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In search of the molecular identities of cold-sensing receptors, we carried out an unbiased genetic screen for cold-sensing mutants in C. elegans and isolated a mutant allele of glr-3 gene that encodes a kainate-type glutamate receptor. While glutamate receptors are best known to transmit chemical synaptic signals in the CNS, we show that GLR-3 senses cold in the peripheral sensory neuron ASER to trigger cold-avoidance behavior. GLR-3 transmits cold signals via G protein signaling independently of its glutamate-gated channel function, suggesting GLR-3 as a metabotropic cold receptor. The vertebrate GLR-3 homolog GluK2 from zebrafish, mouse, and human can all function as a cold receptor in heterologous systems. Mouse DRG sensory neurons express GluK2, and GluK2 knockdown in these neurons suppresses their sensitivity to cold but not cool temperatures. Our study identifies an evolutionarily conserved cold receptor, revealing that a central chemical receptor unexpectedly functions as a thermal receptor in the periphery.
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Proteínas de Caenorhabditis elegans/fisiología , Caenorhabditis elegans/genética , Receptores de Glutamato/fisiología , Receptores de Ácido Kaínico/fisiología , Receptores de Glutamato Metabotrópico/fisiología , Sensación Térmica/fisiología , Animales , Células CHO , Proteínas de Caenorhabditis elegans/genética , Frío , Cricetulus , Humanos , Ratones , Neuronas/metabolismo , Receptores de Glutamato/genética , Receptores de Ácido Kaínico/genética , Receptores de Glutamato Metabotrópico/genética , Sensación Térmica/genéticaRESUMEN
Pain information processing in the spinal cord has been postulated to rely on nociceptive transmission (T) neurons receiving inputs from nociceptors and Aß mechanoreceptors, with Aß inputs gated through feed-forward activation of spinal inhibitory neurons (INs). Here, we used intersectional genetic manipulations to identify these critical components of pain transduction. Marking and ablating six populations of spinal excitatory and inhibitory neurons, coupled with behavioral and electrophysiological analysis, showed that excitatory neurons expressing somatostatin (SOM) include T-type cells, whose ablation causes loss of mechanical pain. Inhibitory neurons marked by the expression of dynorphin (Dyn) represent INs, which are necessary to gate Aß fibers from activating SOM(+) neurons to evoke pain. Therefore, peripheral mechanical nociceptors and Aß mechanoreceptors, together with spinal SOM(+) excitatory and Dyn(+) inhibitory neurons, form a microcircuit that transmits and gates mechanical pain. PAPERCLIP:
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Neuronas/fisiología , Dolor/metabolismo , Médula Espinal/fisiología , Animales , Dinorfinas/metabolismo , Mecanorreceptores/metabolismo , Ratones , Percepción del Dolor , Somatostatina/metabolismoRESUMEN
MOTIVATION: Molecular representation learning is pivotal for advancing deep learning applications in quantum chemistry and drug discovery. Existing methods for molecular representation learning often fall short of fully capturing the intricate interactions within chemical bonds of 2D topological graphs and the multifaceted effects of 3D geometric conformations. RESULTS: To overcome these challenges, we present a novel contrastive learning strategy for molecular representation learning, named Geometric Triangle Awareness Model (GTAM). This method integrates innovative molecular encoders for both 2D graphs and 3D conformations, enabling the accurate capture of geometric dependencies among edges in graph-based molecular structures. Furthermore, GTAM is bolstered by the development of two contrastive training objectives designed to facilitate the direct transfer of edge information between 2D topological graphs and 3D geometric conformations, enhancing the functionality of the molecular encoders. Through extensive evaluations on a range of 2D and 3D downstream tasks, our model has demonstrated superior performance over existing approaches. AVAILABILITY: The test code and data of GTAM are available online at https://github.com/StellaHxy/GTAM. SUPPLEMENTARY INFORMATION: Comprehensive implementation details, datasets, and additional experiments can be found in Supplementary Materials A-C.
