Your browser doesn't support javascript.
loading
Mostrar: 20 | 50 | 100
Resultados 1 - 20 de 33
Filtrar
1.
Plant Cell Environ ; 47(6): 2310-2321, 2024 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-38494960

RESUMO

Grain protein content (GPC) is a crucial quality trait in bread wheat, which is influenced by the key transcription factor TaNAM. However, the regulatory mechanisms of TaNAM have remained largely elusive. In this study, a new role of TaNAM was unveiled in regulating nitrogen remobilisation which impacts GPC. The TaNAM knockout mutants generated by clustered regularly interspaced short palindromic repeats/Cas9 exhibited significantly delayed senescence and lower GPC, while overexpression of TaNAM-6A resulted in premature senility and much higher GPC. Further analysis revealed that TaNAM directly activates the genes TaNRT1.1 and TaNPF5.5s, which are involved in nitrogen remobilisation. This activity aids in the transfer of nitrogen from leaves to grains for protein synthesis. In addition, an elite allele of TaNAM-6A, associated with high GPC, was identified as a candidate gene for breeding high-quality wheat. Overall, our work not only elucidates the potential mechanism of TaNAM-6A affecting bread wheat GPC, but also highlights the significance of nitrogen remobilisation from senescent leaves to grains for protein accumulation. Moreover, our research provides a new target and approach for improving the quality traits of wheat, particularly the GPC.


Assuntos
Nitrogênio , Triticum , Triticum/genética , Triticum/metabolismo , Nitrogênio/metabolismo , Proteínas de Grãos/metabolismo , Proteínas de Plantas/metabolismo , Proteínas de Plantas/genética , Regulação da Expressão Gênica de Plantas , Fatores de Transcrição/metabolismo , Fatores de Transcrição/genética , Grão Comestível/metabolismo , Grão Comestível/genética , Folhas de Planta/metabolismo , Folhas de Planta/genética
2.
BMC Cancer ; 24(1): 624, 2024 May 22.
Artigo em Inglês | MEDLINE | ID: mdl-38778317

RESUMO

BACKGROUND: Hepatocellular carcinoma (HCC) has a high mortality rate, and the mechanisms underlying tumor development and progression remain unclear. However, inactivated tumor suppressor genes might play key roles. DNA methylation is a critical regulatory mechanism for inactivating tumor suppressor genes in HCC. Therefore, this study investigated methylation-related tumor suppressors in HCC to identify potential biomarkers and therapeutic targets. METHODS: We assessed genome-wide DNA methylation in HCC using whole genome bisulfite sequencing (WGBS) and RNA sequencing, respectively, and identified the differential expression of methylation-related genes, and finally screened phosphodiesterase 7B (PDE7B) for the study. The correlation between PDE7B expression and clinical features was then assessed. We then analyzed the changes of PDE7B expression in HCC cells before and after DNA methyltransferase inhibitor treatment by MassArray nucleic acid mass spectrometry. Furthermore, HCC cell lines overexpressing PDE7B were constructed to investigate its effect on HCC cell function. Finally, GO and KEGG were applied for the enrichment analysis of PDE7B-related pathways, and their effects on the expression of pathway proteins and EMT-related factors in HCC cells were preliminarily explored. RESULTS: HCC exhibited a genome-wide hypomethylation pattern. We screened 713 hypomethylated and 362 hypermethylated mCG regions in HCC and adjacent normal tissues. GO analysis showed that the main molecular functions of hypermethylation and hypomethylation were "DNA-binding transcriptional activator activity" and "structural component of ribosomes", respectively, whereas KEGG analysis showed that they were enriched in "bile secretion" and "Ras-associated protein-1 (Rap1) signaling pathway", respectively. PDE7B expression was significantly down-regulated in HCC tissues, and this low expression was negatively correlated with recurrence and prognosis of HCC. In addition, DNA methylation regulates PDE7B expression in HCC. On the contrary, overexpression of PDE7B inhibited tumor proliferation and metastasis in vitro. In addition, PDE7B-related genes were mainly enriched in the PI3K/ATK signaling pathway, and PDE7B overexpression inhibited the progression of PI3K/ATK signaling pathway-related proteins and EMT. CONCLUSION: PDE7B expression in HCC may be regulated by promoter methylation. PDE7B can regulate the EMT process in HCC cells through the PI3K/AKT pathway, which in turn affects HCC metastasis and invasion.


