoramento Vegetal , Animais , Aquicultura/métodos , AlgoritmosRESUMO
PURPOSE OF REVIEW: Cholesterol is delivered to the limiting membrane of late endosomes by Niemann-Pick Type C1 and C2 proteins. This review summarizes recent evidence that cholesterol transfer from endosomes to the endoplasmic reticulum and other organelles is mediated by lipid-binding proteins that localize to membrane contact sites (MCS). RECENT FINDINGS: LDL-cholesterol in the late endosomal/lysosomes is exported to the plasma membrane, where most cholesterol resides, and the endoplasmic reticulum, which harbors the regulatory complexes and enzymes that control the synthesis and esterification of cholesterol. A major advance in dissecting these cholesterol transport pathways was identification of frequent and dynamic MCS between endosomes and the endoplasmic reticulum, peroxisomes and plasma membrane. Positioned at these MCS are members of the oxysterol-binding protein (OSBP) and steroidogenic acute regulatory protein-related lipid-transfer family of lipid transfer proteins that bridge the opposing membranes and directly or indirectly mediate cholesterol transfer. OSBP-related protein 1L (ORP1L), ORP5 and ORP6 mediate cholesterol transfer to the endoplasmic reticulum that regulates cholesterol homeostasis. ORP1L and STARD3 also move cholesterol from the endoplasmic reticulum-to-late endosomal/lysosomes under low-cholesterol conditions to facilitate intraluminal vesicle formation. Cholesterol transport also occurs at MCS with peroxisomes and possibly the plasma membrane. SUMMARY: Frequent contacts between organelles and the endo-lysosomal vesicles are sites for bidirectional transfer of cholesterol.
Assuntos
Membrana Celular/metabolismo , LDL-Colesterol/metabolismo , Retículo Endoplasmático/metabolismo , Endossomos/metabolismo , Lisossomos/metabolismo , Animais , Transporte Biológico Ativo , Proteínas de Transporte/genética , Proteínas de Transporte/metabolismo , LDL-Colesterol/genética , Retículo Endoplasmático/genética , Endossomos/genética , Humanos , Lisossomos/genética , Proteínas de Membrana/genética , Proteínas de Membrana/metabolismo , Proteína C1 de Niemann-Pick/metabolismo , Peroxissomos/genética , Peroxissomos/metabolismo , Receptores de Esteroides/genética , Receptores de Esteroides/metabolismoRESUMO
Microdeletions in 19q12q13.12 cause a rare and complex haploinsufficiency syndrome characterized by intellectual deficiency, developmental delays, and neurological movement disorders. Variability in the size and interval of the deletions makes it difficult to attribute the complex clinical phenotype of this syndrome to an underlying gene(s). As an alternate approach, we examined the biochemical and metabolic features of fibroblasts from an affected individual to derive clues as to the molecular basis for the syndrome. Immunofluorescence and electron microscopy of affected fibroblasts revealed an abnormal endo-lysosomal compartment that was characterized by rapid accumulation of lysosomotropic dyes, elevated LAMP1 and LAMP2 expression and vacuoles containing membrane whorls, common features of lysosomal lipid storage disorders. The late endosomes-lysosomes (LE/LY) of affected fibroblasts accumulated low-density lipoprotein cholesterol, and displayed reduced cholesterol esterification and increased de novo cholesterol synthesis, indicative of defective cholesterol transport to the endoplasmic reticulum. Affected fibroblasts also had increased ceramide and sphingolipid mass, altered glycosphingolipid species and accumulation of a fluorescent lactosylceramide probe in LE/LY. Autophagosomes also accumulated in affected fibroblasts because of decreased fusion with autolysosomes, a defect associated with other lysosomal storage diseases. Attempts to correct the cholesterol/sphingolipid storage defect in fibroblasts with cyclodextrin, sphingolipid synthesis inhibitors or by altering ion transport were unsuccessful. Our data show that 19q13.12 deletion fibroblasts have abnormal accumulation of cholesterol and sphingolipids in the endo-lysosomal system that compromises organelle function and could be an underlying cause of the clinical features of the syndrome.