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Multiscale defects engineering offers a promising strategy for synergistically enhancing the thermoelectric and mechanical properties of thermoelectric semiconductors. However, the specific impact of individual defects, in particular precipitation, on mechanical properties remains ambiguous. In this work, the mechanical and thermoelectric properties of Sn1.03- xMnxTe (x = 0-0.30) semiconductors are systematically studied. Mn-alloying induces dense dislocations and Mn nano-precipitates, resulting in an enhanced compressive strength with x increased to 0.15. Quantitative calculations are performed to assess the strengthening contributions including grain boundary, solid solution, dislocation, and precipitation strengthening. Due to the dominant contribution of precipitation strengthening, the yield strength of the x = 0.10 sample is improved by ≈74.5% in comparison to the Mn-free Sn1.03Te. For x ≥ 0.15, numerous MnTe precipitates lead to a synergistic enhancement of strength-ductility. In addition, multiscale defects induced by Mn alloying can scatter phonons over a wide frequency spectrum. The peak figure of merit ZT of ≈1.3 and an ultralow lattice thermal conductivity of ≈0.35 Wm-1 K-1 are obtained at 873 K for x = 0.10 and x = 0.30 samples respectively. This work reveals tha precipitation evolution optimizes the mechanical and thermoelectric properties of Sn1.03- xMnxTe semiconductors, which may hold potential implications for other thermoelectric systems.
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Seven new lanthanide coordination polymers, namely [Ln(cpt)3H2O)]n(Ln = La (1), Pr (2), Sm (3), Eu (4), Gd (5), Dy (6), and Er (7)), which were synthesized under hydrothermal conditions using 4'-(4-(4-carboxyphenyloxy)phenyl)-4,2':6',4'-tripyridine (Hcpt) as the ligand. The crystal structures of these seven complexes were determined using single-crystal X-ray diffraction, and they were found to be isostructural, crystallizing in the triclinic P1- space group. The Ln(III) ions were nine-coordinated with tricapped trigonal prism coordination geometry. The Ln(III) cations were coordinated by carboxylic and pyridine groups from (cpt)- ligands, forming one-dimensional ring-chain structures. Furthermore, the luminescent properties of complexes 1-7 were investigated using fluorescent spectra in the solid state. The fluorescence sensing experiments demonstrated that complex 4 exhibits high selectivity and sensitivity for detecting Co2+, Cu2+ ions, and nitrobenzene. Moreover, complex 3 shows good capability for detecting Cu2+ ions and nitrobenzene. Additionally, the sensing mechanism was also thoroughly examined through theoretical calculations.
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PURPOSE: This study aims to evaluate the safety and efficacy of ultrasound-guided balloon dilation compared to non-balloon dilation for percutaneous nephrolithotomy (PCNL). MATERIALS AND METHODS: A systematic review and meta-analysis were conducted by searching PubMed, EMBASE, and the Cochrane Library. Results were filtered using predefined inclusion and exclusion criteria as described and meta-analysis was performed using Review Manager 5.4 software. RESULTS: A total of six studies involving 1189 patients who underwent PCNL were included. The meta-analysis results demonstrated that compared to non-balloon dilation, balloon dilation was associated with reduced haemoglobin drop [mean difference (MD) = -0.26, 95% CI = -0.40 ~ -0.12, P = 0.0002], decreased transfusion rate [odds ratio (OR) = 0.47, 95% CI = 0.24 ~ 0.92, P = 0.03], shorter tract establishment time (MD = -1.30, 95% CI = -1.87 ~ -0.72, P < 0.0001) and shorter operation time (MD = -5.23, 95% CI = -10.19 ~ -0.27, P = 0.04). CONCLUSIONS: Overall, ultrasound-guided balloon dilatation offered several advantages in PCNL procedures. It facilitated faster access establishment, as evidenced by shorter access creation time. Additionally, it reduced the risk of kidney injury by minimizing postoperative haemoglobin drop and decreasing the need for transfusions. Moreover, it enhanced the efficiency of surgery by reducing the operation time. However, it is important to note that the quality of some included studies was subpar, as they did not adequately control for confounding factors that may affect the outcomes. Therefore, further research is necessary to validate and strengthen these findings.
