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1.
BMC Genomics ; 21(1): 123, 2020 Feb 04.
Artigo em Inglês | MEDLINE | ID: mdl-32019511

RESUMO

BACKGROUND: Duck hepatitis A virus type 3 (DHAV-3) is one of the most harmful pathogens in the duck industry. However, the molecular mechanism underlying DHAV-3 infection in ducklings remains poorly understood. To study the genetic regulatory network for miRNA-mRNA and the signaling pathways involved in DHAV-3 infection in ducklings, we conducted global miRNA and mRNA expression profiling of duckling liver tissues infected with lethal DHAV-3 by high-throughput sequencing. RESULTS: We found 156 differentially expressed miRNAs (DEMs) and 7717 differentially expressed genes (DEGs) in livers of mock-infected and DHAV-3-infected duckling. A total of 19,606 miRNA-mRNA pairs with negatively correlated expression patterns were identified in miRNA-mRNA networks constructed on the basis of these DEMs and DEGs. Moreover, immune-related pathways, including the cytokine-cytokine receptor interaction, apoptosis, Toll-like receptor, Jak-STAT, and RIG-I-like receptor signaling pathway, were significantly enriched through analyzing functions of mRNAs in the network in response to DHAV-3 infection. Furthermore, apl-miR-32-5p, apl-miR-125-5p, apl-miR-128-3p, apl-miR-460-5p, and novel-m0012-3p were identified as potential regulators in the immune-related signaling pathways during DHAV-3 infection. And some host miRNAs were predicted to target the DHAV-3 genome. CONCLUSIONS: This is the first integrated analysis of miRNA and mRNA in DHAV-3-infected ducklings. The results indicated the important roles of miRNAs in regulating immune response genes and revealed the immune related miRNA-mRNA regulation network in the DHAV-3-infected duckling liver. These findings increase our knowledge of the roles of miRNAs and their target genes in DHAV-3 replication and pathogenesis. They also aid in the understanding of host-virus interactions.

2.
Med Mycol ; 58(2): 181-186, 2020 Feb 01.
Artigo em Inglês | MEDLINE | ID: mdl-31131856

RESUMO

Talaromyces (Penicillium) marneffei is an emerging pathogen that causes significant morbidity and mortality in immunocompromised patients in endemic regions such as southeast Asia. The diagnosis of disseminated T. marneffei infection remains challenging in clinical practice. In the study, a well-validated real-time quantitative polymerase chain reaction (qPCR) target region of ITS1-5.8S-ITS2 and a Platelia galactomannan (GM) assay were compared for their diagnostic performance using serum samples from patients with or without human immunodeficiency virus (HIV). The results showed that this novel qPCR method is highly sensitive and specific for T. marneffei DNA detection in serum samples, and the limit of detection and species-specificity of qPCR were five copies of DNA and 100%, respectively. For detection in serum samples from 36 talaromycosis patients, the sensitivity of qPCR was 86.11% (31/36), including 20/20 (100%) patients with fungemia and 11/16 (68.75%) patients without fungemia. For the GM assay, the sensitivity was 80.56% (29/36) when the GM optical density cutoff index was ≥0.5, including 19/20 (95%) patients with fungemia and 10/16 (62.5%) patients without fungemia. These results indicate that the novel qPCR and GM assays can be used as a valuable tool in the diagnosis of T. marneffei infection. Serum samples are convenient hematological specimens for T. marneffei DNA quantification. Combining the GM assay and qPCR is more scientific and appropriate for diagnosing T. marneffei infection in endemic areas.