Assuntos
Carcinoma Hepatocelular , Nucleotídeo Cíclico Fosfodiesterase do Tipo 7 , Metilação de DNA , Regulação Neoplásica da Expressão Gênica , Neoplasias Hepáticas , Fosfatidilinositol 3-Quinases , Proteínas Proto-Oncogênicas c-akt , Transdução de Sinais , Humanos , Neoplasias Hepáticas/genética , Neoplasias Hepáticas/patologia , Neoplasias Hepáticas/metabolismo , Carcinoma Hepatocelular/genética , Carcinoma Hepatocelular/patologia , Carcinoma Hepatocelular/metabolismo , Nucleotídeo Cíclico Fosfodiesterase do Tipo 7/genética , Nucleotídeo Cíclico Fosfodiesterase do Tipo 7/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , Transdução de Sinais/genética , Fosfatidilinositol 3-Quinases/metabolismo , Fosfatidilinositol 3-Quinases/genética , Linhagem Celular Tumoral , Invasividade Neoplásica/genética , Genes Supressores de Tumor , Masculino , Proliferação de Células/genética , Feminino , Metástase Neoplásica , Movimento Celular/genética
3.
J Exp Bot ; 73(19): 6600-6614, 2022 11 02.
Artigo em Inglês | MEDLINE | ID: mdl-35781562

RESUMO

Heat stress substantially reduces the yield potential of wheat (Triticum aestivum L.), one of the most widely cultivated staple crops, and greatly threatens global food security in the context of global warming. However, few studies have explored the heat stress tolerance (HST)-related genetic resources in wheat. Here, we identified and fine-mapped a wheat HST locus, TaHST2, which is indispensable for HST in both the vegetative and reproductive stages of the wheat life cycle. The studied pair of near isogenic lines (NILs) exhibited diverse morphologies under heat stress, based on which we mapped TaHST2 to a 485 kb interval on chromosome arm 4DS. Under heat stress, TaHST2 confers a superior conversion rate from soluble sugars to starch in wheat grains, resulting in faster grain filling and a higher yield potential. A further exploration of genetic resources indicated that TaHST2 underwent strong artificial selection during wheat domestication, suggesting it is an essential locus for basal HST in wheat. Our findings provide deeper insights into the genetic basis of wheat HST and might be useful for global efforts to breed heat-stress-tolerant cultivars.


Assuntos
Termotolerância , Triticum , Triticum/genética , Melhoramento Vegetal , Resposta ao Choque Térmico/genética , Termotolerância/genética , Grão Comestível/genética
4.
Int J Mol Sci ; 23(10)2022 May 10.
Artigo em Inglês | MEDLINE | ID: mdl-35628114

RESUMO

WRINKLED1 (WRI1), an APETALA2 (AP2) transcription factor (TF), critically regulates the processes related to fatty acid synthesis, storage oil accumulation, and seed development in plants. However, the WRI1 genes remain unknown in allohexaploid bread wheat (Triticum aestivum L.). In this study, based on the sequence of Arabidopsis AtWRI1, two TaWRI1Ls genes of bread wheat, TaWRI1L1 and TaWRI1L2, were cloned. TaWRI1L2 was closely related to monocotyledons and clustered in one subgroup with AtWRI1, while TaWRI1L1 was clustered in another subgroup with AtWRI3 and AtWRI4. Both were expressed highly in the developmental grain, subcellular localized in the nucleus, and showed transcriptional activation activity. TaWRI1L2, rather than TaWRI1L1, promoted oil body accumulation and significantly increased triglyceride (TAG) content in tobacco leaves. Overexpression of TaWRI1L2 compensated for the functional loss of AtWRI1 in an Arabidopsis mutant and restored the wild-type phenotypes of seed shape, generation, and fatty acid synthesis and accumulation. Knockout of TaWRI1L2 reduced grain size, 1000 grain weight, and grain fatty acid synthesis in bread wheat. Conclusively, TaWRI1L2, rather than TaWRI1L1, was the key transcriptional factor in the regulation of grain fatty acid synthesis in bread wheat. This study lays a foundation for gene regulation and genetic manipulation of fatty acid synthesis in wheat genetic breeding programs.


Assuntos
Proteínas de Arabidopsis , Arabidopsis , Arabidopsis/metabolismo , Proteínas de Arabidopsis/genética , Pão , Clonagem Molecular , Grão Comestível/genética , Ácidos Graxos , Regulação da Expressão Gênica de Plantas , Melhoramento Vegetal , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo , Triticum/metabolismo
5.
Respir Res ; 22(1): 8, 2021 Jan 06.
Artigo em Inglês | MEDLINE | ID: mdl-33407472