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Cálculos Renales , Nefrolitotomía Percutánea , Nefrostomía Percutánea , Humanos , Nefrolitotomía Percutánea/métodos , Dilatación , Riñón , Cálculos Renales/cirugía , Ultrasonografía Intervencional , Hemoglobinas , Nefrostomía Percutánea/métodos , Resultado del TratamientoRESUMEN
PURPOSE: To study the analgesic effects and side effects of a transdermal lappaconitine (TLA) patch, ibuprofen suspension (IS), and TLA combined with IS (TLACIS) after adenoidectomy and tonsillectomy. DESIGN: Prospective, randomized clinical trial. METHODS: The patients were randomized into three groups defined by different analgesic drug regimens: the TLA group, the IS group, and the TLACIS group. Pain scores at 2, 12, and 24 hours after surgery and adverse-event reports within the first postoperative week were collected. RESULTS: Ultimately, this study included 102 cases in the TLA group, 101 cases in the IS group, and 101 cases in the TLACIS group. At 2 hours after surgery, the pain scores of the TLA and the TLACIS groups were both significantly lower than that of the IS group (all P < .05). At 12 and 24 hours after surgery, the pain score of the TLACIS group was significantly lower than those of the TLA and IS groups (all P < .05); furthermore, the pain score of the IS group was significantly lower than that of the TLA group (P < .05). Within 1 week after the operation, there was no significant difference in the incidence of adverse events. CONCLUSIONS: The addition of a TLA patch can speed the onset of analgesia. In terms of analgesic effects, IS alone is more advantageous than TLA alone, while the combination of TLA and IS has the best analgesic effect. No significant differences were found in the incidence of adverse events among the three regimens.
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BACKGROUND: Gastric cancer is the fourth leading cause of cancer-related deaths worldwide. And patient outcomes are poor due to tumor relapse and metastasis. To develop new therapeutic strategies, it is of great importance to explore the mechanism underlying the progression of gastric cancer. METHODS: Primary gastric cancer samples with lymph node metastases (LNM) and without LNM were subjected to mRNA microarray assay. The differentially expressed genes were confirmed by RT-qPCR. HRCT1 protein expression was further detected using an immunohistochemistry (IHC) assay. In vitro and in vivo assays were performed to investigate the role of HRCT1 in tumor invasion, metastasis, and proliferation. The expressions of the downstream target genes of HRCT1 were detected by microarray, RT-qPCR and Western blot assays. Dual-luciferase reporter and Western blot assays were carried out to identify miRNAs target to HRCT1. RESULTS: HRCT1 was upregulated in gastric cancer, and high expression of HRCT1 was associated with poor overall survival (OS) and disease-free survival (DFS). Moreover, HRCT1protein expression was an independent predictor for poor OS and DFS. HRCT1 could promote gastric cancer cells' migration, invasion, and proliferation in vitro as well as tumor metastasis and growth in vivo. Notably, our data showed that HRCT1 promoted gastric cancer progression by activating the ERBB2-MAPK signaling pathway. At least partially, the expression of HRCT1 could be negatively regulated by miR-124-3p. CONCLUSIONS: The upregulated expression of HRCT1 predicts poor survival for patients with gastric cancer. HRCT1 promotes tumor progression by activating the ERBB2-MAPK pathway. HRCT1, negatively regulated by miR-124-3p, may be a potential therapeutic target for patients with gastric cancer.
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MicroARNs , Neoplasias Gástricas , Humanos , Neoplasias Gástricas/patología , Proliferación Celular/genética , Línea Celular Tumoral , MicroARNs/genética , Transducción de Señal , Metástasis Linfática , Regulación Neoplásica de la Expresión Génica , Movimiento Celular/genética , Receptor ErbB-2/metabolismoRESUMEN
We report a rare case of esophageal highly differentiated liposarcoma that was safely removed by endoscopic submucosal dissection.
