Produtividade de híbridos de milho pipoca submetidos a diferentes populações na semeadura / Productivity of poppy corn hybrids submitted to different populations in seeding / Productividad de híbridos de maíz poppy presentados a poblaciones diferentes en la sembrada

Quando se trata de milho pipoca o Brasil é o segundo maior produtor, mas necessitando ainda de importações para suprir a demanda interna do país, visto a importância de estudos que melhor explorem as recomendações agronômicas para a cultura do milho pipoca. Assim, o presente estudo objetivou avaliar o impacto da população de plantas sobre algumas variáveis morfológicas e na produtividade final de grãos de dois híbridos de milho pipoca (8203 e 4512). Foram estudadas as populações de 60.000, 65.000, 70.000, 75.000 e 80.000 plantas por ha-1 no espaçamento de 0,45cm entre linha de semeadura. O experimento foi conduzido no ano agrícola 2018/2019, na área experimental do Departamento de Ciências Agronômicas e Ambientais da Universidade Federal de Santa Maria, campus Frederico Westphalen ­ RS, em um delineamento de blocos casualizados em um esquema fatorial (2x5). O diâmetro de colmo, comprimento da espiga e peso de mil sementes diminuíram à medida que se aumentou a população de plantas. Para ambos os híbridos, e para a maioria das variáveis analisadas as densidades populacionais não interferiram de forma significativa na produtividade final de grãos do milho pipoca. Entretanto quando se trabalha a média das populações se observa uma superioridade do híbrido 8203 para as variáveis, altura de planta, altura de inserção da espiga, prolificidade, empalhamento, diâmetro de espiga, número de grãos por espiga, peso de mil sementes e produtividade final de grãos.(AU)

When it comes to popcorn, Brazil is the second largest producer, but still needing imports to supply the country's domestic demand, given the importance of studies that better explore agronomic recommendations for popcorn culture. Thus, the aim of the present study was to evaluate the impact of the plant population on some morphological variables and the final consumption of two hybrid popcorn kernels (8203 and 4512). The populations of 60.000, 65.000, 70.000, 75.000 and 80.000 plants per ha-1 were studied in the 0.45cm spacing between sowing lines. The experiment was carried out in the agricultural year 2018/2019, in the experimental area of the Department of Agricultural and Environmental Sciences of the Federal University of Santa Maria, Frederico Westphalen campus - RS, in a randomized block design in a factorial scheme (2x5). The stem diameter, ear length and weight of a thousand seeds decreased as the plant population increased. For both hybrids, and for most of the variables analyzed, population densities did not significantly affect the final grain yield of popcorn. However, when working with the average population, a superiority of the 8203 hybrid is observed for the variables, plant height, height of ear insertion, prolificacy, stuffing, ear diameter, number of grains per ear, weight of a thousand seeds and final productivity of grain.(AU)

En lo que respecta a las palomitas de maíz, Brasil es el segundo mayor productor, pero aún necesita importaciones para satisfacer la demanda interna del país, dada la importancia de los estudios que exploran mejor las recomendaciones agronómicas para el cultivo de palomitas de maíz. Por lo tanto, el objetivo del presente estudio fue evaluar el impacto de la población de plantas en algunas variables morfológicas y en el rendimiento final de grano de dos híbridos de palomitas de maíz (8203 y 4512). Se estudiaron las poblaciones de 60.000, 65.000, 70.000, 75.000 y 80.000 plantas por ha-1 en el espacio de 0.45cm entre líneas de siembra. El experimento se realizó en el año agrícola 2018/2019, en el área experimental del Departamento de Ciencias Agronómicas y Ambientales de la Universidad Federal de Santa María, campus Frederico Westphalen - RS, en un diseño de bloques al azar en un esquema factorial (2x5). El diámetro del tallo, la longitud de la mazorca y el peso de mil semillas disminuyeron à medida que aumentó la población de plantas. Para ambos híbridos, y para la mayoría de las variables analizadas, las densidades de población no afectaron significativamente el rendimiento final de grano de las palomitas de maíz. Sin embargo, cuando se trabaja con la población promedio, se observa una superioridad del híbrido 8203 para las variables, altura de la planta, altura de inserción de la mazorca, prolificidad, relleno, diámetro de la mazorca, número de granos por mazorca, peso de mil semillas y productividad final de grano.(AU)

Screening of interacting proteins of idiopathic gonadotropin-releasing hormone deficiency pathogenic gene RNF216 / 中华医学遗传学杂志

OBJECTIVE@#To screen proteins interacting with ring finger protein 216(RNF216) through yeast two hybrid experiment, and further clarify the role of RNF216 in the pathogenesis of gonadotropin-releasing hormone deficiency.@*METHODS@#A recombinant expression vector pGBKT7-RNF216 was constructed and transformed into yeast Y2HGold, which was hybridized with a human cDNA library in order to screen proteins interacting with RNF216. The interaction was verified in yeast Y2HGold.@*RESULTS@#A recombinant expression vector pGBKT7-RNF216 was successfully constructed and expressed in yeast Y2HGold. Filamin B (FLNB) was identified by yeast two hybrid experiment, and their interaction was verified in yeast Y2HGold.@*CONCLUSION@#An interaction between FLNB and RNF216 was identified through yeast two hybrid experiment. RNF216 may affect the proliferation and migration of GnRH neurons by regulating FLNB or FLNB/FLNA heterodimers.

