A non-invasive method for silencing gene transcription in honeybees maintained under natural conditions

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Epigenetic Silencing of CRABP2 and MX1 in Head and Neck Tumors

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

The post-transcriptional gene silencing pathway in Eucalyptus

Post-transcriptional gene silencing (PTGS) is a conserved surveillance mechanism that identifies and cleaves double-stranded RNA molecules and their cellular cognate transcripts. The RNA silencing response is actually used as a powerful technique (named RNA interference) for potent and specific inhibition of gene expression in several organisms. To identify gene products in Eucalyptus sharing similarities with enzymes involved in the PTGS pathway, we queried the expressed sequence tag database of the Brazilian Eucalyptus Genome Sequence Project Consortium (FORESTs) with the amino acid sequences of known PTGS-related proteins. Among twenty-six prospected genes, our search detected fifteen assembled sequences encoding products presenting high level of similarity (E value < 10 -40) to proteins involved in PTGS in plants and other organisms. We conclude that most of the genes known to be involved in the PTGS pathway are represented in the FORESTs database. Copyright by the Brazilian Society of Genetics.

Dioctadecyldimethylammonium:Monoolein Nanocarriers for Efficient in Vitro Gene Silencing

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Estudos funcionais da rgs-CaM, uma calmodulina envolvida na supressão de silenciamento por RNA

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Silencing mitogen-activated protein kinase-activated protein kinase-2 arrests inflammatory bone loss

p38 mitogen-activated protein kinases (MAPKs) are critical for innate immune signaling and subsequent cytokine expression in periodontal inflammation and bone destruction. In fact, previous studies show that systemic p38 MAPK inhibitors block periodontal disease progression. However, development of p38 MAPK inhibitors with favorable toxicological profiles is difficult. Here, we report our findings regarding the contribution of the downstream p38 MAPK substrate, mitogen-activated protein kinase-activated protein kinase 2 (MK2 or MAPKAPK-2), in immune response modulation in an experimental model of pathogen-derived lipopolysaccharide (LPS)-induced periodontal bone loss. To determine whether small interfering RNA (siRNA) technology has intraoral applications, we initially validated MK2 siRNA specificity. Then, gingival tissue surrounding maxillary molars of rats was injected with MK2 siRNA or scrambled siRNA at the palatal regions of bone loss. Intraoral tissues treated with MK2 siRNA had significantly less MK2 mRNA expression compared with scrambled siRNA-treated tissues. MK2 siRNA delivery arrested LPS-induced inflammatory bone loss, decreased inflammatory infiltrate, and decreased osteoclastogenesis. This proof-of-concept study suggests a novel target using an intraoral RNA interference strategy to control periodontal inflammation.

Efeito da aplicação de extratos aquosos em couve na alimentação de largatas de Ascia monuste orseis

Os efeitos de extratos aquosos de amêndoas de Azadirachta indica e frutos de Sapindus saponaria, aplicados em discos de folhas de couve (Brassica oleracea var. acephala) nas concentrações de 0,0117% e 1,0342% (p/v), respectivamente, foram estudados sobre a alimentação das lagartas de Ascia monuste orseis. Avaliou-se a atratividade e o consumo de área foliar de lagartas de primeiro e terceiro ínstar durante 24 horas, em condições de laboratório (T = 25 ± 2ºC, UR = 60 ± 10% e fotofase = 12 horas). Nos testes com e sem chance de escolha, para lagartas de primeiro ínstar e teste sem chance de escolha para lagartas de terceiro ínstar, não houve diferença quanto à atratividade das lagartas. No teste com chance de escolha para lagartas de terceiro ínstar, houve menor atratividade das lagartas pelos discos de folhas tratadas com S. saponaria, diferindo da testemunha. No decorrer de 24 horas de avaliações, pode-se observar 58,3% de lagartas atraídas na testemunha, não diferindo de A. indica e diferindo de S. saponaria, com 39,3% e 2,4% das lagartas atraídas, respectivamente. Quanto ao consumo de área foliar, o extrato de S. saponaria diminuiu o consumo de lagartas, em todos os testes realizados. Quando não tinham opção de escolha para se alimentar de folhas sem os extratos, as lagartas consumiram as folhas tratadas nas concentrações testadas, porém em menor quantidade. Os extratos testados neste experimento demonstram ter efeitos sobre a alimentação das lagartas de A. monuste orseis, possivelmente com propriedades deterrentes e/ou supressoras de alimentação.

Quantitative control of Lettuce mosaic virus fitness and host defence inhibition by P1-HCPro

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

DNA methylation in the CTCF-binding site I and the expression pattern of the H19 gene: Does positive expression predict poor prognosis in early stage head and neck carcinomas?

