RESUMO
Starch properties can be modified by mutating genes responsible for the synthesis of amylose and amylopectin in the endosperm. However, little is known about the effects of such targeted modifications on the overall starch biosynthesis pathway and broader metabolism. Here we investigated the effects of mutating the OsSBEIIb gene encoding starch branching enzyme IIb, which is required for amylopectin synthesis in the endosperm. As anticipated, homozygous mutant plants, in which OsSBEIIb was completely inactivated by abolishing the catalytic center and C-terminal regulatory domain, produced opaque seeds with depleted starch reserves. Amylose content in the mutant increased from 19.6 to 27.4% and resistant starch (RS) content increased from 0.2 to 17.2%. Many genes encoding isoforms of AGPase, soluble starch synthase, and other starch branching enzymes were up-regulated, either in their native tissues or in an ectopic manner, whereas genes encoding granule-bound starch synthase, debranching enzymes, pullulanase, and starch phosphorylases were largely down-regulated. There was a general increase in the accumulation of sugars, fatty acids, amino acids, and phytosterols in the mutant endosperm, suggesting that intermediates in the starch biosynthesis pathway increased flux through spillover pathways causing a profound impact on the accumulation of multiple primary and secondary metabolites. Our results provide insights into the broader implications of perturbing starch metabolism in rice endosperm and its impact on the whole plant, which will make it easier to predict the effect of metabolic engineering in cereals for nutritional improvement or the production of valuable metabolites.
Assuntos
Enzima Ramificadora de 1,4-alfa-Glucana/genética , Enzima Ramificadora de 1,4-alfa-Glucana/metabolismo , Oryza/metabolismo , Enzima Ramificadora de 1,4-alfa-Glucana/química , Amilopectina/biossíntese , Amilopectina/química , Amilose/biossíntese , Amilose/química , Metabolismo dos Carboidratos , Grão Comestível/genética , Endosperma/metabolismo , Mutação , Oryza/genética , Proteínas de Plantas/genética , Plantas Geneticamente Modificadas/genética , Sementes/metabolismo , Amido/biossíntese , Sintase do Amido/química , Sintase do Amido/genética , Sintase do Amido/metabolismoRESUMO
The fight against infectious diseases often focuses on epidemics and pandemics, which demand urgent resources and command attention from the health authorities and media. However, the vast majority of deaths caused by infectious diseases occur in endemic zones, particularly in developing countries, placing a disproportionate burden on underfunded health systems and often requiring international interventions. The provision of vaccines and other biologics is hampered not only by the high cost and limited scalability of traditional manufacturing platforms based on microbial and animal cells, but also by challenges caused by distribution and storage, particularly in regions without a complete cold chain. In this review article, we consider the potential of molecular farming to address the challenges of endemic and re-emerging diseases, focusing on edible plants for the development of oral drugs. Key recent developments in this field include successful clinical trials based on orally delivered dried leaves of Artemisia annua against malarial parasite strains resistant to artemisinin combination therapy, the ability to produce clinical-grade protein drugs in leaves to treat infectious diseases and the long-term storage of protein drugs in dried leaves at ambient temperatures. Recent FDA approval of the first orally delivered protein drug encapsulated in plant cells to treat peanut allergy has opened the door for the development of affordable oral drugs that can be manufactured and distributed in remote areas without cold storage infrastructure and that eliminate the need for expensive purification steps and sterile delivery by injection.
Assuntos
Artemisia annua , Doenças Transmissíveis , Preparações Farmacêuticas , Animais , Humanos , Agricultura Molecular , Plantas ComestíveisRESUMO
Infectious diseases, also known as transmissible or communicable diseases, are caused by pathogens or parasites that spread in communities by direct contact with infected individuals or contaminated materials, through droplets and aerosols, or via vectors such as insects. Such diseases cause Ë17% of all human deaths and their management and control places an immense burden on healthcare systems worldwide. Traditional approaches for the prevention and control of infectious diseases include vaccination programmes, hygiene measures and drugs that suppress the pathogen, treat the disease symptoms or attenuate aggressive reactions of the host immune system. The provision of vaccines and biologic drugs such as antibodies is hampered by the high cost and limited scalability of traditional manufacturing platforms based on microbial and animal cells, particularly in developing countries where infectious diseases are prevalent and poorly controlled. Molecular farming, which uses plants for protein expression, is a promising strategy to address the drawbacks of current manufacturing platforms. In this review article, we consider the potential of molecular farming to address healthcare demands for the most prevalent and important epidemic and pandemic diseases, focussing on recent outbreaks of high-mortality coronavirus infections and diseases that disproportionately affect the developing world.