Produtividade de híbridos de milho pipoca submetidos a diferentes populações na semeadura / Productivity of popcorn hybrids submitted to different populations in seeding / Productividad de híbridos de maíz para palomitas sometidos a diferentes poblaciones en la siembra

Quando se trata de milho pipoca o Brasil é o segundo maior produtor. A carência de recomendações agronômicas específicas para a cultura tem sido um limitante no avanço sobre as áreas de cultivo, dessa maneira se observa a necessidade de realizar estudos voltados ao melhor manejo para a cultura do milho pipoca. Assim, o presente estudo objetivou avaliar o impacto da população de plantas sobre algumas variáveis morfológicas e na produtividade final de grãos de dois híbridos de milho pipoca (8203 e 4512). Foram estudadas as populações de 60.000, 65.000, 70.000, 75.000 e 80.000 plantas por ha-1 no espaçamento de 0,45cm entre linha de semeadura. O experimento foi conduzido no ano agrícola 2018/2019, na área experimental do Departamento de Ciências Agronômicas e Ambientais da Universidade Federal de Santa Maria, campus Frederico Westphalen ­ RS, em um delineamento de blocos casualizados em um esquema fatorial (2x5). O diâmetro de colmo, comprimento da espiga e peso de mil sementes diminuíram à medida que se aumentou a população de plantas. Para ambos os híbridos, e para a maioria das variáveis analisadas, as densidades populacionais não interferiram de forma significativa na produtividade final de grãos do milho pipoca. Entretanto quando se trabalha a média das populações se observa uma superioridade do híbrido 8203 para as variáveis, altura de planta, altura de inserção da espiga, prolificidade, empalhamento, diâmetro de espiga, número de grãos por espiga, peso de mil sementes e produtividade final de grãos.(AU)

When it comes to popcorn, Brazil is the second largest producer of this type of maze. The lack of specific agronomic recommendations for the crop has been a limiting factor in the advance on the cultivation areas. Therefore, the need to carry out studies aimed at the best management for the culture of popcorn is essential. Thus, this study aimed at evaluating the impact of the plant population on some morphological variables and on the final grain yield of two popcorn hybrids (8203 and 4512). The populations of 60,000; 65,000; 70,000; 75,000; and 80,000 plants per ha-1 were studied in the 0.45-cm spacing between sowing lines. The experiment was carried out in the 2018/2019 agricultural year in the experimental area of the Department of Agricultural and Environmental Sciences at the Federal University of Santa Maria, in the Frederico Westphalen campus, state of Rio Grande do Sul, in Brazil, in a randomized block design in a (2x5) factorial scheme. The stem diameter, ear length and weight of a thousand seeds decreased as the plant population increased. For both hybrids, and for most of the variables analyzed, population densities did not significantly affect the final grain yield of popcorn. However, when working with the average population, a superiority of the 8203 hybrid is observed for the variables plant height, height of ear insertion, prolificacy, stuffing, ear diameter, number of grains per ear, weight of a thousand seeds, and final grain productivity.(AU)

Cuando se trata de maíz para palomitas, Brasil es el segundo mayor productor. La falta de recomendaciones agronómicas específicas para el cultivo ha sido un factor limitante en el avance de las áreas de cultivo, por lo que se observa la necesidad de realizar estudios encaminados al mejor manejo para el cultivo de maíz para palomitas. Así, el presente estudio tuvo como objetivo evaluar el impacto de la población de plantas sobre algunas variables morfológicas y sobre el rendimiento final de grano de dos híbridos de maíz para palomitas (8203 y 4512). Se estudiaron las poblaciones de 60.000, 65.000, 70.000, 75.000 y 80.000 plantas por ha-1 en el espacio de 0.45cm entre líneas de siembra. El experimento se realizó en el año agrícola 2018/2019, en el área experimental del Departamento de Ciencias Agronómicas y Ambientales de la Universidad Federal de Santa María, campus Frederico Westphalen - RS, en un diseño de bloques al azar en un esquema factorial (2x5). El diámetro del tallo, la longitud de la mazorca y el peso de mil semillas disminuyeron a medida que aumentó la población de plantas. Para ambos híbridos, y para la mayoría de las variables analizadas, las densidades de población no afectaron significativamente el rendimiento final de granos del maíz para palomitas. Sin embargo, cuando se trabaja con la población promedio, se observa una superioridad del híbrido 8203 para las variables, altura de la planta, altura de inserción de la mazorca, prolificidad, chalas, diámetro de la mazorca, número de granos por mazorca, peso de mil semillas y productividad final de granos.(AU)