Loss of allele-specific expression by the imprinted genes IGF2 and H19 has been correlated with a differentially methylated region (DMR) upstream to the H19 gene. The H19-DMR contains seven potential CCCTC-binding factor (CTCF) binding sites. CTCF is a chromatin insulator and a multifunctional transcription factor whose binding to the H19-DMR is suppressed by DNA methylation. Our study included a group of 41 head and neck squamous cell carcinoma (HNSCC) samples. The imprinting status of the H19 gene was analyzed in 11 out of 35 positive cases for H19 gene expression, and only 1 of them showed loss of imprinting. We detected a significant correlation (P=0.041, Fisher's exact test) between H19 expression and tumor recurrence. Among H19 positive cases, six were T2, in which five developed recurrence and/or metastasis. Inversely, in the group of tumors that showed no H19 gene expression, 5 out of 24 were T2 and only I presented regional recurrence. These data support the hypothesis that H19 expression could be used as a prognostic marker to indicate recurrence in early stage tumors. We also examined the methylation of the CTCF binding site 1 in a subgroup of these samples. The H19 gene silencing and loss of imprinting were not correlated with the methylation pattern of the CTCF binding site 1. However, the significant correlation between H19 expression and tumor recurrence suggest that this transcript could be a marker for the progression of HNSCC. (c) 2005 Wiley-Liss, Inc.

Loss of Expression and Function of SOCS3 Is an Early Event in HNSCC: Altered Subcellular Localization as a Possible Mechanism Involved in Proliferation, Migration and Invasion

Background: Suppressor of cytokine signaling 3 (SOCS3) is an inducible endogenous negative regulator of signal transduction and activator of transcription 3 (STAT3). Epigenetic silencing of SOCS3 has been shown in head and neck squamous cell carcinoma (HNSCC), which is associated with increased activation of STAT3. There is scarce information on the functional role of the reduction of SOCS3 expression and no information on altered subcellular localization of SOCS3 in HNSCC.Methodology/Principal Findings: We assessed endogenous SOCS3 expression in different HNSCC cell lines by RT-qPCR and western blot. Immunofluorescence and western blot were used to study the subcellular localization of endogenous SOCS3 induced by IL-6. Overexpression of SOCS3 by CMV-driven plasmids and siRNA-mediated inhibition of endogenous SOCS3 were used to verify the role of SOCS3 on tumor cell proliferation, viability, invasion and migration in vitro. In vivo relevance of SOCS3 expression in HNSCC was studied by quantitative immunohistochemistry of commercially-available tissue microarrays. Endogenous expression of SOCS3 was heterogeneous in four HNSCC cell lines and surprisingly preserved in most of these cell lines. Subcellular localization of endogenous SOCS3 in the HNSCC cell lines was predominantly nuclear as opposed to cytoplasmic in non-neoplasic epithelial cells. Overexpression of SOCS3 produced a relative increase of the protein in the cytoplasmic compartment and significantly inhibited proliferation, migration and invasion, whereas inhibition of endogenous nuclear SOCS3 did not affect these events. Analysis of tissue microarrays indicated that loss of SOCS3 is an early event in HNSCC and was correlated with tumor size and histological grade of dysplasia, but a considerable proportion of cases presented detectable expression of SOCS3.Conclusion: Our data support a role for SOCS3 as a tumor suppressor gene in HNSCC with relevance on proliferation and invasion processes and suggests that abnormal subcellular localization impairs SOCS3 function in HNSCC cells.

DNA methylation patterns of the CDHI, RARB, and SFN genes in choroid plexus tumors

Genetic and epigenetic alterations in choroid plexus tumors, a rare neuroepithelial neoplasm most frequently detected in children, are poorly characterized. Epigenetic silencing associated with aberrant CpG island methylation is one mechanism leading to the loss of tumor suppressor functions in cancer cells. Using methylation-specific polymerase chain reaction, the methylation patterns of the genes CDH1 (E-cadherin), RARB (retinoic acid receptor, beta), and SFN (stratifin; 14-3-3 sigma) were retrospectively investigated in eight choroid plexus tumors (five papillomas, two atypical papillomas, and one carcinoma), as well as in two normal cortexes obtained after autopsy from male individuals aged 6 months and 64 years. Among the six pediatric tumors, the mean age at diagnosis was 1.8 years old (range, 0.2-6) and the two adult tumors were detected in a 66-year-old man and a 45-year-old woman. A high frequency of hypermethylation was detected in CDH1 and SFN genes in tumoral and normal cortex tissues. Tumor-specific RARB hypermethylation was observed in four papillomas. Further studies are required to evaluate the role of aberrant methylation in choroid plexus tumor progression. (c) 2007 Elsevier B.V. All rights reserved.

Strand Analysis, a free online program for the computational identification of the best RNA interference (RNAi) targets based on Gibbs free energy

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Analysis of plant LTR-retrotransposons at the fine-scale family level reveals individual molecular patterns

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Evidence of epigenetic regulation of the tumor suppressor gene cluster flanking RASSF1 in breast cancer cell lines

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)