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Resección Endoscópica de la Mucosa , Neoplasias Esofágicas , Liposarcoma , Humanos , Neoplasias Esofágicas/diagnóstico por imagen , Neoplasias Esofágicas/cirugía , Liposarcoma/diagnóstico por imagen , Liposarcoma/cirugíaRESUMEN
BACKGROUND: Cystinuria and xanthinuria are both rare genetic diseases involving urinary calculi. However, cases combining these two disorders have not yet been reported. CASE PRESENTATION: In this study, we report a case of cystinuria with xanthine stones and hyperuricemia. The 23-year-old male patient was diagnosed with kidney and ureteral stones, solitary functioning kidney and hyperuricemia after admission to the hospital. The stones were removed by surgery and found to be composed of xanthine. CONCLUSION: Genetic testing by next-generation sequencing technology showed that the patient carried the homozygous nonsense mutation c.1113 C> A (p.Tyr371*) in the SLC3A1 gene, which was judged to be a functionally pathogenic variant. Sanger sequencing revealed that the patient's parents carried this heterozygous mutation, which is a pathogenic variant that can cause cystinuria. The 24-h urine metabolism analysis showed that the cystine content was 644 mg (<320 mg/24 h), indicating that the patient had cystinuria, consistent with the genetic test results. This case shows that cystinuria and xanthine stones can occur simultaneously, and provides evidence of a possible connection between the two conditions. Furthermore, our findings demonstrate the potential value of genetic testing using next-generation sequencing to effectively assist in the clinical diagnosis and treatment of patients with urinary calculi.
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Sistemas de Transporte de Aminoácidos , Cistinuria , Humanos , Masculino , Adulto Joven , Cistinuria/genética , Sistemas de Transporte de Aminoácidos/genética , Xantina , Cálculos Renales , Hiperuricemia , Codón sin Sentido , Pruebas Genéticas , Linaje , FemeninoRESUMEN
5-Methylcytosine (m5C) is a RNA modification that exists in tRNAs and rRNAs and was recently found in mRNAs. Although it has been suggested to regulate diverse biological functions, whether m5C RNA modification influences adult stem cell development remains undetermined. In this study, we show that Ypsilon schachtel (YPS), a homolog of human Y box binding protein 1 (YBX1), promotes germ line stem cell (GSC) maintenance, proliferation, and differentiation in the Drosophila ovary by preferentially binding to m5C-containing RNAs. YPS is genetically demonstrated to function intrinsically for GSC maintenance, proliferation, and progeny differentiation in the Drosophila ovary, and human YBX1 can functionally replace YPS to support normal GSC development. Highly conserved cold-shock domains (CSDs) of YPS and YBX1 preferentially bind to m5C RNA in vitro. Moreover, YPS also preferentially binds to m5C-containing RNAs, including mRNAs, in germ cells. The crystal structure of the YBX1 CSD-RNA complex reveals that both hydrophobic stacking and hydrogen bonds are critical for m5C binding. Overexpression of RNA-binding-defective YPS and YBX1 proteins disrupts GSC development. Taken together, our findings show that m5C RNA modification plays an important role in adult stem cell development.