Assuntos
COVID-19 , Doenças Transmissíveis , Doenças Transmissíveis/epidemiologia , Humanos , Pandemias/prevenção & controle , SARS-CoV-2RESUMO
Genome-editing technologies offer unprecedented opportunities for crop improvement with superior precision and speed. This review presents an analysis of the current state of genome editing in the major cereal crops- rice, maize, wheat and barley. Genome editing has been used to achieve important agronomic and quality traits in cereals. These include adaptive traits to mitigate the effects of climate change, tolerance to biotic stresses, higher yields, more optimal plant architecture, improved grain quality and nutritional content, and safer products. Not all traits can be achieved through genome editing, and several technical and regulatory challenges need to be overcome for the technology to realize its full potential. Genome editing, however, has already revolutionized cereal crop improvement and is poised to shape future agricultural practices in conjunction with other breeding innovations.
Assuntos
Sistemas CRISPR-Cas , Produtos Agrícolas/genética , Grão Comestível/genética , Edição de Genes , Genoma de Planta , Melhoramento Vegetal/métodos , Plantas Geneticamente Modificadas/genética , Marcação de GenesRESUMO
The aim of this meta-analysis is to generate large-scale evidence on whether common vascular endothelial growth factor (VEGF) gene polymorphisms (-2578C/A [dbSNP: rs699947] and -460C/T [dbSNP: rs833061]) are associated with lung cancer. A literature search of PubMed, Embase, Web of Science, Cochrane Library, and CBM databases was conducted to identify all eligible studies published before May 3, 2013. Crude odds ratios (ORs) with their corresponding confidence intervals (95% CIs) were used to evaluate the strength of the association. Eleven case-control studies were included with a total of 3,861 lung cancer cases and 3,676 controls in this meta-analysis. For the VEGF -2578C/A polymorphism, the combined results showed that there exist highly significant risk factors for individuals carrying the A allele resulting in lung cancer, and the magnitude of this effect was similar in smoker patients and squamous cell carcinoma (SCC) patients. Unlike the situation with the -2578C/A polymorphism, the VEGF -460C/T polymorphism is not associated with the risk of lung cancer in neither Asians nor Caucasians. However, when stratified according to smoking status and histological types of lung cancer, we found that the T allele (-460C/T) was associated with decreased lung cancer risk among nonsmoker patients and SCC patients. Our findings showed that the -2578C/A polymorphism may increase lung cancer risk, especially in smoker patients and SCC patients, whereas the -460C/T polymorphism may decrease lung cancer risk, especially in nonsmoker patients and SCC patients.
Assuntos
Estudos de Associação Genética , Neoplasias Pulmonares/genética , Polimorfismo de Nucleotídeo Único , Fator A de Crescimento do Endotélio Vascular/genética , Alelos , Estudos de Casos e Controles , Frequência do Gene , Predisposição Genética para Doença , Genótipo , Humanos , Neoplasias Pulmonares/etnologia , Neoplasias Pulmonares/patologia , Razão de Chances , Viés de Publicação , RiscoRESUMO
HMGB3 overexpression has been reported in a variety of human cancers. However, the role of HMGB3 in human non-small cell lung cancer (NSCLC) remains unclear. In this study, the HMGB3 expression was examined at mRNA and protein levels by quantitative real-time reverse transcriptase-polymerase chain reaction (qRT-PCR), Western blotting, and immunohistochemistry in NSCLC tissues and adjacent non-cancerous tissues. Statistical analyses were applied to test the associations between HMGB3 expression, clinicopathologic factors, and prognosis. Western blotting and qRT-PCR showed that the expression levels of HMGB3 mRNA and protein were both significantly higher in NSCLC tissues than those in non-cancerous tissues. Immunohistochemistry analysis showed that HMGB3 expression was significantly correlated with tumor grade, tumor size, clinical stage, and lymph node metastases. The results of Kaplan-Meier analysis indicated that a high expression level of HMGB3 resulted in a significantly poor prognosis of NSCLC patients. Importantly, multivariate analysis showed that high HMGB3 expression was an independent prognostic factor for NSCLC patients. In sum, our data suggest that HMGB3 plays an important role in NSCLC progression, and that overexpression of HMGB3 in tumor tissues could be used as a potential prognostic marker for patients with NSCLC.