RESUMO

BACKGROUND: Pulmonary arterial smooth muscle cell (PASMC) proliferation plays a crucial role in hypoxia-induced pulmonary hypertension (HPH). Previous studies have found that resistin-like molecule ß (RELM-ß) is upregulated de novo in response to hypoxia in cultured human PASMCs (hPASMCs). RELM-ß has been reported to promote hPASMC proliferation and is involved in pulmonary vascular remodeling in patients with PAH. However, the expression pattern, effects, and mechanisms of action of RELM-ß in HPH remain unclear. METHODS: We assessed the expression pattern, mitogenetic effect, and mechanism of action of RELM-ß in a rat HPH model and in hPASMCs. RESULTS: Overexpression of RELM-ß caused hemodynamic changes in a rat model of HPH similar to those induced by chronic hypoxia, including increased mean right ventricular systolic pressure (mRVSP), right ventricular hypertrophy index (RVHI) and thickening of small pulmonary arterioles. Knockdown of RELM-ß partially blocked the increases in mRVSP, RVHI, and vascular remodeling induced by hypoxia. The phosphorylation levels of the PI3K, Akt, mTOR, PKC, and MAPK proteins were significantly up- or downregulated by RELM-ß gene overexpression or silencing, respectively. Recombinant RELM-ß protein increased the intracellular Ca2+ concentration in primary cultured hPASMCs and promoted hPASMC proliferation. The mitogenic effects of RELM-ß on hPASMCs and the phosphorylation of PI3K, Akt, mTOR, PKC, and MAPK were suppressed by a Ca2+ inhibitor. CONCLUSIONS: Our findings suggest that RELM-ß acts as a cytokine-like growth factor in the development of HPH and that the effects of RELM-ß are likely to be mediated by the Ca2+-dependent PI3K/Akt/mTOR and PKC/MAPK pathways.


Assuntos
Hormônios Ectópicos/biossíntese , Hipertensão Pulmonar/metabolismo , Sistema de Sinalização das MAP Quinases/fisiologia , Fosfatidilinositol 3-Quinases/biossíntese , Proteína Quinase C/biossíntese , Proteínas Proto-Oncogênicas c-akt/biossíntese , Animais , Cálcio/metabolismo , Células Cultivadas , Técnicas de Silenciamento de Genes/métodos , Substâncias de Crescimento/biossíntese , Substâncias de Crescimento/genética , Hormônios Ectópicos/antagonistas & inibidores , Hormônios Ectópicos/genética , Hipertensão Pulmonar/genética , Masculino , Ratos , Ratos Sprague-Dawley
6.
Yeast ; 37(7-8): 359-371, 2020 07.
Artigo em Inglês | MEDLINE | ID: mdl-32491201

RESUMO

In the budding yeast Saccharomyces cerevisiae, Svl3 and Pam1 proteins work as functional homologues. Loss of their function causes increased levels of chitin deposition in the cell wall and temperature sensitivity, suggesting their involvement in cell wall formation. We found that the N- and C-termini of these proteins have distinctive and critical functions. They contain an N-terminal part that has a probable 2-dehydropantoate 2-reductase domain. In Svl3, this part can be replaced with the yeast 2-dehydropantoate 2-reductase, Pan5, suggesting that Svl3 and its homologues may be able to mediate 2-dehydropantoate 2-reductase function. On the other hand, Svl3 is recruited to the bud tip and bud neck via multiple localization signals in the C-terminal part. One of such signals is the lysine-rich region located in the C-terminal end. The function and localization of Svl3 are significantly disrupted by the loss of this lysine-rich region; however, its localization is not completely abolished by the mutation because another localization signal enables appropriate transport. Svl3 and Pam1 orthologues are found in cells across fungal species. The Svl3 orthologues of Candida glabrata can complement the loss of Svl3 and Pam1 in S. cerevisiae. C. glabrata cells lacking the SVL3 and PAM1 orthologue genes exhibit phenotypes similar to those observed in svl3∆pam1∆ S. cerevisiae cells. Thus, Svl3 homologues may be generally required for the assembly of the cell wall in fungal cells.


Assuntos
Parede Celular/genética , Parede Celular/metabolismo , Proteínas Fúngicas/genética , Proteínas Fúngicas/metabolismo , Oxirredutases do Álcool , Candida glabrata , Quitina/metabolismo , Genes Fúngicos/genética , Mutação , Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/metabolismo , Proteínas de Saccharomyces cerevisiae/genética , Proteínas de Saccharomyces cerevisiae/metabolismo
7.
Mol Cell Proteomics ; 16(11): 1889-1905, 2017 11.
Artigo em Inglês | MEDLINE | ID: mdl-28821602

RESUMO

Potassium (K+) is the most abundant inorganic cation in plants, and molecular dissection of K+ deficiency has received considerable interest in order to minimize K+ fertilizer input and develop high quality K+-efficient crops. However, the molecular mechanism of plant responses to K+ deficiency is still poorly understood. In this study, 2-week-old bread wheat seedlings grown hydroponically in Hoagland solution were transferred to K+-free conditions for 8 d, and their root and leaf proteome profiles were assessed using the iTRAQ proteome method. Over 4000 unique proteins were identified, and 818 K+-responsive protein species showed significant differences in abundance. The differentially expressed protein species were associated with diverse functions and exhibited organ-specific differences. Most of the differentially expressed protein species related to hormone synthesis were involved in jasmonic acid (JA) synthesis and the upregulated abundance of JA synthesis-related enzymes could result in the increased JA concentrations. Abundance of allene oxide synthase (AOS), one key JA synthesis-related enzyme, was significantly increased in K+-deficient wheat seedlings, and its overexpression markedly increased concentrations of K+ and JA, altered the transcription levels of some genes encoding K+-responsive protein species, as well as enhanced the tolerance of rice plants to low K+ or K+ deficiency. Moreover, rice AOS mutant (osaos) exhibited more sensitivity to low K+ or K+ deficiency. Our findings could highlight the importance of JA in K+ deficiency, and imply a network of molecular processes underlying plant responses to K+ deficiency.