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5-Metilcitosina/metabolismo , Proteínas de Unión al ADN/metabolismo , Proteínas de Drosophila/metabolismo , Óvulo/crecimiento & desarrollo , ARN/metabolismo , Animales , Proliferación Celular , Proteínas de Unión al ADN/genética , Proteínas de Drosophila/genética , Drosophila melanogaster/genética , Drosophila melanogaster/crecimiento & desarrollo , Drosophila melanogaster/metabolismo , Femenino , Humanos , Ovario/metabolismo , Óvulo/metabolismo , ARN/genética , Células Madre/citología , Células Madre/metabolismo , Proteína 1 de Unión a la Caja Y/genética , Proteína 1 de Unión a la Caja Y/metabolismoRESUMEN
Horizontal gene transfer (HGT) is a major driving force for bacterial evolution. To avoid the deleterious effects due to the unregulated expression of newly acquired foreign genes, bacteria have evolved specific proteins named xenogeneic silencers to recognize foreign DNA sequences and suppress their transcription. As there is considerable diversity in genomic base compositions among bacteria, how xenogeneic silencers distinguish self- from nonself DNA in different bacteria remains poorly understood. This review summarizes the progress in studying the DNA binding preferences and the underlying molecular mechanisms of known xenogeneic silencer families, represented by H-NS of Escherichia coli, Lsr2 of Mycobacterium, MvaT of Pseudomonas, and Rok of Bacillus. Comparative analyses of the published data indicate that the differences in DNA recognition mechanisms enable these xenogeneic silencers to have clear characteristics in DNA sequence preferences, which are further correlated with different host genomic features. These correlations provide insights into the mechanisms of how these xenogeneic silencers selectively target foreign DNA in different genomic backgrounds. Furthermore, it is revealed that the genomic AT contents of bacterial species with the same xenogeneic silencer family proteins are distributed in a limited range and are generally lower than those species without any known xenogeneic silencers in the same phylum/class/genus, indicating that xenogeneic silencers have multifaceted roles on bacterial genome evolution. In addition to regulating horizontal gene transfer, xenogeneic silencers also act as a selective force against the GC to AT mutational bias found in bacterial genomes and help the host genomic AT contents maintained at relatively low levels.
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Proteínas Bacterianas , Proteínas de Unión al ADN , Proteínas Bacterianas/genética , ADN , ADN Bacteriano , Proteínas de Unión al ADN/genética , Silenciador del Gen , Transferencia de Gen Horizontal , Genoma Bacteriano , HumanosRESUMEN
The lack of an effective printable ink preparation method and the usual mechanically weak performance obstruct the functional 3D printing hydrogel exploitation and application. Herein, we propose a gentle pre-cross-linking strategy to enable a loosely cross-linked cellulose network for simultaneously achieving favorable printability and a strong hydrogel network via mediating the cellulose self-assembly. A small amount of epichlorohydrin is applied to (i) slightly pre-cross-link the cellulose chains for forming the percolating network to regulate the rheological properties and (ii) form the loosely cross-linked points to mediate the cellulose chains' self-assembly for achieving superior mechanical properties. The fabrication of the complex 3D structures verifies the design flexibility. The printed cellulose hydrogels exhibit a biomimetic nanofibrous topology, remarkable tensile and compressive strength (5.22 and 11.80 MPa), as well as toughness (1.81 and 2.16 MJ/m3). As a demonstration, a bilayer scaffold (mimicking the osteochondral structure) consisting of a top pristine cellulose and a bottom cellulose/bioactive glass hydrogel is printed and exhibits superior osteochondral defect repair performance, showing a potential in tissue engineering. We anticipate that our loose pre-cross-linking 3D printing ink preparation concept can inspire the development of other polymeric inks and strong 3D printing functional hydrogels, eventually spreading the applications in diverse fields.
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Biomimética , Celulosa , Celulosa/química , Hidrogeles/química , Impresión Tridimensional , Ingeniería de Tejidos/métodos , Andamios del Tejido/químicaRESUMEN
GapR is a nucleoid-associated protein that is an essential regulator of chromosome replication in the cell cycle model Caulobacter crescentus. Here, we demonstrate that free GapR is a homotetramer, but not a dimer as previously reported (Guo et al., Cell 175: 583-597, 2018). We have determined the crystal structure of GapR in complex with a 10-bp A-tract DNA, which has an open tetrameric conformation, different from the closed clamp conformation in the previously reported crystal structure of GapR/DNA complex. The free GapR adopts multiple conformations in dynamic exchange equilibrium, with the major conformation resembling the closed tetrameric conformation, while the open tetrameric conformation is a representative of minor conformers. As it is impossible for the circular genomic DNA to get into the central DNA binding tunnel of the major conformation, we propose that GapR initially binds DNA through the open conformation, and then undergoes structural rearrangement to form the closed conformation which fully encircles the DNA. GapR prefers to bind DNA with 10-bp consecutive A/T base pairs nonselectively (Kd â¼12 nM), while it can also bind GC-rich DNA sequence with a reasonable affinity of about 120 nM. Besides, our results suggest that GapR binding results in widening the minor groove of AT-rich DNA, instead of overtwisting DNA.