Physical and Functional Interaction between 5-HT₆ Receptor and Nova-1

5-HT₆ receptor (5-HT₆R) is implicated in cognitive dysfunction, mood disorder, psychosis, and eating disorders. However, despite its significant role in regulating the brain functions, regulation of 5-HT₆R at the molecular level is poorly understood. Here, using yeast two-hybrid assay, we found that human 5-HT₆R directly binds to neuro-oncological ventral antigen 1 (Nova-1), a brain-enriched splicing regulator. The interaction between 5-HT₆R and Nova-1 was confirmed using GST pull-down and co-immunoprecipitation assays in cell lines and rat brain. The splicing activity of Nova-1 was decreased upon overexpression of 5-HT₆R, which was examined by detecting the spliced intermediates of gonadotropin-releasing hormone (GnRH), a known pre-mRNA target of Nova-1, using RT-PCR. In addition, overexpression of 5-HT₆R induced the translocation of Nova-1 from the nucleus to cytoplasm, resulting in the reduced splicing activity of Nova-1. In contrast, overexpression of Nova-1 reduced the activity and the total protein levels of 5-HT₆R. Taken together, these results indicate that when the expression levels of 5-HT₆R or Nova-1 protein are not properly regulated, it may also deteriorate the function of the other.

Identificación de estructuras de infección de Phytophthora palmivora en hojas de clones de palma de aceite Elaeis guinennsis Jacq / Identification of infection structures of Phytophthora palmivora in leaves of oil palm clones Elaeis guineensis Jacq

Este trabajo investigó el modelo de interacción planta-patógeno mediante discos de hojas de clones de palma de aceite inoculados en condiciones ex situ con un aislamiento de Phytophthora palmivora. Las inoculaciones se realizaron en condiciones controladas en cámara de crecimiento. En total, seis diferentes ortets fueron evaluados en seis tiempos de infección (2, 4, 6, 12, 24 y 48 horas postinfección, o hpi). Se determinó la presencia de estructuras de patogenicidad de P. palmivora como quiste, apresorio y tubos germinativos. Los quistes fueron identificados principalmente a las 2, 4 y 6 hpi. A partir de las 48 hpi no hubo presencia de zoosporas enquistadas para ningún ortet evaluado. En cuanto a los apresorios, estos se empezaron a desarrollar a las 4 horas de realizada la inoculación (siendo las 12 y 24 hpi los tiempos de mayor registro de estas estructuras). 121 Identificación de estructuras de infección de Phytophthora palmivora en hojas de clones de palma de aceite (Elaeis guineensis Jacq.) Méndez, K. et al. Introducción La Pudrición del cogollo (pc) es una de las principales enfermedades que afecta al cultivo de palma de aceite, destruyendo plantaciones desde 1964. En Colombia la enfermedad se presenta en las cuatro zonas palmeras y ha alcanzado proporciones epidémicas (Sarria et al., 2013a, 2016). A finales de 2004 en la Zona Suroccidental se registró un incremento en el número de casos, con un crecimiento exponencial de esta enfermedad, y a partir de 2007 se vieron afectadas más de 30.000 ha de cultivos de palma de aceite. En la Zona Central un comportamiento similar de la enfermedad fue registrado para la región de Puerto Wilches (Norte de Santander) entre 2009 y 2013, periodo en el cual se perdieron más de 40.000 ha como consecuencia de una epidemia de pc (Sanz, 2016). Por su parte, los productores de la Zona Norte actualmente hacen frente a la amenaza de una expansión epidémica de la pc con síntoma de hoja clorótica, la cual da cuenta de un estado avanzado de la enfermedad. Una amplia revisión del impacto de esta enfermedad desde sus inicios se encuentra en Benítez & García (2015), Sundram & Intan-Nur (2017) y Torres et al. (2016). La enfermedad de la Pudrición del cogollo es causada por el oomiceto hemibiótrofo P. palmivora (Sarria et al., 2008, 2013). Desde su identificación como agente causal de la pc en 2008, el Centro de Investigación en Palma de Aceite (Cenipalma) ha liderado diferentes investigaciones que han dado como resultado el desarrollo de estrategias de manejo integrado del cultivo de palma, así como la descripción del patógeno por medio de inoculación en condiciones in vitro en foliolos inmaduros de palma (Martínez et al., 2013, 2014a), entre otras. En cuanto a la presencia de esta enfermedad en otros cultivos, Mohamed-Azni et al. (2017) emplearon la técnica de foliolo inmaduro, logrando infectar foliolos de palma de aceite con P. palmivora aislada proveniente de cultivos de cacao y durián. Teniendo en cuenta que actualmente no se conocen fuentes probadas de resistencia de cultivares de Elaeis guineensis a la pc, y que un ciclo de mejoramiento genético de palma puede durar más de 25 años Por su parte, los tubos germinativos se encontraron a partir de las 48 hpi únicamente. Finalmente, se pudo establecer que el patógeno logra colonizar tejidos de foliolo no lignificados de clones de palma. Además, se encontró una relación entre el número de estructuras del patógeno con el comportamiento del cultivar de la palma donor (ramet)