Assuntos
Ciclopentanos/metabolismo , Oryza/genética , Oxilipinas/metabolismo , Proteínas de Plantas/metabolismo , Plantas Geneticamente Modificadas/crescimento & desenvolvimento , Potássio/metabolismo , Proteômica/métodos , Triticum/genética , Produtos Agrícolas/genética , Produtos Agrícolas/crescimento & desenvolvimento , Produtos Agrícolas/metabolismo , Perfilação da Expressão Gênica/métodos , Regulação da Expressão Gênica de Plantas , Redes Reguladoras de Genes , Especificidade de Órgãos , Oryza/crescimento & desenvolvimento , Oryza/metabolismo , Folhas de Planta/metabolismo , Proteínas de Plantas/genética , Raízes de Plantas/metabolismo , Plantas Geneticamente Modificadas/metabolismo , Plântula/crescimento & desenvolvimento , Triticum/crescimento & desenvolvimento , Triticum/metabolismo
8.
Blood ; 127(21): 2575-86, 2016 05 26.
Artigo em Inglês | MEDLINE | ID: mdl-26941401

RESUMO

Herein we demonstrate that oncolytic herpes simplex virus-1 (HSV-1) potently activates human peripheral blood mononuclear cells (PBMCs) to lyse leukemic cell lines and primary acute myeloid leukemia samples, but not healthy allogeneic lymphocytes. Intriguingly, we found that UV light-inactivated HSV-1 (UV-HSV-1) is equally effective in promoting PBMC cytolysis of leukemic cells and is 1000- to 10 000-fold more potent at stimulating innate antileukemic responses than UV-inactivated cytomegalovirus, vesicular stomatitis virus, reovirus, or adenovirus. Mechanistically, UV-HSV-1 stimulates PBMC cytolysis of leukemic cells, partly via Toll-like receptor-2/protein kinase C/nuclear factor-κB signaling, and potently stimulates expression of CD69, degranulation, migration, and cytokine production in natural killer (NK) cells, suggesting that surface components of UV-HSV-1 directly activate NK cells. Importantly, UV-HSV-1 synergizes with interleukin-15 (IL-15) and IL-2 in inducing activation and cytolytic activity of NK cells. Additionally, UV-HSV-1 stimulates glycolysis and fatty acid oxidation-dependent oxygen consumption in NK cells, but only glycolysis is required for their enhanced antileukemic activity. Last, we demonstrate that T cell-depleted human PBMCs exposed to UV-HSV-1 provide a survival benefit in a murine xenograft model of human acute myeloid leukemia (AML). Taken together, our results support the preclinical development of UV-HSV-1 as an adjuvant, alone or in combination with IL-15, for allogeneic donor mononuclear cell infusions to treat AML.


Assuntos
Herpesvirus Humano 1/imunologia , Imunidade Celular , Células Matadoras Naturais/imunologia , Leucemia/imunologia , Raios Ultravioleta , Inativação de Vírus/efeitos da radiação , Degranulação Celular/imunologia , Movimento Celular/imunologia , Feminino , Humanos , Interleucina-15/imunologia , Interleucina-2/imunologia , Células Jurkat , Masculino , NF-kappa B/imunologia , Proteína Quinase C/imunologia , Transdução de Sinais/imunologia , Receptor 2 Toll-Like/imunologia
9.
Molecules ; 23(4)2018 Mar 24.
Artigo em Inglês | MEDLINE | ID: mdl-29587341

RESUMO

Wheat aphids are major pests during the seed filling stage of wheat. Plant lectins are toxic to sap-sucking pests such as wheat aphids. In this study, Pinellia pedatisecta agglutinin (ppa), a gene encoding mannose binding lectin, was cloned, and it shared 92.69% nucleotide similarity and 94% amino acid similarity with Pinellia ternata agglutinin (pta). The ppa gene, driven by the constitutive and phloem-specific ribulose bisphosphate carboxylase small subunit gene (rbcs) promoter in pBAC-rbcs-ppa expression vector, was transferred into the wheat cultivar Baofeng104 (BF104) by particle bombardment transformation. Fifty-four T0 transgenic plants were generated. The inheritance and expression of the ppa gene were confirmed by PCR and RT-PCR analysis respectively, and seven homozygous transgenic lines were obtained. An aphid bioassay on detached leaf segments revealed that seven ppa transgenic wheat lines had lower aphid growth rates and higher inhibition rates than BF104. Furthermore, two-year aphid bioassays in isolated fields showed that aphid numbers per tiller of transgenic lines were significantly decreased, compared with wild type BF104. Therefore, ppa could be a strong biotechnological candidate to produce aphid-resistant wheat.