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Proteínas Bacterianas/genética , ADN/genética , Conformación Molecular , Secuencia de Aminoácidos/genética , Proteínas Bacterianas/ultraestructura , Caulobacter crescentus/genética , Caulobacter crescentus/ultraestructura , División Celular/genética , ADN/ultraestructura , Regulación Bacteriana de la Expresión Génica/genética , Proteínas de la Membrana/genética , Proteínas de la Membrana/ultraestructura , Conformación de Ácido Nucleico , Multimerización de Proteína/genéticaRESUMEN
BACKGROUND: The inhibitory effect of Tetrandrine (Tet) on rheumatoid arthritis (RA) is well established. However, its exact molecular mechanism remains unknown. METHODS: RT-qPCR coupled with western blotting was employed to analyze the expression of NEAT1, miR-17-5p, and STAT3 in RA tissues and/or RA-fibroblast-like synoviocytes (RA-FLS) treated with 3 µmol/L of Tet for 48 h. Cell Counting Kit-8 assay and flow cytometry were performed to assess RA-FLS proliferation and apoptosis. Luciferase reporter assays were used to validate the interactions between miR-17-5p and STAT3 or NEAT1. RESULTS: The expression of NEAT1 decreased in a time-dependent manner upon Tet treatment. Tet significantly inhibited RA-FLS proliferation and triggered apoptosis by downregulating NEAT1 expression. Additionally, NEAT1 directly targeted miR-17-5p to upregulate STAT3 expression. Tet-induced low NEAT1 expression impaired RA-FLS growth by targeting miR-17-5p and inhibiting STAT3. CONCLUSION: Tet exerts its inhibitory role in RA progression by regulating the NEAT1/miR-17-5p/STAT3 pathway.
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Artritis Reumatoide , MicroARNs , ARN Largo no Codificante , Humanos , ARN Largo no Codificante/genética , Artritis Reumatoide/tratamiento farmacológico , Artritis Reumatoide/genética , MicroARNs/genética , Factor de Transcripción STAT3/genéticaRESUMEN
Methylation of DNA at CpG sites is a major mark for epigenetic regulation, but how transcription factors are influenced by CpG methylation is not well understood. Here, we report the molecular mechanisms of how the TCF (T-cell factor) and GEF (glucose transporter 4 enhancer factor) families of proteins selectively target unmethylated DNA sequences with a C-clamp type zinc finger domain. The structure of the C-clamp domain from human GEF family protein HDBP1 (C-clampHDBP1) in complex with DNA was determined using NMR spectroscopy, which adopts a unique zinc finger fold and selectively binds RCCGG (R = A/G) DNA sequences with an "Arg···Trp-Lys-Lys" DNA recognition motif inserted in the major groove. The CpG base pairs are central to the binding due to multiple hydrogen bonds formed with the backbone carbonyl groups of Trp378 and Lys379, as well as the side chain ε-amino groups of Lys379 and Lys380 from C-clampHDBP1. Consequently, methylation of the CpG dinucleotide almost abolishes the binding. Homology modeling reveals that the C-clamp domain from human TCF1E (C-clampTCF1E) binds DNA through essentially the same mechanism, with a similar "Arg···Arg-Lys-Lys" DNA recognition motif. The substitution of tryptophan by arginine makes C-clampHDBP1 prefer RCCGC DNA sequences. The two signature DNA recognition motifs are invariant in the GEF and TCF families of proteins, respectively, from fly to human. The recognition of the CpG dinucleotide through two consecutive backbone carbonyl groups is the same as that of the CXXC type unmethylated CpG DNA binding domains, suggesting a common mechanism shared by unmethylated CpG binding proteins.