Assuntos
Deleção Cromossômica , Cromossomos Humanos Par 19/genética , Fibroblastos/patologia , Doenças por Armazenamento dos Lisossomos/patologia , Doenças Raras/patologia , Células Cultivadas , Criança , Colesterol/metabolismo , Retículo Endoplasmático/metabolismo , Endossomos/metabolismo , Endossomos/patologia , Endossomos/ultraestrutura , Fibroblastos/citologia , Fibroblastos/metabolismo , Fibroblastos/ultraestrutura , Humanos , Doenças por Armazenamento dos Lisossomos/diagnóstico , Doenças por Armazenamento dos Lisossomos/genética , Lisossomos/metabolismo , Lisossomos/patologia , Lisossomos/ultraestrutura , Masculino , Microscopia Eletrônica de Transmissão , Doenças Raras/diagnóstico , Doenças Raras/genética , Esfingolipídeos/metabolismo , Síndrome , Sequenciamento Completo do GenomaRESUMO
The repair of DNA damage is fundamental to normal cell development and replication. Hydrogen sulfide (H2S) is a novel gasotransmitter that has been reported to protect cellular aging. Here, we show that H2S attenuates DNA damage in human endothelial cells and fibroblasts by S-sulfhydrating MEK1 at cysteine 341, which leads to PARP-1 activation. H2S-induced MEK1 S-sulfhydration facilitates the translocation of phosphorylated ERK1/2 into nucleus, where it activates PARP-1 through direct interaction. Mutation of MEK1 cysteine 341 inhibits ERK phosphorylation and PARP-1 activation. In the presence of H2S, activated PARP-1 recruits XRCC1 and DNA ligase III to DNA breaks to mediate DNA damage repair, and cells are protected from senescence.
Assuntos
Dano ao DNA , Reparo do DNA , MAP Quinase Quinase 1/metabolismo , Poli(ADP-Ribose) Polimerases/metabolismo , Dano ao DNA/efeitos dos fármacos , Reparo do DNA/efeitos dos fármacos , Ativação Enzimática/efeitos dos fármacos , Humanos , Sulfeto de Hidrogênio/farmacologia , MAP Quinase Quinase 1/genética , Sistema de Sinalização das MAP Quinases , Modelos Biológicos , Fosforilação , Poli(ADP-Ribose) Polimerase-1 , Processamento de Proteína Pós-TraducionalRESUMO
Oxygen-sensitive accumulation and degradation, two opposite but intrinsically linked events, of heme proteins in mitochondria affect mitochondrial functions, including bioenergetics and oxygen-sensing processes. Cystathionine ß-synthase (CBS) contains a prosthetic heme group and catalyzes the production of hydrogen sulfide in mammalian cells. Here we show that CBS proteins were present in liver mitochondria at a low level under normoxia conditions. Ischemia/hypoxia increased the accumulation of CBS proteins in mitochondria. The normalization of oxygen partial pressure accelerated the degradation of CBS proteins. Lon protease, a major degradation enzyme in mitochondrial matrix, recognized and degraded mitochondrial CBS by specifically targeting at the oxygenated heme group of CBS proteins. The accumulation of CBS in mitochondria increased hydrogen sulfide production, which prevented Ca(2+)-mediated cytochrome c release from mitochondria and decreased reactive oxygen species generation. Mitochondrial accumulation of heme oxygenase-1, another heme protein, was also regulated by oxygen level and Lon protease in the same mechanism as for CBS. Our findings provide a fundamental and general mechanism for oxygen-sensitive regulation of mitochondrial functions by linking oxygenation level to the accumulation/degradation of mitochondrial heme proteins.
Assuntos
Cistationina beta-Sintase/metabolismo , Mitocôndrias Hepáticas/metabolismo , Proteínas Mitocondriais/metabolismo , Oxigênio/metabolismo , Protease La/metabolismo , Proteólise , Animais , Células HEK293 , Heme/metabolismo , Heme Oxigenase (Desciclizante)/metabolismo , Humanos , Sulfeto de Hidrogênio/metabolismo , Masculino , Ratos , Ratos Sprague-Dawley , Espécies Reativas de Oxigênio/metabolismoRESUMO
Androgen receptor (AR) signaling is indispensable for the development of prostate cancer from the initial androgen-dependent state to a later aggressive androgen-resistant state. This study examined the role of hydrogen sulfide (H(2)S), a novel gasotransmitter, in the regulation of AR signaling as well as its mediation in androgen-independent cell growth in prostate cancer cells. Here we found that H(2)S inhibits cell proliferation of both androgen-dependent (LNCaP) and antiandrogen-resistant prostate cancer cells (LNCaP-B), with more significance on the latter, which was established by long term treatment of parental LNCaP cells with bicalutamide. The expression of cystathionine γ-lyase (CSE), a major H(2)S producing enzyme in prostate tissue, was reduced in both human prostate cancer tissues and LNCaP-B cells. LNCaP-B cells were resistant to bicalutamide-induced cell growth inhibition, and CSE overexpression could rebuild the sensitivity of LNCaP-B cells to bicalutamide. H(2)S significantly repressed the expression of prostate-specific antigen (PSA) and TMPRSS2, two AR-targeted genes. In addition, H(2)S inhibited AR binding with PSA promoter and androgen-responsive element (ARE) luciferase activity. We further found that AR is post-translationally modified by H(2)S through S-sulfhydration. Mutation of cysteine 611 and cysteine 614 in the second zinc finger module of AR-DNA binding domain diminished the effects of H(2)S on AR S-sulfhydration and AR dimerization. These data suggest that reduced CSE/H2S signaling contributes to antiandrogen-resistant status, and sufficient level of H(2)S is able to inhibit AR transactivation and treat castration-resistant prostate cancer.