Assuntos
Afídeos/crescimento & desenvolvimento , Lectinas de Ligação a Manose/genética , Pinellia/metabolismo , Triticum/crescimento & desenvolvimento , Animais , Sequência de Bases , Clonagem Molecular , Lectinas de Ligação a Manose/metabolismo , Controle Biológico de Vetores , Pinellia/genética , Folhas de Planta/genética , Folhas de Planta/crescimento & desenvolvimento , Folhas de Planta/parasitologia , Lectinas de Plantas/genética , Lectinas de Plantas/metabolismo , Plantas Geneticamente Modificadas/crescimento & desenvolvimento , Plantas Geneticamente Modificadas/metabolismo , Regiões Promotoras Genéticas , Triticum/genética , Triticum/parasitologia
10.
Int J Mol Sci ; 17(10)2016 Sep 23.
Artigo em Inglês | MEDLINE | ID: mdl-27669224

RESUMO

The function of a wheat starch regulator 1 (TaRSR1) in regulating the synthesis of grain storage starch was determined using the barley stripe mosaic virus-virus induced gene-silencing (BSMV-VIGS) method in field experiments. Chlorotic stripes appeared on the wheat spikes infected with barley stripe mosaic virus-virus induced gene-silencing- wheat starch regulator 1 (BSMV-VIGS-TaRSR1) at 15 days after anthesis, at which time the transcription levels of the TaRSR1 gene significantly decreased. Quantitative real-time PCR was also used to measure the transcription levels of 26 starch synthesis-related enzyme genes in the grains of BSMV-VIGS-TaRSR1-silenced wheat plants at 20, 27, and 31 days after anthesis. The results showed that the transcription levels of some starch synthesis-related enzyme genes were markedly induced at different sampling time points: TaSSI, TaSSIV, TaBEIII, TaISA1, TaISA3, TaPHOL, and TaDPE1 genes were induced at each of the three sampling time points and TaAGPS1-b, TaAGPL1, TaAGPL2, TaSSIIb, TaSSIIc, TaSSIIIb, TaBEI, TaBEIIa, TaBEIIb, TaISA2, TaPHOH, and TaDPE2 genes were induced at one sampling time point. Moreover, both the grain starch contents, one thousand kernel weights, grain length and width of BSMV-VIGS-TaRSR1-infected wheat plants significantly increased. These results suggest that TaRSR1 acts as a negative regulator and plays an important role in starch synthesis in wheat grains by temporally regulating the expression of specific starch synthesis-related enzyme genes.


Assuntos
Proteínas de Plantas/metabolismo , Amido/biossíntese , Fatores de Transcrição/metabolismo , Triticum/metabolismo , Inativação Gênica , Vetores Genéticos/genética , Vetores Genéticos/metabolismo , Vírus do Mosaico/genética , Fenótipo , Proteínas de Plantas/antagonistas & inibidores , Proteínas de Plantas/genética , Plantas Geneticamente Modificadas/genética , Plantas Geneticamente Modificadas/metabolismo , Fatores de Transcrição/antagonistas & inibidores , Fatores de Transcrição/genética , Triticum/genética
11.
Talanta ; 273: 125950, 2024 Jun 01.
Artigo em Inglês | MEDLINE | ID: mdl-38521024

RESUMO

The residue of ampicillin (AMP) in food and ecological environment poses a potential harm to human health. Therefore, a reliable system for detecting AMP is in great demand. Herein, a label-free and sensitive electrochemical sensor utilizing NH2-Co-MOF as an electrocatalytic active material for methylene blue (MB) was developed for rapid and facile AMP detection by combining hybridization chain reaction (HCR), catalytic hairpin assembly (CHA) with CRISPR/Cas12a. The surface of glassy carbon electrode modified with NH2-Co-MOF was able to undergo HCR independent of the AMP, forming long dsDNA complexes to load MB, resulting in strong original electrochemical signal. The presence of AMP could trigger upstream CHA circuit to activate the CRISPR/Cas12a system, thereby achieving rapid non-specific cleavage of the trigger ssDNA of HCR on the electrode surface, hindering the occurrence of HCR and reducing the load of MB. Significant signal change triggered by the target was ultimately obtained, thus achieving sensitive detection of the AMP with a detection limit as low as 1.60 pM (S/N = 3). The proposed sensor exhibited good stability, selectivity, and stability, and achieved reliable detection of AMP in milk and livestock wastewater samples, demonstrating its promising application prospects in food safety and environmental monitoring.