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Cisteína/química , ADN/química , Islas de CpG , HumanosRESUMEN
OBJECTIVE: Dorsal root ganglion stimulation (DRGS) is an effective therapy for chronic pain, though its mechanisms of action are unknown. Currently, we do not understand how clinically controllable parameters (e.g., electrode position, stimulus pulse width) affect the direct neural response to DRGS. Therefore, the goal of this study was to utilize a computational modeling approach to characterize how varying clinically controllable parameters changed neural activation profiles during DRGS. MATERIALS AND METHODS: We coupled a finite element model of a human L5 DRG to multicompartment models of primary sensory neurons (i.e., Aα-, Aß-, Aδ-, and C-neurons). We calculated the stimulation amplitudes necessary to elicit one or more action potentials in each neuron, and examined how neural activation profiles were affected by varying clinically controllable parameters. RESULTS: In general, DRGS predominantly activated large myelinated Aα- and Aß-neurons. Shifting the electrode more than 2 mm away from the ganglion abolished most DRGS-induced neural activation. Increasing the stimulus pulse width to 500 µs or greater increased the number of activated Aδ-neurons, while shorter pulse widths typically only activated Aα- and Aß-neurons. Placing a cathode near a nerve root, or an anode near the ganglion body, maximized Aß-mechanoreceptor activation. Guarded active contact configurations did not activate more Aß-mechanoreceptors than conventional bipolar configurations. CONCLUSIONS: Our results suggest that DRGS applied with stimulation parameters within typical clinical ranges predominantly activates Aß-mechanoreceptors. In general, varying clinically controllable parameters affects the number of Aß-mechanoreceptors activated, although longer pulse widths can increase Aδ-neuron activation. Our data support several Neuromodulation Appropriateness Consensus Committee guidelines on the clinical implementation of DRGS.
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Dolor Crónico , Ganglios Espinales , Potenciales de Acción , Humanos , NeuronasRESUMEN
The nucleoid-associated protein GapR found in Caulobacter crescentus is crucial for DNA replication, transcription, and cell division. Associated with overtwisted DNA in front of replication forks and the 3' end of highly-expressed genes, GapR can stimulate gyrase and topo IV to relax (+) supercoils, thus facilitating the movement of the replication and transcription machines. GapR forms a dimer-of-dimers structure in solution that can exist in either an open or a closed conformation. It initially binds DNA through the open conformation and then undergoes structural rearrangement to form a closed tetramer, with DNA wrapped in the central channel. Here, we show that the DNA binding domain of GapR (residues 1-72, GapRΔC17) exists as a dimer in solution and adopts the same fold as the two dimer units in the full-length tetrameric protein. It binds DNA at the minor groove and reads the spatial distribution of DNA phosphate groups through a lysine/arginine network, with a preference towards AT-rich overtwisted DNA. These findings indicate that the dimer unit of GapR has an intrinsic DNA binding preference. Thus, at the initial binding step, the open tetramer of GapR with two relatively independent dimer units can be more efficiently recruited to overtwisted regions.