3.
Emerg Microbes Infect ; 8(1): 367-376, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31851879

RESUMO

The dimorphic fungus Talaromyces marneffei (TM) is a common cause of HIV-associated opportunistic infections in Southeast Asia. Cotrimoxazole (CTX) inhibits folic acid synthesis which is important for the survival of many bacteria, protozoa, and fungi and has been used to prevent several opportunistic infections among HIV/AIDS patients. We question whether CTX is effective in preventing TM infection. To investigate this question, we conducted an 11-year (2005-2016) retrospective observational cohort study of all patients on the Chinese national antiretroviral therapy (ART) programme in Guangxi, a province with high HIV and TM burden in China. Survival analysis was conducted to investigate TM cumulative incidence, and Cox regression and propensity score matching (PSM) were used to evaluate the effect of CTX on TM incidence. Of the 3359 eligible individuals contributing 10,504.66 person-years of follow-up, 81.81% received CTX within 6 months after ART initiation, and 4.73% developed TM infection, contributing 15.14/1,000 person-year TM incidence rate. CTX patients had a significantly lower incidence of TM infection than non-CTX patients (4.11% vs. 7.53%; adjusted hazard ratio (aHR) = 0.50, 95% CI 0.35-0.73). CTX reduced TM incidence in all CD4+ cell subgroups (<50 cells/µL, 50-99 cells/µL, 100-199 cells/µL), with the highest reduction observed in patients with a baseline CD4+ cell count <50 cells/µL in both Cox regression and the PSM analyses. In conclusion, in addition to preventing other HIV-associated opportunistic infections, CTX prophylaxis has the potential to prevent TM infection in HIV/AIDS patients receiving ART.

4.
Sci Rep ; 9(1): 7816, 2019 May 24.
Artigo em Inglês | MEDLINE | ID: mdl-31127157

RESUMO

Previous studies investigating HIV-infected patients suggested a direct link between underweight and the mortality rate of AIDS. However, there was a lack of evidence showing the optimal range of initial body mass index (BMI) patients maintain during antiretroviral therapy (ART). We aimed to evaluate associations of the BMI values pre-ART and during the entire ART duration with mortality among HIV-positive individuals. In total, 5101 HIV/AIDS patients, including 1439 (28.2%) underweight, 3047 (59.7%) normal-weight, 548 (10.7%) overweight and 67 (1.3%) obese patients, were included in this cohort. The cumulative mortality of underweight, normal-weight, and overweight were 2.4/100 person-years (95% CI 1.9-2.9), 1.1/100 person-years (95% CI 0.9-1.3), and 0.5/100 person-years (95% CI 0.1-0.9), respectively. Cumulative mortality was lower in both the normal-weight and overweight populations than in the underweight population, with an adjusted hazard ratio (AHR) of 0.5 (95% CI 0.4-0.7, p < 0.001) and 0.3 (95% CI 0.1-0.6, p = 0.002), respectively. Additionally, in the 1176 patients with available viral load data, there was significant difference between the underweight and normal-weight groups after adjustment for all factors, including viral load (p = 0.031). This result suggests that HIV-infected patients in Guangxi maintaining a BMI of 19-28 kg/m2, especially 24-28 kg/m2, have a reduced risk of death.

5.
Mol Med Rep ; 18(1): 77-86, 2018 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-29749506

RESUMO

Lymph node (LN) fibrosis resulting in cluster of differentiation (CD) 4+ T cell reduction following human immunodeficiency virus (HIV) infection is an important step in the pathogenesis of acquired immunodeficiency syndrome. The mechanisms mediating LN fibrosis following HIV infection have not been completely elucidated. In order to investigate the mechanism of LN fibrosis, the expression of transforming growth factor (TGF)­ß1 was determined in the LNs of HIV­infected individuals by immunohistochemistry and fluorescence­based flow cytometry. The effect of stimulated CD8+ T cells on collagen secretion by fibroblasts was detected using immunofluorescence staining and western blot analysis. The results demonstrated that the LNs of HIV­infected individuals exhibited a significantly increased proportion of CD8+ T cells with high TGF­ß1 expression. These CD8+ T cells demonstrated increased CD38 and programmed cell death protein 1 expression and decreased CD127 expression compared with the controls. CD8+ T cells from the LNs of non­HIV infected individuals expressed a high TGF­ß1 level following stimulation with phorbol­12­myristate 13­acetate. These CD8+T cells subsequently induced the secretion of a large amount of type I collagen in human lymphatic fibroblasts. The results of the present study indicated that CD8+ T cells with high TGF­ß1 expression served an important role in LN fibrosis following HIV infection.