Plant-pathogen model interaction was studied using leaf disks of oil palm clones inoculated ex situ with a Phytophthora palmivora isolate. The inoculation process was performed under growth chamber conditions. Six ortets were evaluated at six post-inoculation times (2, 4, 6, 12, 24 y 48 hours post infection, or hpi). Pathogen's infection structures as cyst, apressorium and germinative tubes were found. Cysts were identified mainly at 2, 4 and 6 hpi. After 48 hpi there were not any cysts for the evaluated ortets. The apresoria started to develop at 4 hpi, with the highest presence of these structures at 12 and 24 hpi. Germinative tubes were found only after 48 hpi. Therefore, it was established that the pathogen can colonize no-lignified tissue of oil palm clones. Finally, we found a relation between the pathogen's structures number and the response of the ortet related to its susceptibility and resistance response. Thus, it was found that the susceptible cultivar showed the highest number of germinative tubes

Construction of yeast two-hybrid library of Salvia miltiorrhiza and screening of SmJAZ8 interaction protein / 中国中药杂志

The study is aimed to construct high quality Salvia miltiorrhiza cDNA library and obtain the SmJAZ8 gene of S. miltiorrhiza by yeast two-hybrid system. In this study， full-length cDNA was synthesized from roots， stems， leaves， flowers and hairy roots of S. miltiorrhiza. The full-length cDNA library was synthesized by SMART method and constructed with DSN homogenization technique. The results showed that the library capacity was 1.45×10⁶， the recombination rate was 100%， and the average size of the insert was 500-2 000 bp. The recombinant vector of pDEST-pGADT7-SmJAZ8 was constructed and transformed into Y2HGold strain. The interaction protein was screened by yeast two-hybrid system. The DnaJ protein and UBQ protein were screened by yeast two-hybrid system. This study has successfully constructed a full-length cDNA library of S. miltiorrhiza， and laid the foundation for the follow-up study on functional gene screening and gene function of S. miltiorrhiza.

The modified bacterial two-hybrid system / 生物工程学报

Bacterial two-hybrid system is a newly developed method for studying protein-protein interactions. However, in our studies of the interaction of regulatory proteins in Streptomyces, it was found that the bacterial two-hybrid system is not sensitive enough by the blue-and-white selection on X-gal plate. To overcome this drawback, the reason of false positive clone was firstly determined, which was the disturbance of other direct or indirect regulation on lacZ promoter. Then the disturbance was diluted by introducing multicopy lacZ promoter, which drive another reporter gene gfp. By such design, the sensitivity of the modified bacterial two-hybrid system was significantly inproved and the two different reporters also help to decrease the rate of the false positive clones. Further the evaluation of the modifiedd bacterial two-hybrid system indicated that the sensitivity was significantly improved.

Identification of proteins interacting with the circadian clock protein PER1 in tumors using bacterial two-hybrid system technique / 中华医学遗传学杂志

<p><b>OBJECTIVE</b>To identify protein-protein interaction partners of PER1 (period circadian protein homolog 1), key component of the molecular oscillation system of the circadian rhythm in tumors using bacterial two-hybrid system technique.</p><p><b>METHODS</b>Human cervical carcinoma cell Hela library was adopted. Recombinant bait plasmid pBT-PER1 and pTRG cDNA plasmid library were cotransformed into the two-hybrid system reporter strain cultured in a special selective medium. Target clones were screened. After isolating the positive clones, the target clones were sequenced and analyzed.</p><p><b>RESULTS</b>Fourteen protein coding genes were identified, 4 of which were found to contain whole coding regions of genes, which included optic atrophy 3 protein (OPA3) associated with mitochondrial dynamics and homo sapiens cutA divalent cation tolerance homolog of E. coli (CUTA) associated with copper metabolism. There were also cellular events related proteins and proteins which are involved in biochemical reaction and signal transduction-related proteins.</p><p><b>CONCLUSION</b>Identification of potential interacting proteins with PER1 in tumors may provide us new insights into the functions of the circadian clock protein PER1 during tumorigenesis.</p>

Construction and Identification of the Bait Vector Containing Duck Circovirus Cap Gene for the Yeast Two-hybrid System / 病毒学报

To construct a bait expression vector containing the duck circovirus Cap gene for use in the yeast two-hybrid system, the whole cap codon-optimized gene was inserted into pGBKT7 vector and confirmed by PCR, restriction enzyme digestion, and sequence analysis. After transformation into a Y2HGold yeast strain, the expression of Cap protein was analyzed by Western blotting. Toxicity and self-activation of the bait protein were detected using different dropout minimal base. PCR reaction, restriction enzyme digestion, and sequencing analyses indicated that the duck circovirus Cap gene was correctly inserted into pG- BKT7. Western blotting showed that the whole Cap protein was expressed. The recombinant bait protein had no toxicity and self-activation. Therefore, the bait vector with the Cap gene was constructed successfully, providing a foundation for future screening for interacting proteins in the yeast two-hybrid system.