Assuntos
Técnicas Biossensoriais , Ácidos Nucleicos , Humanos , Sistemas CRISPR-Cas , Hibridização de Ácido Nucleico , Ampicilina , Técnicas Biossensoriais/métodos
12.
Anal Chim Acta ; 1304: 342562, 2024 May 22.
Artigo em Inglês | MEDLINE | ID: mdl-38637031

RESUMO

A sensitive electrochemical platform was constructed with NH2-Cu-MOF as electrochemical probe to detect antibiotics using CRISPR/Cas12a system triggered by hybridization chain reaction (HCR). The sensing system consists of two HCR systems. HCR1 occurred on the electrode surface independent of the target, generating long dsDNA to connect signal probes and producing a strong electrochemical signal. HCR2 was triggered by target, and the resulting dsDNA products activated the CRISPR/Cas12a, thereby resulting in effective and rapid cleavage of the trigger of HCR1, hindering the occurrence of HCR1, and reducing the number of NH2-Cu-MOF on the electrode surface. Eventually, significant signal change depended on the target was obtained. On this basis and with the help of the programmability of DNA, kanamycin and ampicillin were sensitively detected with detection limits of 60 fM and 10 fM (S/N = 3), respectively. Furthermore, the sensing platform showed good detection performance in milk and livestock wastewater samples, demonstrating its great application prospects in the detection of antibiotics in food and environmental water samples.


Assuntos
Antibacterianos , Técnicas Biossensoriais , Técnicas Eletroquímicas/métodos , Sistemas CRISPR-Cas , Técnicas Biossensoriais/métodos , Hibridização de Ácido Nucleico
13.
Anal Chim Acta ; 1278: 341715, 2023 Oct 16.
Artigo em Inglês | MEDLINE | ID: mdl-37709458

RESUMO

Herein, powerful DNA strand displacement reaction and sensitive electrochemical analysis method were ingeniously integrated to develop a programmable biosensing platform. Using DNA as the detection model, a cascade amplification system based on catalytic hairpin assembly and entropy-driven catalytic was constructed, and the reaction rate and signal amplification effect were significantly improved. The product of the cascade amplification circuit could undergo strand displacement reaction with the signal probe on the electrode surface to obtain sensitive electrochemical signal changes and realize highly sensitive detection of the target. In addition, without redesigning the DNA sequences in the cascade amplification circuit, the by-product strand typically wasted in traditional entropy-driven catalytic reactions can be fully utilized to construct a single-signal output biosensing system and even a dual-signal output ratiometric biosensing platform, improving the detection repeatability and reliability of the system, and expanding the application of DNA strand displacement reaction in electrochemical biosensing. Furthermore, benefiting from the design flexibility of the DNA molecules, the constructed biosensing platform realized the sensitive detection of aptamer substrate (kanamycin as an example) and certain metal ion (mercury as an example) by simply recoding the corresponding recognition sequence, demonstrating the good versatility of the biosensing platform.


Assuntos
Técnicas Eletroquímicas , Entropia , Reprodutibilidade dos Testes , Catálise , Eletrodos
14.
Microorganisms ; 11(5)2023 Apr 26.
Artigo em Inglês | MEDLINE | ID: mdl-37317099

RESUMO

Herein, we describe a yeast cell-based assay system to analyze SNAP25-targeting botulinum neurotoxins (BoNTs). BoNTs are protein toxins, and, upon incorporation into neuronal cells, their light chains (BoNT-LCs) target specific synaptosomal N-ethylmaleimide-sensitive attachment protein receptor (SNARE) proteins, including synaptosomal-associated protein 25 (SNAP25). BoNT-LCs are metalloproteases, and each BoNT-LC recognizes and cleaves conserved domains in SNAREs termed the SNARE domain. In the budding yeast Saccharomyces cerevisiae, the SNAP25 ortholog Spo20 is required for production of the spore plasma membrane; thus, defects in Spo20 cause sporulation deficiencies. We found that chimeric SNAREs in which SNARE domains in Spo20 are replaced with those of SNAP25 are functional in yeast cells. The Spo20/SNAP25 chimeras, but not Spo20, are sensitive to digestion by BoNT-LCs. We demonstrate that spo20∆ yeasts harboring the chimeras exhibit sporulation defects when various SNAP25-targeting BoNT-LCs are expressed. Thus, the activities of BoNT-LCs can be assessed by colorimetric measurement of sporulation efficiencies. Although BoNTs are notorious toxins, they are also used as therapeutic and cosmetic agents. Our assay system will be useful for analyzing novel BoNTs and BoNT-like genes, as well as their manipulation.