Assuntos
Síndrome de Imunodeficiência Adquirida/imunologia , Linfócitos T CD8-Positivos/imunologia , Regulação da Expressão Gênica/imunologia , Linfonodos/imunologia , Fator de Crescimento Transformador beta1/imunologia , Síndrome de Imunodeficiência Adquirida/patologia , Adulto , Linfócitos T CD8-Positivos/patologia , Feminino , Fibrose , Humanos , Linfonodos/patologia , Masculino , Pessoa de Meia-Idade
6.
BMC Plant Biol ; 17(1): 198, 2017 Nov 13.
Artigo em Inglês | MEDLINE | ID: mdl-29132307

RESUMO

BACKGROUND: Shikonin is a naphthoquinone secondary metabolite with important medicinal value and is found in Lithospermum erythrorhizon. Considering the limited knowledge on the membrane transport mechanism of shikonin, this study investigated such molecular mechanism. RESULTS: We successfully isolated an ATP-binding cassette protein gene, LeMDR, from L. erythrorhizon. LeMDR is predominantly expressed in L. erythrorhizon roots, where shikonin accumulated. Functional analysis of LeMDR by using the yeast cell expression system revealed that LeMDR is possibly involved in the shikonin efflux transport. The accumulation of shikonin is lower in yeast cells transformed with LeMDR-overexpressing vector than that with empty vector. The transgenic hairy roots of L. erythrorhizon overexpressing LeMDR (MDRO) significantly enhanced shikonin production, whereas the RNA interference of LeMDR (MDRi) displayed a reverse trend. Moreover, the mRNA expression level of LeMDR was up-regulated by treatment with shikonin and shikonin-positive regulators, methyl jasmonate and indole-3-acetic acid. There might be a relationship of mutual regulation between the expression level of LeMDR and shikonin biosynthesis. CONCLUSIONS: Our findings demonstrated the important role of LeMDR in transmembrane transport and biosynthesis of shikonin.


Assuntos
Transportadores de Cassetes de Ligação de ATP/fisiologia , Lithospermum/metabolismo , Naftoquinonas/metabolismo , Transportadores de Cassetes de Ligação de ATP/genética , Transporte Biológico , Southern Blotting , Clonagem Molecular , Regulação da Expressão Gênica de Plantas , Genes de Plantas/genética , Genes de Plantas/fisiologia , Raízes de Plantas/metabolismo , Plantas Geneticamente Modificadas , Análise de Sequência de DNA
7.
Sci Rep ; 7(1): 4477, 2017 06 30.
Artigo em Inglês | MEDLINE | ID: mdl-28667265

RESUMO

Shikonin and its derivatives extracted from Lithospermeae plants' red roots have current applications in food and pharmaceutical industries. Previous studies have cloned some genes related to shikonin biosynthesis. However, most genes related to shikonin biosynthesis remain unclear, because the lack of the genome/transcriptome of the Lithospermeae plants. Therefore, in order to provide a new understanding of shikonin biosynthesis, we obtained transcriptome data and unigenes expression profiles in three shikonin-producing Lithospermeae plants, i.e., Lithospermum erythrorhizon, Arnebia euchroma and Echium plantagineum. As a result, two unigenes (i.e., G10H and 12OPR) that are involved in "shikonin downstream biosynthesis" and "methyl jasmonate biosynthesis" were deemed to relate to shikonin biosynthesis in this study. Furthermore, we conducted a Lamiids phylogenetic model and identified orthologous unigenes under positive selection in above three Lithospermeae plants. The results indicated Boraginales was more relative to Solanales/Gentianales than to Lamiales.