Assuntos
Carcinoma Pulmonar de Células não Pequenas/metabolismo , Proteína HMGB3/metabolismo , Neoplasias Pulmonares/metabolismo , Carcinoma Pulmonar de Células não Pequenas/mortalidade , Carcinoma Pulmonar de Células não Pequenas/patologia , Progressão da Doença , Feminino , Expressão Gênica , Proteína HMGB3/genética , Humanos , Estimativa de Kaplan-Meier , Neoplasias Pulmonares/mortalidade , Neoplasias Pulmonares/patologia , Masculino , Pessoa de Meia-Idade , Análise Multivariada , PrognósticoRESUMO
OBJECTIVE: To investigate the effect of selective phosphodiesterase (PDE) 4 inhibitors on nuclear factor kappa B (NF-κB), tumor necrosis factor-α (TNFα) and interleukin-8 (IL-8) secreted by peripheral blood mononuclear cells (PBMCs) in patients diagnosed as rheumatoid arthritis with interstitial lung disease (RA-ILD). METHODS: PBMCs isolated from 15 healthy volunteers (group A) and 20 patients with untreated active RA-ILD (group B) were cultured in vitro. PBMCs from healthy subjects were considered as normal control. PBMCs from RA-ILD patients were divided into four groups with different treatment: blank group (B1), theophylline group (B2), selective PDE4 inhibitor rolipram group (B3), and glucocorticoid group (B4) with dexamethasone. The expression of NF-κB was determined by immunocytochemical staining, and the levels of TNFα and IL-8 in the culture supernatant were detected by enzyme linked immunosorbent assay (ELISA). RESULTS: (1) The activity of NF-κB and the levels of TNFα and IL-8 in group B1 were significant higher than that in group A (P < 0.01). Compared with group B1, three parameters above were similar to those in group B2 (P > 0.05), while group B3 and group B4 had significant decreased levels of three parameters (P < 0.01); IL-8 level in group B4 was significantly lower than that in group B3 (P < 0.05). (2) TNFα and IL-8 levels were positively correlated with NF-κB activity in group B (r = 0.902 and 0.735, P < 0.01 respectively). (3) The reduction of TNFα and IL-8 levels were positively correlated with reduction of NF-κB activity after intervention of rolipram in group B3 (r = 0.874, P < 0.01; r = 0.561, P < 0.05 respectively). CONCLUSION: NF-κB activation and proinflammatory cytokines were involved in the pathogenesis of RA-ILD. selective PDE4 inhibitors may inhibit the production of inflammatory cytokines by inhibiting the activity of the transcription factor NF-κB in PBMC, thus inhibiting the inflammatory reaction of RA-ILD.