15.
Sci Rep ; 13(1): 10185, 2023 06 22.
Artigo em Inglês | MEDLINE | ID: mdl-37349536

RESUMO

Hyperuricemia (HUA) endangers human health, and its prevalence has increased rapidly in recent decades. The current study investigated HUA's prevalence and influencing factors in Gongcheng, southern China. A cross-sectional investigation was conducted; 2128 participants aged 30-93 years were included from 2018 to 2019. Univariate and multivariate logistic regression models were used to screen HUA variables. A Bayesian network model was constructed using the PC algorithm to evaluate the association between influencing factors and HUA. The prevalence of HUA was 15.6% (23.2% in men, 10.7% in women). After screening the variables using a logistic regression analysis model, fatty liver disease (FLD), dyslipidemia, abdominal obesity, creatinine (CREA), somatotype, bone mass, drinking, and physical activity level at work were included in the Bayesian network model. The model results showed that dyslipidemia, somatotype, CREA, and drinking were directly related to HUA. Bone mass and FLD were indirectly associated with HUA by affecting the somatotype. The prevalence of HUA in Gongcheng was high in China. The prevalence of HUA was related to somatotype, drinking, bone mass, physical activity level at work, and other metabolic diseases. A good diet and moderate exercise are recommended to maintain a healthy somatotype and reduce the prevalence rate of HUA.


Assuntos
Hiperuricemia , Masculino , Pessoa de Meia-Idade , Humanos , Feminino , Idoso , Hiperuricemia/epidemiologia , Estudos Transversais , Prevalência , Teorema de Bayes , Fatores de Risco , China/epidemiologia
16.
Sci China Life Sci ; 66(7): 1647-1664, 2023 07.
Artigo em Inglês | MEDLINE | ID: mdl-36802319

RESUMO

Grain development is a crucial determinant of yield and quality in bread wheat (Triticum aestivum L.). However, the regulatory mechanisms underlying wheat grain development remain elusive. Here we report how TaMADS29 interacts with TaNF-YB1 to synergistically regulate early grain development in bread wheat. The tamads29 mutants generated by CRISPR/Cas9 exhibited severe grain filling deficiency, coupled with excessive accumulation of reactive oxygen species (ROS) and abnormal programmed cell death that occurred in early developing grains, while overexpression of TaMADS29 increased grain width and 1,000-kernel weight. Further analysis revealed that TaMADS29 interacted directly with TaNF-YB1; null mutation in TaNF-YB1 caused grain developmental deficiency similar to tamads29 mutants. The regulatory complex composed of TaMADS29 and TaNF-YB1 exercises its possible function that inhibits the excessive accumulation of ROS by regulating the genes involved in chloroplast development and photosynthesis in early developing wheat grains and prevents nucellar projection degradation and endosperm cell death, facilitating transportation of nutrients into the endosperm and wholly filling of developing grains. Collectively, our work not only discloses the molecular mechanism of MADS-box and NF-Y TFs in facilitating bread wheat grain development, but also indicates that caryopsis chloroplast might be a central regulator of grain development rather than merely a photosynthesis organelle. More importantly, our work offers an innovative way to breed high-yield wheat cultivars by controlling the ROS level in developing grains.


Assuntos
Pão , Triticum , Espécies Reativas de Oxigênio/metabolismo , Melhoramento Vegetal , Grão Comestível/metabolismo
17.
Mol Ther Oncolytics ; 28: 334-348, 2023 Mar 16.
Artigo em Inglês | MEDLINE | ID: mdl-36938544

RESUMO

VG2025 is a recombinant oncolytic herpes simplex virus type 1 (HSV-1) that uses transcriptional and translational dual regulation (TTDR) of critical viral genes to enhance virus safety and promote tumor-specific virus replication without reducing virulence. The TTDR platform is based on transcriptional control of the essential HSV-1 immediate-early protein ICP27 using a tumor-specific carcinoembryonic antigen (CEA) promoter, coupled with translational control of the neurovirulence factor ICP34.5 using multiple microRNA (miR)-binding sites. VG2025 further incorporates IL-12 and the IL-15/IL-15 receptor alpha subunit complex to enhance the antitumor and immune stimulatory properties of oncolytic HSVs. The TTDR strategy was verified in vitro and shown to be highly selective. Strong in vivo antitumor efficacy was observed following both intratumoral and intravenous administration. Clear abscopal and immune memory effects were also evident, indicating a robust antitumor immune response. Gene expression profiling of treated tumors revealed increased immune cell infiltration and activation of multiple immune-signaling pathways when compared with the backbone virus. Absence of neurotoxicity was verified in mice and in rhesus monkeys. Taken together, the enhanced tumor clearance, excellent safety profile, and positive correlation between CEA levels and viral replication efficiency may provide an opportunity for using biomarker-based precision medicine in oncolytic virotherapy.