Assuntos
Evolução Biológica , Vias Biossintéticas/genética , Regulação da Expressão Gênica de Plantas , Lithospermum/genética , Lithospermum/metabolismo , Naftoquinonas/metabolismo , Transcriptoma , Boraginaceae/genética , Boraginaceae/metabolismo , Cromatografia Líquida de Alta Pressão , Biologia Computacional/métodos , Perfilação da Expressão Gênica , Sequenciamento de Nucleotídeos em Larga Escala , Lithospermum/classificação , Anotação de Sequência Molecular , Naftoquinonas/análise , Filogenia , Seleção Genética
8.
PLoS One ; 11(7): e0159810, 2016.
Artigo em Inglês | MEDLINE | ID: mdl-27463682

RESUMO

MicroRNAs (miRNAs) play an important role in plant growth, development, and response to environment. For identifying and comparing miRNAs and their targets in seed development between two maize inbred lines (i.e. PH6WC and PH4CV), two sRNAs and two degradome libraries were constructed. Through high-throughput sequencing and miRNA identification, 55 conserved and 24 novel unique miRNA sequences were identified in two sRNA libraries; moreover, through degradome sequencing and analysis, 137 target transcripts corresponding to 38 unique miRNA sequences were identified in two degradome libraries. Subsequently, 16 significantly differentially expressed miRNA sequences were verified by qRT-PCR, in which 9 verified sequences obviously target 30 transcripts mainly involved with regulation in flowering and development in embryo. Therefore, the results suggested that some miRNAs (e.g. miR156, miR171, miR396 and miR444) related reproductive development might differentially express in seed development between the PH6WC and PH4CV maize inbred lines in this present study.


Assuntos
MicroRNAs/genética , RNA de Plantas/genética , Sementes/genética , Zea mays/genética , Sequência Conservada , Regulação da Expressão Gênica no Desenvolvimento , Regulação da Expressão Gênica de Plantas , Endogamia , Sementes/crescimento & desenvolvimento , Zea mays/crescimento & desenvolvimento
9.
BMC Plant Biol ; 16(1): 121, 2016 05 26.
Artigo em Inglês | MEDLINE | ID: mdl-27230755

RESUMO

BACKGROUND: The phytohormone ethylene (ET) is a key signaling molecule for inducing the biosynthesis of shikonin and its derivatives, which are secondary metabolites in Lithospermum erythrorhizon. Although ETHYLENE INSENSITIVE3 (EIN3)/EIN3-like proteins (EILs) are crucial transcription factors in ET signal transduction pathway, the possible function of EIN3/EIL1 in shikonin biosynthesis remains unknown. In this study, by targeting LeEIL-1 (L. erythrorhizon EIN3-like protein gene 1) at the expression level, we revealed the positive regulatory effect of LeEIL-1 on shikonin formation. RESULTS: The mRNA level of LeEIL-1 was significantly up-regulated and down-regulated in the LeEIL-1-overexpressing hairy root lines and LeEIL-1-RNAi hairy root lines, respectively. Specifically, LeEIL-1 overexpression resulted in increased transcript levels of the downstream gene of ET signal transduction pathway (LeERF-1) and a subset of genes for shikonin formation, excretion and/or transportation (LePAL, LeC4H-2, Le4CL-1, HMGR, LePGT-1, LeDI-2, and LePS-2), which was consistent with the enhanced shikonin contents in the LeEIL-1-overexpressing hairy root lines. Conversely, LeEIL-1-RNAi dramatically repressed the expression of the above genes and significantly reduced shikonin production. CONCLUSIONS: The results revealed that LeEIL-1 is a positive regulator of the biosynthesis of shikonin and its derivatives in L. erythrorhizon hairy roots. Our findings gave new insights into the molecular regulatory mechanism of ET in shikonin biosynthesis. LeEIL-1 could be a crucial target gene for the genetic engineering of shikonin biosynthesis.