Assuntos
Artrite Reumatoide/sangue , Interleucina-8/metabolismo , Doenças Pulmonares Intersticiais/sangue , Subunidade p50 de NF-kappa B/metabolismo , Inibidores da Fosfodiesterase 4/farmacologia , Fator de Necrose Tumoral alfa/metabolismo , Adulto , Idoso , Artrite Reumatoide/complicações , Estudos de Casos e Controles , Células Cultivadas , Feminino , Humanos , Leucócitos Mononucleares/efeitos dos fármacos , Leucócitos Mononucleares/metabolismo , Doenças Pulmonares Intersticiais/complicações , Masculino , Pessoa de Meia-IdadeRESUMO
The engineering of nitrogen fixation in plants requires assembly of an active prokaryotic nitrogenase complex, which is yet to be achieved. Nitrogenase biogenesis relies on NifB, which catalyzes the formation of the [8Fe-9S-C] metal cluster NifB-co. This is the first committed step in the biosynthesis of the iron-molybdenum cofactor (FeMo-co) found at the nitrogenase active site. The production of NifB in plants is challenging because this protein is often insoluble in eukaryotic cells, and its [Fe-S] clusters are extremely unstable and sensitive to O2. As a first step to address this challenge, we generated transgenic rice plants expressing NifB from the Archaea Methanocaldococcus infernus and Methanothermobacter thermautotrophicus. The recombinant proteins were targeted to the mitochondria to limit exposure to O2 and to have access to essential [4Fe-4S] clusters required for NifB-co biosynthesis. M. infernus and M. thermautotrophicus NifB accumulated as soluble proteins in planta, and the purified proteins were functional in the in vitro FeMo-co synthesis assay. We thus report NifB protein expression and purification from an engineered staple crop, representing a first step in the biosynthesis of a functional NifDK complex, as required for independent biological nitrogen fixation in cereals.
Assuntos
Nitrogenase , Oryza , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Compostos de Ferro , Molibdoferredoxina/química , Molibdoferredoxina/metabolismo , Nitrogenase/metabolismo , Oryza/genética , Proteínas Recombinantes/metabolismoRESUMO
Engineering cereals to express functional nitrogenase is a long-term goal of plant biotechnology and would permit partial or total replacement of synthetic N fertilizers by metabolization of atmospheric N2. Developing this technology is hindered by the genetic and biochemical complexity of nitrogenase biosynthesis. Nitrogenase and many of the accessory proteins involved in its assembly and function are O2 sensitive and only sparingly soluble in non-native hosts. We generated transgenic rice plants expressing the nitrogenase structural component, Fe protein (NifH), which carries a [4Fe-4S] cluster in its active form. NifH from Hydrogenobacter thermophilus was targeted to mitochondria together with the putative peptidyl prolyl cis-trans isomerase NifM from Azotobacter vinelandii to assist in NifH polypeptide folding. The isolated NifH was partially active in electron transfer to the MoFe protein nitrogenase component (NifDK) and in the biosynthesis of the nitrogenase iron-molybdenum cofactor (FeMo-co), two fundamental roles for NifH in N2 fixation. NifH functionality was, however, limited by poor [4Fe-4S] cluster occupancy, highlighting the importance of in vivo [Fe-S] cluster insertion and stability to achieve biological N2 fixation in planta. Nevertheless, the expression and activity of a nitrogenase component in rice plants represents the first major step to engineer functional nitrogenase in cereal crops.
Assuntos
Molibdoferredoxina , Oryza , Fertilizantes , Molibdoferredoxina/genética , Molibdoferredoxina/metabolismo , Nitrogenase/genética , Nitrogenase/metabolismo , Oryza/genética , Oryza/metabolismo , Oxirredutases , cis-trans-Isomerases/metabolismoRESUMO
In maize, the shank is a unique tissue linking the stem to the ear. Shank length (SL) mainly affects the transport of photosynthetic products to the ear and the dehydration of kernels via regulated husk morphology. The limited studies on SL revealed it is a highly heritable quantitative trait controlled by significant additive and additive-dominance effects. However, the genetic basis of SL remains unclear. In this study, we analyzed three maize recombinant inbred line (RIL) populations to elucidate the molecular mechanism underlying the SL. The data indicated the SL varied among the three RIL populations and was highly heritable. Additionally, the SL was positively correlated with the husk length (HL), husk number (HN), ear length (EL), and ear weight (EW) in the BY815/K22 (BYK) and CI7/K22 (CIK) RIL populations, but was negatively correlated with the husk width (HW) in the BYK RIL population. Moreover, 10 quantitative trait loci (QTL) for SL were identified in the three RIL populations, five of which were large-effect QTL. The percentage of the total phenotypic variation explained by the QTL for SL was 13.67 %, 20.45 %, and 30.81 % in the BY815/DE3 (BYD), BYK, and CIK RIL populations, respectively. Further analyses uncovered some genetic overlap between SL and EL, SL and ear row number (ERN), SL and cob weight (CW), and SL and HN. Unlike the large-effect QTL qSL BYK-2-2, which spanned the centromere, the other four large-effect QTL were delimited to a single peak bin via bin map. Furthermore, 2, 5, 6, and 12 genes associated with SL were identified for qSL BYK-2-1, qSL CIK-2-1, qSL CIK-9-1, and qSL CIK-9-2, respectively. Five of the candidate genes for SL may contribute to the hormone metabolism and sphingolipid biosynthesis regulating cell elongation, division, differentiation, and expansion. These results may be relevant for future studies on the genetic basis of SL and for the molecular breeding of maize based on marker-assisted selection to develop new varieties with an ideal SL.