18.
Vaccines (Basel) ; 11(12)2023 Dec 02.
Artigo em Inglês | MEDLINE | ID: mdl-38140209

RESUMO

The development of effective cancer vaccines remains a significant challenge due to immune tolerance and limited clinical benefits. Oncolytic herpes simplex virus type 1 (oHSV-1) has shown promise as a cancer therapy, but efficacy is often limited in advanced cancers. In this study, we constructed and characterized a novel oHSV-1 virus (VG22401) expressing the human epidermal growth factor receptor 2 (HER2), a transmembrane glycoprotein overexpressed in many carcinomas. VG22401 exhibited efficient replication and HER2 payload expression in both human and mouse colorectal cancer cells. Mice immunized with VG22401 showed significant binding of serum anti-HER2 antibodies to HER2-expressing tumor cells, inducing antibody-dependent cell-mediated cytotoxicity (ADCC) and complement-dependent cytotoxicity (CDC). Furthermore, mice primed with VG22401 and intratumorally boosted with the same virus showed enhanced antitumor efficacy in a bilateral syngeneic HER2(+) tumor model, compared to HER2-null backbone virus. This effect was accompanied by the induction of anti-HER2 T cell responses. Our findings suggest that peripheral priming with HER2-expressing oHSV-1 followed by an intratumoral boost with the same virus can significantly enhance antitumor immunity and efficacy, presenting a promising strategy for cancer immunotherapy.

19.
Commun Biol ; 5(1): 824, 2022 08 16.
Artigo em Inglês | MEDLINE | ID: mdl-35974093

RESUMO

In mammals, both professional phagocytes and nonprofessional phagocytes (NPPs) can perform phagocytosis. However, limited targets are phagocytosed by NPPs, and thus, the mechanism remains unclear. We find that spores of the yeast Saccharomyces cerevisiae are internalized efficiently by NPPs. Analyses of this phenomenon reveals that RNA fragments derived from cytosolic RNA species are attached to the spore wall, and these fragments serve as ligands to induce spore internalization. Furthermore, we show that a multiligand receptor, RAGE (receptor for advanced glycation end-products), mediates phagocytosis in NPPs. RAGE-mediated phagocytosis is not uniquely induced by spores but is an intrinsic mechanism by which NPPs internalize macromolecules containing RAGE ligands. In fact, artificial particles labeled with polynucleotides, HMGB1, or histone (but not bovine serum albumin) are internalized in NPPs. Our findings provide insight into the molecular basis of phagocytosis by NPPs, a process by which a variety of macromolecules are targeted for internalization.


Assuntos
Fagócitos , Receptores Imunológicos , Animais , Ligantes , Mamíferos , Fagocitose , RNA , Receptor para Produtos Finais de Glicação Avançada
20.
Microbiol Spectr ; 10(4): e0089722, 2022 08 31.
Artigo em Inglês | MEDLINE | ID: mdl-35924934

RESUMO

Cells of the budding yeast Saccharomyces cerevisiae form spores or stationary cells upon nutrient starvation. These quiescent cells are known to resume mitotic growth in response to nutrient signals, but the mechanism remains elusive. Here, we report that quiescent yeast cells are equipped with a negative regulatory mechanism which suppresses the commencement of mitotic growth. The regulatory process involves a glycolytic enzyme, triosephosphate isomerase (Tpi1), and its product, glyceraldehyde-3-phosphate (GAP). GAP serves as an inhibitory signaling molecule; indeed, the return to growth of spores or stationary cells is suppressed by the addition of GAP even in nutrient-rich growth media, though mitotic cells are not affected. Reciprocally, dormancy is abolished by heat treatment because of the heat sensitivity of Tpi1. For example, spores commence germination merely upon heat treatment, which indicates that the negative regulatory mechanism is actively required for spores to prevent premature germination. Stationary cells of Candida glabrata are also manipulated by heat and GAP, suggesting that the regulatory process is conserved in the pathogenic yeast. IMPORTANCE Our results suggest that, in quiescent cells, nutrient signals do not merely provoke a positive regulatory process to commence mitotic growth. Exit from the quiescent state in yeast cells is regulated by balancing between the positive and negative signaling pathways. Identifying the negative regulatory pathway would provide new insight into the regulation of the transition from the quiescent to the mitotic state. Clinically, quiescent cells are problematic because they are resistant to environmental stresses and antibiotics. Given that the quiescent state is modulated by manipulation of the negative regulatory mechanism, understanding this process is important not only for its biological interest but also as a potential target for antifungal treatment.


Assuntos
Saccharomyces cerevisiae , Triose-Fosfato Isomerase , Gliceraldeído , Gliceraldeído 3-Fosfato , Fosfatos , Triose-Fosfato Isomerase/metabolismo
SELEÇÃO DE REFERÊNCIAS
DETALHE DA PESQUISA