Assuntos
Regulação da Expressão Gênica de Plantas , Lithospermum/genética , Naftoquinonas/metabolismo , Proteínas de Plantas/genética , Fatores de Transcrição/genética , Lithospermum/metabolismo , Proteínas de Plantas/metabolismo , Raízes de Plantas/metabolismo , Plantas Geneticamente Modificadas/genética , Plantas Geneticamente Modificadas/metabolismo , Interferência de RNA , Fatores de Transcrição/metabolismo
10.
Int J Mol Sci ; 17(2): 219, 2016 Feb 06.
Artigo em Inglês | MEDLINE | ID: mdl-26861313

RESUMO

The revelation of mechanisms of photodynamic therapy (PDT) at the cellular level as well as singlet oxygen (¹O2) as a second messengers requires the quantification of intracellular ¹O2. To detect singlet oxygen, directly measuring the phosphorescence emitted from ¹O2 at 1270 nm is simple but limited for the low quantum yield and intrinsic efficiency of ¹O2 emission. Another method is chemically trapping ¹O2 and measuring fluorescence, absorption and Electron Spin Resonance (ESR). In this paper, we used indocyanine green (ICG), the only near-infrared (NIR) probe approved by the Food and Drug Administration (FDA), to detect ¹O2 in vitro. Once it reacts with ¹O2, ICG is decomposed and its UV absorption at 780 nm decreases with the laser irradiation. Our data demonstrated that ICG could be more sensitive and accurate than Singlet Oxygen Sensor Green reagent(®) (SOSG, a commercialized fluorescence probe) in vitro, moreover, ICG functioned with Eosin Y while SOSG failed. Thus, ICG would reasonably provide the possibility to sense ¹O2 in vitro, with high sensitivity, selectivity and suitability to most photosensitizers.


Assuntos
Corantes Fluorescentes , Verde de Indocianina , Raios Infravermelhos , Oxigênio Singlete/análise , Fotoquimioterapia , Fármacos Fotossensibilizantes , Reprodutibilidade dos Testes , Sensibilidade e Especificidade , Espectrofotometria Ultravioleta
11.
Plant Mol Biol ; 90(4-5): 345-58, 2016 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-26780904

RESUMO

The phytohormone ethylene (ET) is a crucial signaling molecule that induces the biosynthesis of shikonin and its derivatives in Lithospermum erythrorhizon shoot cultures. However, the molecular mechanism and the positive regulators involved in this physiological process are largely unknown. In this study, the function of LeACS-1, a key gene encoding the 1-aminocyclopropane-1-carboxylic acid synthase for ET biosynthesis in L. erythrorhizon hairy roots, was characterized by using overexpression and RNA interference (RNAi) strategies. The results showed that overexpression of LeACS-1 significantly increased endogenous ET concentration and shikonin production, consistent with the up-regulated genes involved in ET biosynthesis and transduction, as well as the genes related to shikonin biosynthesis. Conversely, RNAi of LeACS-1 effectively decreased endogenous ET concentration and shikonin production and down-regulated the expression level of above genes. Correlation analysis showed a significant positive linear relationship between ET concentration and shikonin production. All these results suggest that LeACS-1 acts as a positive regulator of ethylene-induced shikonin biosynthesis in L. erythrorhizon hairy roots. Our work not only gives new insights into the understanding of the relationship between ET and shikonin biosynthesis, but also provides an efficient genetic engineering target gene for secondary metabolite production in non-model plant L. erythrorhizon.