Identification of Host Factors Interacting with the Movement Protein of Apple Chlorotic Leaf Spot Virus by Yeast Two-Hybrid System / 病毒学报

In order to identify host factors which interact with the movement protein (MP) of Apple chlorotic leaf spot virus (ACLSV), ACLSV MP was cloned into the bait vector pGBKT7 and used to screen a cDNA library of Malus sylvestris cv. R12740-7A, which had previously been constructed by yeast two-hybrid sequencing transformation. The protein functions of the identified host factors were determined according to their gene annotations in GenBank. The result showed that the bait plasmid pGBKT7-MP showed no virulence or self-activating effect on yeast strain Y2H Gold. Sixty-nine interactor proteins were identified, which were divided into the following 10 classes according to their described functions: hydrolases; pathogenesis-related proteins; DNA binding proteins; phosphatases; ligases; proteins with catalytic activity; phenylalanine ammonialyases; peroxidases; NAD binding proteins; and proteins of unknown function. Bioinformatic analysis of gene homology suggested that phosphatases, pathogenesis-related proteins and glyceraldehyde-3-phosphate dehydrogenase A may play an important role in the interaction between virus and host. This study may provide a theoretical basis for the further study of viral pathogenesis and virus-host interaction mechanisms.

Occurrence of Lutzomyia longipalpis (Phlebotominae) and canine visceral leishmaniasis in a rural area of Ilha Solteira, SP, Brazil / Ocorrência de Lutzomyia longipalpis (Phlebotominae) e leishmaniose visceral canina em uma área rural de Ilha Solteira, SP, Brasil

This study aimed to investigate the occurrence of Lutzomyia longipalpis and also the canine visceral leishmaniasis (CVL) in a rural area of Ilha Solteira, state of São Paulo. Blood samples were collected from 32 dogs from different rural properties (small farms) and were analyzed by ELISA and the indirect immunofluorescence antibody test (IFAT) in order to diagnose CVL. From these serological tests, 31.25% of the dogs were positive for CVL and these were distributed in 66.7% (8/12) of the rural properties, which were positive for L. longipalpis. CDC (Center for Disease Control and Prevention) light traps were installed in 12 properties (one per property) and insects were caught on three consecutive days per month for one year. L. longipalpis was present on 100% of the rural properties visited, at least once during the twelve-month interval, totaling 64 males and 25 females. The insects were more numerous after the peak of the rain, but the association between prevalence of peridomestic vectors and the climatic data (precipitation, relative air humidity and temperature) and the occurrences of CVL among dogs on each rural property were not statistical significant (p <0.05). However, the occurrence of CVL cases in dogs and the presence of L. longipalpis indicate that more attention is necessairy for the control of this disease in the rural area studied.

O objetivo desse trabalho foi o estudo da prevalência de Lutzomyia longipalpis e da leishmaniose visceral canina (LVC) em uma área rural do município de Ilha Solteira do estado de São Paulo. Amostras de sangue foram coletadas de 32 cães provenientes de pequenas propriedades rurais e analisadas por meio dos métodos sorológicos ELISA (imunoensaio enzimático indireto) e RIFI (reação de imunofluorescência indireta) para o diagnóstico da LVC. Pelos exames sorológicos, dos 32 cães avaliados, 31,25% foram diagnosticados positivos para LVC, os quais estavam diostribuídos em 66,67% (8/12) das propriedades positivas para Lutzomyia longipalpis. Armadilhas luminosas do tipo CDC (Center for Disease Control and Prevention) foram instaladas em 12 propriedades, sendo uma por propriedade, e as coletas dos insetos foram realizadas três dias consecutivos a cada mês, durante um ano. O inseto L. longipalpis foi encontrado em 100% das propriedades visitadas, pelo menos uma vez no ano, totalizando 65 machos e 25 fêmeas. A maior quantidade de insetos foi observada principalmente após a ocorrência dos maiores picos de precipitação pluvial, mas a associação entre a prevalência dos vetores peridomiciliares e os dados climáticos (precipitação, umidade relativa do ar e temperatura) assim como a ocorrência da CVL em cães em cada propriedade não foi estatisticamente significante (p<0.05). No entanto, alerta-se que pela presença dos casos de LVC nos cães amostrados e também de L. longipalpis, maior atenção deve ser dada durante as investigações epidemiológicas para o controle dessa doença nessa área rural estudada.