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Caulobacter crescentus/metabolismo , ADN Bacteriano/metabolismo , Transactivadores/química , Transactivadores/metabolismo , Secuencias de Aminoácidos , Proteínas Bacterianas/química , Proteínas Bacterianas/genética , Proteínas Bacterianas/metabolismo , Sitios de Unión , Caulobacter crescentus/química , Caulobacter crescentus/genética , Cristalografía por Rayos X , ADN Bacteriano/química , Regulación Bacteriana de la Expresión Génica , Modelos Moleculares , Mutación , Unión Proteica , Conformación Proteica , Multimerización de Proteína , Transactivadores/genéticaRESUMEN
Autophagy is typically a prosurvival cellular process that promotes the turnover of long-lived proteins and damaged organelles, but it can also induce cell death. We have previously reported that the small molecule Z36 induces autophagy along with autophagic cell death in HeLa cells. In this study, we analyzed differential gene expression in Z36-treated HeLa cells and found that Z36-induced endoplasmic reticulum-specific autophagy (ER-phagy) results in ER stress and the unfolded protein response (UPR). This result is in contrast to the common notion that autophagy is generally activated in response to ER stress and the UPR. We demonstrate that Z36 up-regulates the expression levels of FAM134B, LC3, and Atg9, which together mediate excessive ER-phagy, characterized by forming increased numbers of autophagosomes with larger sizes. We noted that the excessive ER-phagy accelerates ER degradation and impairs ER homeostasis and thereby triggers ER stress and the UPR as well as ER-phagy-dependent cell death. Interestingly, overexpression of FAM134B alone in HeLa cells is sufficient to impair ER homeostasis and cause ER stress and cell death. These findings suggest a mechanism involving FAM134B activity for ER-phagy to promote cell death.
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Autofagia , Estrés del Retículo Endoplásmico , Péptidos y Proteínas de Señalización Intracelular/metabolismo , Proteínas de la Membrana/metabolismo , Respuesta de Proteína Desplegada , Adenina/análogos & derivados , Adenina/farmacología , Apoptosis/efectos de los fármacos , Autofagosomas/metabolismo , Autofagia/efectos de los fármacos , Proteínas Relacionadas con la Autofagia/genética , Proteínas Relacionadas con la Autofagia/metabolismo , Retículo Endoplásmico/metabolismo , Estrés del Retículo Endoplásmico/efectos de los fármacos , Endorribonucleasas/metabolismo , Células HeLa , Humanos , Indoles/farmacología , Péptidos y Proteínas de Señalización Intracelular/antagonistas & inhibidores , Péptidos y Proteínas de Señalización Intracelular/genética , Proteínas de la Membrana/antagonistas & inhibidores , Proteínas de la Membrana/genética , Proteínas Asociadas a Microtúbulos/genética , Proteínas Asociadas a Microtúbulos/metabolismo , Proteínas Serina-Treonina Quinasas/metabolismo , Interferencia de ARN , ARN Interferente Pequeño/metabolismo , Respuesta de Proteína Desplegada/efectos de los fármacos , Regulación hacia Arriba/efectos de los fármacos , Proteínas de Transporte Vesicular/genética , Proteínas de Transporte Vesicular/metabolismoRESUMEN
BACKGROUND: Accumulating evidence demonstrated that long noncoding RNAs (lncRNAs) played important regulatory roles in many cancer types. However, the role of lncRNAs in gastric cancer (GC) progression remains unclear. METHODS: RT-qPCR assay was performed to detect the expression of HNF1A-AS1 in gastric cancer tissues and the non-tumourous gastric mucosa. Overexpression and RNA interference approaches were used to investigate the effects of HNF1A-AS1 on GC cells. Insight into competitive endogenous RNA (ceRNA) mechanisms was gained via bioinformatics analysis, luciferase assays and an RNA-binding protein immunoprecipitation (RIP) assay, RNA-FISH co-localisation analysis combined with microRNA (miRNA)-pulldown assay. RESULTS: This study displayed that revealed expression of HNF1A-AS1 was associated with positive lymph node metastasis in GC. Moreover, HNF1A-AS1 significantly promoted gastric cancer invasion, metastasis, angiogenesis and lymphangiogenesis in vitro and in vivo. In addition, HNF1A-AS1 was demonstrated to function as a ceRNA for miR-30b-3p. HNF1A-AS1 abolished the function of the miRNA-30b-3p and resulted in the derepression of its target, PIK3CD, which is a core oncogene involved in the progression of GC. CONCLUSION: This study demonstrated that HNF1A-AS1 worked as a ceRNA and promoted PI3K/AKT signalling pathway-mediated GC metastasis by sponging miR-30b-3p, offering novel insights of the metastasis mechanism in GC.