Assuntos
Locos de Características Quantitativas/genética , Zea mays/genética , Genes de Plantas/genética , Estudos de Associação Genética , Ligação Genética/genética , Melhoramento Vegetal , Característica Quantitativa Herdável , Zea mays/anatomia & histologiaRESUMO
Light is an essential regulator of many developmental processes in higher plants. We investigated the effect of 4-hydroxy-3-methylbut-2-enyl diphosphate reductase 1/2 genes (OsHDR1/2) and isopentenyl diphosphate isomerase 1/2 genes (OsIPPI1/2) on the biosynthesis of chlorophylls, carotenoids, and phytosterols in 14-day-old etiolated rice (Oyza sativa L.) leaves during de-etiolation. However, little is known about the effect of isoprenoid biosynthesis genes on the corresponding metabolites during the de-etiolation of etiolated rice leaves. The results showed that the levels of α-tocopherol were significantly increased in de-etiolated rice leaves. Similar to 1-deoxy-D-xylulose-5-phosphate synthase 3 gene (OsDXS3), both OsDXS1 and OsDXS2 genes encode functional 1-deoxy-D-xylulose-5-phosphate synthase (DXS) activities. Their expression patterns and the synthesis of chlorophyll, carotenoid, and tocopherol metabolites suggested that OsDXS1 is responsible for the biosynthesis of plastidial isoprenoids in de-etiolated rice leaves. The expression analysis of isoprenoid biosynthesis genes revealed that the coordinated expression of the MEP (2-C-methyl-D-erythritol 4-phosphate) pathway, chlorophyll, carotenoid, and tocopherol pathway genes mirrored the changes in the levels of the corresponding metabolites during de-etiolation. The underpinning mechanistic basis of coordinated light-upregulated gene expression was elucidated during the de-etiolation process, specifically the role of light-responsive cis-regulatory motifs in the promoter region of these genes. In silico promoter analysis showed that the light-responsive cis-regulatory elements presented in all the promoter regions of each light-upregulated gene, providing an important link between observed phenotype during de-etiolation and the molecular machinery controlling expression of these genes.
RESUMO
In this study, we conducted a meta-analysis on high-throughput gene expression data to identify TNF-α-mediated genes implicated in lung cancer. We first investigated the gene expression profiles of two independent TNF-α/TNFR KO murine models. The EGF receptor signaling pathway was the top pathway associated with genes mediated by TNF-α. After matching the TNF-α-mediated mouse genes to their human orthologs, we compared the expression patterns of the TNF-α-mediated genes in normal and tumor lung tissues obtained from humans. Based on the TNF-α-mediated genes that were dysregulated in lung tumors, we developed a prognostic gene signature that effectively predicted recurrence-free survival in lung cancer in two validation cohorts. Resampling tests suggested that the prognostic power of the gene signature was not by chance, and multivariate analysis suggested that this gene signature was independent of the traditional clinical factors and enhanced the identification of lung cancer patients at greater risk for recurrence.