New ligands of the Cellular Retinoic Acid-Binding Protein 2 (CRABP2) suggest a role for this protein in chromatin remodeling

Retinoic acid (RA) regulates the transcription of a series of genes involved in cell proliferation, differentiation and apoptosis by binding to the RA Receptor (RAR) and Retinoid X Receptor (RXR) heterodimers. The cellular retinoic acid-binding protein 2 (CRABP2) is involved in the transport of RA from the cytosol to specific RA receptors in the nucleus, acting as a coactivator of nuclear retinoid receptors. In order to have a better understanding of the role of CRABP2 in RA signaling, we used the yeast two-hybrid system as a tool for the identification of physical protein-protein interactions. Twenty-three putative CRABP2-interacting proteins were identified by screening in the presence of RA, five of which are related to transcription regulation or, more specifically, to the process of chromatin remodeling: t-complex 1 (TCP1); H3 histone, family 3A (H3F3A); H3 histone, family 3B (H3F3B); β-tubulin (TUBB) and SR-related CTD-associated factor 1 (SCAF1). These results suggest a more direct role for CRABP2 in chromatin remodeling and may be a starting point for the elucidation of the fine-tuning control of transcription by RA receptors...

Determinants of emergency contraception non-use among women in unplanned or ambivalent pregnancies / Determinantes del no uso de la anticoncepción de emergencia entre mujeres con embarazo no planeado u ambivalente / Determinantes do não uso da anticoncepção de emergência entre mulheres com gravidez não planejada ou ambivalente

Objective To analyze the determinants of emergency contraception non-use among women in unplanned and ambivalent pregnancies. Method Cross-sectional study with a probabilistic sample of 366 pregnant women from 12 primary health care units in the city of São Paulo, Brazil. A multinomial logistic regression was performed, comparing three groups: women who used emergency contraception to prevent ongoing pregnancies (reference); women who made no use of emergency contraception, but used other contraceptive methods; and women who made no use of any contraceptive methods at all. Results Cohabitation with a partner was the common determinant of emergency contraception non-use. No pregnancy risk awareness, ambivalent pregnancies and no previous use of emergency contraception also contributed to emergency contraception non-use. Conclusion Apart from what is pointed out in the literature, knowledge of emergency contraception and the fertile period were not associated to its use. .

Objetivo Analizar los determinantes del no uso de la anticoncepción de emergencia entre las mujeres con embarazo no planeado o ambivalente. Método Estudio transversal en una muestra probabilística de 366 mujeres embarazadas de 12 Unidades Básicas de Salud de São Paulo. Mediante regresión logística multinomial, se comparó tres grupos de mujeres: aquellas que usaron la anticoncepción de emergencia para prevenir el embarazo en curso (referencia), aquellas que usaron algún método anticonceptivo, pero no la anticoncepción de emergência; y aquellas que no usaron ningún método. Resultados Los hallazgos mostraron que vivir com la pareja fue el determinante común del no uso de la anticoncepción de emergencia. No tener conciencia del riesgo de embarazo, estar en un embarazo ambivalente y nunca tener utilizado la anticoncepción de emergencia también fueron associados con su no uso para prevenir el embarazo en curso. Conclusión Contrariamente a lo que reporta la literatura, el conocimiento de la anticoncepción de emergencia y el período fértil no mostró asociación con el no uso. .

Objetivo Analisar os determinantes do não uso da anticoncepção de emergência entre mulheres com gravidez não planejada ou ambivalente. Método Estudo transversal com amostra probabilística de 366 gestantes de 12 Unidades Básicas de Saúde da cidade de São Paulo. Por meio de regressão logística multinomial, compararam-se três grupos de mulheres: as que usaram anticoncepção de emergência para prevenir a gravidez em curso (referência); as que usaram algum método contraceptivo, mas não anticoncepção de emergência; e as que não usaram nenhum método. Resultados Os achados mostraram que morar com o parceiro foi o determinante comum do não uso da anticoncepção de emergência. Não ter consciência do risco de engravidar, estar em uma gravidez ambivalente e nunca ter usado anticoncepção de emergência também foram associados ao seu não uso para prevenir a gravidez em curso. Conclusão Diferentemente do que relata a literatura, o conhecimento sobre anticoncepção de emergência e sobre o período fértil não mostrou qualquer associação ao não uso. .