Assuntos
Gasotransmissores/farmacologia , Sulfeto de Hidrogênio/farmacologia , Proteínas de Neoplasias/metabolismo , Neoplasias da Próstata/metabolismo , Receptores Androgênicos/metabolismo , Ativação Transcricional/efeitos dos fármacos , Animais , Linhagem Celular Tumoral , Humanos , Calicreínas/genética , Calicreínas/metabolismo , Masculino , Camundongos , Camundongos Knockout , Mutação , Proteínas de Neoplasias/genética , Antígeno Prostático Específico/genética , Antígeno Prostático Específico/metabolismo , Neoplasias da Próstata/genética , Neoplasias da Próstata/terapia , Multimerização Proteica/efeitos dos fármacos , Multimerização Proteica/genética , Receptores Androgênicos/genética , Serina Endopeptidases/genética , Serina Endopeptidases/metabolismo , Ativação Transcricional/genética , Dedos de ZincoRESUMO
P2X receptors are commonly known as plasma membrane cation channels involved in a wide variety of cell functions. The properties of these channels have been extensively studied on the plasma membrane. However, studies in amoeba suggest that P2X receptors are also present intracellularly and involved in vesicle fusion with the plasma membrane. Recently, it was shown that in addition to plasma membrane expression, mammalian P2X4 was also localized intracellularly in lysosomes. However, it was not clear whether the lysosomal P2X4 receptors function as channels and how they are activated and regulated. In this paper, we show that both P2X4 and its natural ligand, ATP, are enriched in lysosomes of COS1 and HEK293 cells. By directly recording membrane currents from enlarged lysosomal vacuoles, we demonstrated that lysosomal P2X4 formed channels activated by ATP from the luminal side in a pH-dependent manner. While the acidic pH at the luminal side inhibited P2X4 activity, increasing the luminal pH in the presence of ATP caused P2X4 activation. We further showed that, as for the plasma membrane P2X4, the lysosomal P2X4 was potentiated by ivermectin but insensitive to suramin and PPADS, and it permeated the large cation N-methyl-d-glucamine upon activation. Our data suggest that P2X4 forms functional ATP-activated cation channels on lysosomal membranes regulated by luminal pH. Together with the reported fusion effect of intracellular P2X in lower organisms, we speculate that the lysosome-localized P2X4 may play specific roles in membrane trafficking of acidic organelles in mammalian cells.
Assuntos
Membranas Intracelulares/metabolismo , Lisossomos/metabolismo , Receptores Purinérgicos P2X4/metabolismo , Trifosfato de Adenosina/genética , Trifosfato de Adenosina/metabolismo , Animais , Antinematódeos/farmacologia , Transporte Biológico Ativo/efeitos dos fármacos , Transporte Biológico Ativo/fisiologia , Células COS , Chlorocebus aethiops , Glutamatos/farmacologia , Células HEK293 , Humanos , Concentração de Íons de Hidrogênio , Lisossomos/genética , Inibidores da Agregação Plaquetária/farmacologia , Fosfato de Piridoxal/análogos & derivados , Fosfato de Piridoxal/farmacologia , Ratos , Receptores Purinérgicos P2X4/genética , Suramina/farmacologiaRESUMO
Lysosomes contain abundant ATP, which is released through lysosomal exocytosis following exposure to various stimuli. However, the molecular mechanisms underlying lysosomal ATP accumulation remain unknown. The vesicular nucleotide transporter, also known as solute carrier family 17 member 9 (SLC17A9), has been shown to function in ATP transport across secretory vesicles/granules membrane in adrenal chromaffin cells, T cells, and pancreatic cells. Here, using mammalian cell lines, we report that SLC17A9 is highly enriched in lysosomes and functions as an ATP transporter in those organelles. SLC17A9 deficiency reduced lysosome ATP accumulation and compromised lysosome function, resulting in cell death. Our data suggest that SLC17A9 activity mediates lysosomal ATP accumulation and plays an important role in lysosomal physiology and cell viability.