Assuntos
Etilenos/farmacologia , Regulação da Expressão Gênica de Plantas/fisiologia , Lithospermum/metabolismo , Liases/metabolismo , Naftoquinonas/metabolismo , Raízes de Plantas/metabolismo , Clonagem Molecular , Biologia Computacional , DNA Complementar/genética , DNA Complementar/metabolismo , Liases/genética , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Raízes de Plantas/efeitos dos fármacos , Plantas Geneticamente Modificadas , Transdução de Sinais/fisiologia
12.
Mol Med Rep ; 13(1): 731-43, 2016 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-26647762

RESUMO

Arginase I (Arg I) and inducible nitric oxide synthase (iNOS) are important in regulating immune functions through their metabolites. Previous studies have revealed that the expression of Arg I is increased and the expression of iNOS is reduced in the serum and peripheral blood mononuclear cells of human immunodeficiency virus (HIV)­infected patients. As one of the most important immune organs and HIV replication sites, whether similar changes are present in the lymph nodes following HIV infection remains to be elucidated. To investigate this, the present study collected lymph node and blood specimens from 52 HIV­infected patients to measure the expression levels of Arg I and iNOS by immunohistochemistry and fluoresence­based flow cytometry. Compared with control subjects without HIV infection, the patients with HIV had significantly higher expression levels of Arg I in the lymph nodes and higher frequencies of Arg I+ CD4+ T cells and CD8+ T cells in the blood and lymph nodes, and these results were contrary the those of iNOS in the corresponding compartments. The expression levels of Arg I in the lymph nodes and blood were negatively associated with peripheral CD4+ T cell count and positively associated with viral load. However, the expression levels of iNOS in the lymph nodes and blood were positively associated with peripheral CD4+ T cell count and negatively associated with viral load. These results showed that alterations in the expression levels of Arg I and iNOS in the peripheral T cells and peripheral nodes of HIV infected patients are associated with disease progression in these patients. These results indicate a potential to therapeutic strategy for delaying disease progression through regulating and manipulating the expression levels of Arg I and iNOS in patients infected with HIV.


Assuntos
Arginase/sangue , Soropositividade para HIV/sangue , Soropositividade para HIV/enzimologia , Linfonodos/enzimologia , Óxido Nítrico Sintase Tipo II/sangue , Adulto , Idoso , Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD8-Positivos/imunologia , Feminino , Citometria de Fluxo , Soropositividade para HIV/imunologia , Humanos , Contagem de Linfócitos , Masculino , Pessoa de Meia-Idade , Carga Viral , Adulto Jovem
13.
Mol Biol Rep ; 41(3): 1623-30, 2014 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-24398552

RESUMO

The gene encoding cathepsin D of silkworm, Bombyx mori (BmCatD) is specifically expressed in the larval fat body and pupal gut, and plays an important role in the programmed cell death during metamorphosis. To identify element involved in this transcription-dependent spatial restriction, truncation and deletion of the 5' terminal from the BmCatD promoter were conducted in vivo. The recombinant AcMNPV vector (Autographa californica multiple nucleopolyhedrovirus) with a dual-luciferase quantitative assay system was used as the transfer. A 289 bp DNA sequence (-1,214 to -925) upstream of the transcriptional start site is found to be responsible for promoting tissue-specific transcription. Further analysis of a series of deletion within the 289 bp region of overlapping deletion showed that a 33 bp region (-1,071 to -1,038) sequence suppresses the ectopic expression of the BmCatD promoter. These results suggest that this 33 bp region could function as a promoter element in the tissue-specificity expression.


Assuntos
Bombyx/genética , Catepsina D/genética , Regiões Promotoras Genéticas , Animais , Catepsina D/biossíntese , Regulação da Expressão Gênica no Desenvolvimento , Larva/genética , Especificidade de Órgãos/genética
14.
World J Microbiol Biotechnol ; 28(5): 2029-38, 2012 May.
Artigo em Inglês | MEDLINE | ID: mdl-22806024