Development of a novel screening assay for inhibitors targeting HIF-1alpha and P300 interaction / 药学学报

Hypoxia is a general characteristic of most solid malignancies and intimately related to cancer progression. Homeostatic response to hypoxia is primarily mediated by hypoxia inducible factor-1alpha (HIF-1alpha) that elicits transcriptional activity through recruitment P300 coactivator. Targeting the interaction of HIF- alpha and P300 would thus constitute a novel approach for cancer treatment by suppressing tumor angiogenesis and metastasis. Here, a screening assay was developed for inhibitors targeting the interaction between HIF-1alpha and P300. The nucleotide sequence of human HIF-1alpha and P300 were cloned into pBIND and pACT vectors, named pBIND-HIF1alpha and pACT-P300. The interaction of HIF-1alpha and P300 was identified in HEK293 cell using mammalian two-hybrid system. And compound chetomin decreased their interaction in this mammalian two-hybrid system. We further verified HIF-1 inhibition effect of chetomin in U251-HRE cells. Therefore, we established a screening assay combined HIF-1alpha and P300 mammalian two-hybrid system and U251-HRE reporter assay for HIF-1 selective inhibitors.

Research methods in protein-protein and protein-nucleic acid interactions and application in the study of human enterovirus A71 / 病毒学报

Methods for analyses of protein-protein interactions include: yeast two hybrid (Y2H), phage dis- play (PD), co-immunoprecipitation (Co-IP), glutathione S-transferase pull-down (GST pull-down), cellular co-localization, far-western blotting, virus overlay protein binding assay (VOPBA), surface plasmon resonance (SPR), and fluorescence resonance energy transfer (FRET). Technologies for the detection of protein-nucleic acid interactions include: yeast one hybrid (Y1H), chromatin immunoprecipitation (ChIP), electrophoretic mobility shift assay (EMSA), Southwestern blotting, reporter gene, Co-IP, GST pull-down, and PD. These methods are often used in the study of the human enterovirus A71 (EV-A71) by our research team. Reviews in the Chinese literature in this field are lacking, so we reviewed applications of these methods in the study of EV-A71. This review may impart important knowledge in the research of other viruses with regard to protein-protein and protein-nucleic acid interactions.

Location of the binding position for HBV X protein functional interaction with cytochrome C oxidase III / 中华肝脏病杂志

<p><b>OBJECTIVE</b>To identify the binding site position of the hepatitis B virus (HBV) X protein (HBx) functional interaction with the cytochrome C oxidase subunit III (COX III, a key regulator of mitochondrial function) by using a yeast two-hybrid system.</p><p><b>METHODS</b>Two fragments of HBx mutants (X1 1-72aa and X2 1-117aa) were amplified by PCR and inserted into the bait plasmid pAS2-1.The resultant mutant plasmids were transfected into yeast cells using the lithium acetate-method.PCR and gene sequencing were used to confirm that the mutant fragments were expressed properly in yeast cells.Western blotting was used to verify that the mutant proteins were translated accurately in the yeast cells.Filter assay was used to exclude autoactivated mutants.Hybridization in solid medium and beta-gal activity detection were used to determine the precise position of the binding site for HBx and COX III interaction.</p><p><b>RESULTS</b>The two mutant plasmids containing HBx 1-72aa and 1-117aa respectively were successfully constructed and the mutants were both properly expressed and translated in yeast cells; no autoactivated mutants were detected throughout the experimental process.The binding site of HBx and COX III was found to be encompass the amino acids 72 through 117 of HBx.</p><p><b>CONCLUSION</b>Amino acids 72 through 117 of HBx are the key domain of the HBx functional interaction With COX III; this domain may represent a useful target for molecular-based therapies to treat HBV-related diseases.</p>

Protein Phosphatase 2C of Toxoplasma Gondii Interacts with Human SSRP1 and Negatively Regulates Cell Apoptosis / 生物医学与环境科学（英文）

<p><b>OBJECTIVE</b>The protozoan Toxoplasma gondii expresses large amounts of a 37 kDa Type 2C serine-threonine phosphatase, the so-called TgPP2C which has been suggested to contribute to parasite growth regulation. Ectopic expression in mammalian cells also indicated that the enzyme could regulate growth and survival. In this study, we aimed to investigate the interaction of TgPP2C with human SSRP1 (structure-specific recognition protein 1) and the effects of TgPP2C on cell viability.</p><p><b>METHODS</b>The yeast two hybrid system, His-tag pull-down and co-immunoprecipitation assays were used to confirm the interaction of TgPP2C with SSRP1 and determine the binding domain on SSRP1. The evaluation of cell apoptosis was performed using cleaved caspase-3 antibody and Annexin-V/PI kit combined with flow cytometry.</p><p><b>RESULTS</b>We identified human SSRP1 as an interacting partner of TgPP2C. The C-terminal region of SSRP1 including the amino acids 471 to 538 was specifically mapped as the region responsible for interaction with TgPP2C. The overexpression of TgPP2C down-regulated cell apoptosis and negatively regulated apoptosis induced by DRB, casein kinase II (CKII) inhibitor, through enhanced interaction with SSRP1.</p><p><b>CONCLUSION</b>TgPP2C may be a parasitic factor capable of promoting cell survival through interaction with the host protein SSRP1, thereby creating a favorable environment for parasite growth.</p>

Establishment of a hMSH2/hMSH6 protein interaction system and functional evaluation of hMSH2 gene missense mutations / 中华医学遗传学杂志

<p><b>OBJECTIVE</b>To construct a hMSH2/hMSH6 protein interaction system, and to use it for evaluating missense mutations detected in hMSH2 gene.</p><p><b>METHODS</b>Recombinant plasmids pGADT7-hMSH2, pGBKT7-hMSH6 and 7 recombinant pGBKT7 plasmids with different hMSH6 domains were constructed through genetic engineering. Subsequently, site-directed mutagenesis was used to construct 10 mutant pGADT7-hMSH2 plasmids, which were transformed into yeast AH109. The growth of transformants was observed on a histidine-deficient culture.</p><p><b>RESULTS</b>Both hMSH6 MutSII-V and MutSIII-V could interact with hMSH2 in yeast AH109. Yeast two-hybrid transformants pGADT7-hMSH2/pGBKT7-hMSH6 MutSII-V were used to construct a hMSH2/hMSH6 protein interaction system. Compared with wild-type hMSH2, yeast two-hybrid transformants c.505A>G, c.1168C>T, c.1255C>A, c.1261C>A could grow normally, c.1223A>G, c.1886A>G, c.2108C>A and c.2516A>G grew slowly, c.518T>G and c.1664 delA could not grow in a histidine-deficient medium in yeast AH109.</p><p><b>CONCLUSION</b>A hMSH2/hMSH6 protein interaction system has been constructed with yeast two-hybrid system, which has been used for functional evaluation of hMSH2 gene missense mutations. c.518T>G is a pathological mutation. c.1223A>G, c.1886A>G, c.2108C>A, c.2516A>G may in part affect the hMSH2 function. And c.505A>G, c.1168C>T, c.1255C>A, c.1261C>A were innocuous variants.</p>

Screening of hepatocyte proteins interacting with hepatitis B virus X protein using CytoTrap yeast two-hybrid technique / 南方医科大学学报

<p><b>OBJECTIVE</b>To screen the hepatocyte proteins that interact with hepatitis B virus X protein (HBx).</p><p><b>METHODS</b>The recombinant plasmid pSos-HBx was constructed by inserting Sos-HBx fragment into the bait vector, and after sequence verification the plasmid was transformed into competent yeast cells. The expression and self-activation of Sos-HBx protein was detected in the yeast cells. The hepatocyte proteins interacting with the bait protein was screened with CytoTrap yeast two-hybrid technique.</p><p><b>RESULTS</b>The reconstructed plasmid harboring HBx gene expressed Sos-HBx protein in the yeast cells without self-activation of the protein. CytoTrap yeast two-hybrid system identified 6 hepatocyte proteins that interacted with HBx, including fibronectin 1, translationally controlled tumor protein, IQ motif and WD repeats 1, follistatin, orosomucoid 1, and disulfide isomerase family A member 3.</p><p><b>CONCLUSION</b>Six HBx-binding hepatocyte proteins have been identified using the CytoTrap yeast two-hybrid system, which provides clues for further investigation of the role of HBx protein in hepatitis and liver cancer.</p>

Functional Interaction of HIF-1 and NF-kappaB Increasing the Transcriptional Activation of TNF-alpha Gene in Monocytes / 대한수혈학회지

BACKGROUND: Tumor necrosis factor alpha (TNF-alpha) is a pleiotropic cytokine fulfilling a broad variety of immunoregulatory functions. Monocytes and macrophages play a pivotal role in inflammation and immune regulation. NF-kappaB and HIF-1 are known to increase expression of the TNF-alpha gene in a separate way. METHODS: Human monocytic leukemia, U937 cells, were transfected using the standard electroporation method for intracellular expression of NF-kappaB and HIF-1. We performed analysis using the mammalian two-hybrid assay and co-immunoprecipitation assay for detection of protein interaction of both proteins. In addition, chromatin immunoprecipitation analysis was performed for examination of NF-kappaB and HIF-1 binding on the TNF-alpha gene promoter. RESULTS: Here we show that NF-kappaB and HIF-1 cooperatively induced an increase in expression of the TNF-alpha gene dependent on promoter activity by the direct protein interaction of these two transcription factors. Hypoxia signaling induced marked enhancement of the transactivation of TNF-alpha promoter by HIF-1 and NF-kappaB. A tandem NF-kappaB/HIF-1 binding site was identified within the TNF-alpha promoter, which acted as a strong enhancer element. Physical association of the Rel domain of NF-kappaB and the N-TD domain of HIF-1 was required. Hypoxia treatment also resulted in a significant increase in the protein interaction of NF-kappaB and HIF-1 in vivo. Both transcription factors were recruited on the chromatin TNF-alpha promoter dependent on hypoxia stimuli. CONCLUSION: The results of this study indicate that a variety of extracellular signals for activation of TNF-alpha gene expression might converge on the transcriptional regulation through the NF-kappaB/HIF-1 signaling pathway.