RESUMO

Two acidic polysaccharide fractions, CM-jd-CPS2 and CM-jd(Y)-CPS2, were isolated from the fruiting bodies of cultured Cordyceps militaris grown on solid rice medium and silkworm pupa, respectively, by hot-water extraction, ethanol precipitation and fractionation using ion-exchange column (DEAE-cellulose-52) and gel-filtration column (Sephadex G-100) chromatography. Their structural characterizations were performed by gas chromatography and fourier-transform infrared spectroscopy. Some differences existed between their structures, which indicated that culture media could influence the structure of polysaccharides of C. militaris. The antioxidant activities of CM-jd-CPS2 and CM-jd(Y)-CPS2 were evaluated by various methods in vitro. They had strong 2,2-diphenyl-1-picrylhydrazyl radical-scavenging activity and ferrous ion-chelating capacity, but moderate reducing power. The antioxidant activities of CM-jd(Y)-CPS2 were slightly higher than those of CM-jd-CPS2. These two acidic fractions were evaluated for proliferation of mouse splenocyte activity in vitro. They both possessed does-dependent mitogenic effects on mouse splenocytes, and could synergistically promote murine T- and B-lymphocytes induced by Con A and LPS. CM-jd(Y)-CPS2 exhibited stronger stimulatory activities upon immunomodulation than CM-jd-CPS2. These results are beneficial for the interpretation of the connection between polysaccharide structures and their biological activities.


Assuntos
Antioxidantes/química , Antioxidantes/isolamento & purificação , Cordyceps/química , Cordyceps/crescimento & desenvolvimento , Polissacarídeos/química , Polissacarídeos/isolamento & purificação , Animais , Antioxidantes/farmacologia , Bombyx/microbiologia , Proliferação de Células/efeitos dos fármacos , Quelantes/química , Quelantes/isolamento & purificação , Quelantes/farmacologia , Cromatografia Gasosa , Cromatografia em Gel , Cromatografia por Troca Iônica , Depuradores de Radicais Livres/química , Depuradores de Radicais Livres/isolamento & purificação , Depuradores de Radicais Livres/farmacologia , Leucócitos Mononucleares/efeitos dos fármacos , Camundongos , Oryza/microbiologia , Polissacarídeos/farmacologia , Pupa/microbiologia , Espectroscopia de Infravermelho com Transformada de Fourier
15.
Biochem Biophys Res Commun ; 425(1): 113-8, 2012 Aug 17.
Artigo em Inglês | MEDLINE | ID: mdl-22828514

RESUMO

Bombyx mori Cathepsin D (BmCatD) is specifically expressed in the fat body, and plays a critical role for the programmed cell death of the larval fat body and pupal gut during metamorphosis. To better understand the transcriptional control of BmCatD expression, we conducted this study to identify the ecdysone response elements (EcREs) in the BmCatD promoter and clarify their regulational functions. We inserted EcREs into a recombinant AcMNPV (Autographa californica multiple nucleopolyhedrovirus) vector and performed luciferase assay with a dual-luciferase quantitative assay system. Three putative EcREs were located at positions -109 to -99, -836 to -826 and -856 to -846 relative to the transcription start site. Overlapping deletion studies of this EcRE region showed that the three EcREs could suppress the ectopic expression of the BmCatD promoter. EcRE mutations resulted in the loss of the fat body-specific expression of the BmCatD gene. These results suggest that the EcREs are vital for activation of the promoter by 20-hydroxyecdysone (20E) in the larval fat body and further support the crucial role of ecdysone signaling to control cathepsin D gene transcription. It may suggest that the heterodimeric complex EcR/USP mediates the activation of ecdysone-dependent BmCatD transcription in the larval fat body of B. mori.


Assuntos
Bombyx/genética , Catepsina D/genética , Ecdisona/fisiologia , Elementos de Resposta/genética , Ativação Transcricional , Animais , Sequência de Bases , Bombyx/crescimento & desenvolvimento , Ecdisona/farmacologia , Ecdisterona/farmacologia , Luciferases/genética , Dados de Sequência Molecular , Mutação , Regiões Promotoras Genéticas/efeitos dos fármacos , Regiões Promotoras Genéticas/genética , Elementos de Resposta/efeitos dos fármacos , Sítio de Iniciação de Transcrição
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