Composition and Role of Extracellular Polymers in Methanogenic Granules

Methanobacterium formicicum and Methanosarcina mazeii are two prevalent species isolated from an anaerobic granular consortium grown on a fatty acid mixture. The extracellular polysaccharides (EPS) were extracted from Methanobacterium formicicum and Methanosarcina mazeii and from the methanogenic granules to examine their role in granular development. The EPS made up approximately 20 to 14% of the extracellular polymer extracted from the granules, Methanobacterium formicicum, and Methanosarcina mazeii. The EPS produced by Methanobacterium formicicum was composed mainly of rhamnose, mannose, galactose, glucose, and amino sugars, while that produced by Methanosarcina mazeii contained ribose, galactose, glucose, and glucosamine. The same sugars were also present in the EPS produced by the granules. These results indicate that the two methanogens, especially Methanobacterium formicicum, contributed significantly to the production of the extracellular polymer of the anaerobic granules. Growth temperature, substrates (formate and H(inf2)-CO(inf2)), and the key nutrients (nitrogen and phosphate concentrations) affected polymer production by Methanobacterium formicicum.     

A butenolide, isolated from smoke, can overcome the detrimental effects of extreme temperatures during tomato seed germination

The butenolide, 3-methyl-2H-furo[2, 3-c]pyran-2-one, is an highly active compound isolated from plant-derived smoke. This compound is known to stimulate seed germination in a wide range of plants akin to smoke or aqueous extracts of smoke. The present study attempted to elucidate the role of the butenolide in overcoming detrimental effects of low and high temperatures on tomato seed germination and seedling growth. The germination percentage followed a parabolic curve for temperatures ranging from 10 to 40°C, with 25°C being the optimum for all treatments. Control seeds showed radicle emergence at two extreme temperatures (10 and 40°C) and seedlings failed to develop further, even upon prolonged incubation. By comparison the butenolide-treated seeds grew into phenotypically normal seedlings at these non-optimum temperatures. The smoke–water-treated seeds had an intermediate response as only a fraction of germinated seed developed into normal seedlings. Seedling vigour indices as well as seedling weight were significantly higher (p ≤ 0.05) for butenolide-treated seeds at all temperatures. Furthermore, seedlings developed in the presence of the butenolide had about a 1:1 correspondence between root and shoot length. Butenolide-treated seeds grew better than the control seeds in the temperature shift experiments. A gradual decline in the vigour index values was recorded with an increased duration of incubation at the extreme temperatures. Results of the present study are very important from an horticultural point of view as they indicate the potential use of the butenolide compound in restoring normal seed germination and seedling establishment in tomato below and above optimum temperatures.     

非生物胁迫下植物表观遗传变异的研究进展

植物在整个生命过程中固着生长,不能主动躲避外界不良环境的危害,需要通过自身的防御机制来抵御和适应外界胁迫,而表观遗传修饰在调控植物应对不良环境胁迫中起重要作用。该文从DNA甲基化、组蛋白修饰、染色质重塑和非编码RNA等方面进行了综述,主要阐述了近年来国内外有关非生物胁迫下植物的表观遗传变化,以期为利用表观遗传变异提高植物的抗胁迫能力提供参考。     

Nuclear import and export signals in control of Nrf2

Nrf2 binds to the antioxidant response element and regulates expression and antioxidant induction of a battery of chemopreventive genes. In this study, we have identified nuclear import and export signals of Nrf2 and show that the nuclear import and export of Nrf2 is regulated by antioxidants. We demonstrate that Nrf2 contains a bipartite nuclear localization signal (NLS) and a leucine-rich nuclear export signal, which regulate Nrf2 shuttling in and out of the nucleus. Immunofluorescence and immunoblot analysis revealed that Nrf2 accumulates in the nucleus within 15 min of antioxidant treatment and is exported out of nucleus by 8 h after treatment. Nrf2 mutant lacking the NLS failed to enter the nucleus and displayed diminished expression and induction of the downstream NAD(P)H:quinone oxidoreductase 1 gene. The Nrf2 NLS sequence, when fused to green fluorescence protein, resulted in the nuclear accumulation of green fluorescence protein, indicating that this signal sequence was sufficient to direct nuclear localization of Nrf2. A nuclear export signal (NES) was characterized in the C terminus of Nrf2, the deletion of which caused Nrf2 to accumulate predominantly in the nucleus. The Nrf2 NES was sensitive to leptomycin B and could function as an independent export signal when fused to a heterologous protein. Further studies demonstrate that NES-mediated nuclear export of Nrf2 is required for degradation of Nrf2 in the cytosol. These results led to the conclusion that Nrf2 localization between cytosol and nucleus is controlled by both nuclear import and export of Nrf2, and the overall distribution of Nrf2 is probably the result from a balance between these two processes. Antioxidants change this balance in favor of nuclear accumulation of Nrf2, leading to activation of chemopreventive proteins. Once this is achieved, Nrf2 exits the nucleus for binding to INrf2 and degradation.     

Evaluation of cyclooxygenase 2 derived endogenous prostacyclin in mouse preimplantation embryo development in vitro

Cyclooxygenase (COX) plays an important role in prostaglandin (PG) synthesis and has two isoforms, COX1 and COX2. PGI synthase (PGIS) catalyzes the isomeization of PGH(2) to prostacyclin (PGI(2)). It is reported that COX2 derived PGI2(2) plays a critical role in blastocyst implantation and decidualization and PGI2 mediates its function via PPARdelta receptor. It is also known that cyclooxygenase derived prostaglandins play an important role in mouse blastocyst hatching in vitro. In this study we hypothesized that COX2 derived PGI2 plays an important role in preimplantation embryonic development by increasing the cell number. To examine this hypothesis, 8-cell stage mouse embryos were cultured in the presence of selective inhibitors of COX1 (SC560), COX2 (NS398) and PGIS (U51605) respectively. COX2 and PGIS inhibitor significantly reduced the blastocyst development and presence of PGI2 analogue along with these inhibitors restored the blastocyst development by increasing the total number of embryonic cells. Our immunohistochemical analysis showed that COX1 is expressed at 2-cell, 8-cell, compaction and blastocyst stage whereas COX2 expression starts from eight cell stage embryos. PGIS and PPARdelta expression starts at 2-cell stage of development. Our results suggest that PGI(2) may affect blastomeres number via the so called hypothesis of PPARdelta nuclear receptor in autocrine manner.     

Bayesian-based selection of metabolic objective functions

MOTIVATION: A critical component of in silico analysis of underdetermined metabolic systems is the identification of the appropriate objective function. A common assumption is that the objective of the cell is to maximize growth. This objective function has been shown to be consistent in a few limited experimental cases, but may not be universally appropriate. Here a method is presented to quantitatively determine the most probable objective function. RESULTS: The genome-scale metabolism of Escherichia coli growing on succinate was used as a case-study for analysis. Five different objective functions, including maximization of growth rate, were chosen based on biological plausibility. A combination of flux balance analysis and linear programming was used to simulate cellular metabolism, which was then compared to independent experimental data using a Bayesian objective function discrimination technique. After comparing rates of oxygen uptake and acetate production, minimization of the production rate of redox potential was determined to be the most probable objective function. Given the appropriate reaction network and experimental data, the discrimination technique can be applied to any bacterium to test a variety of different possible objective functions. SUPPLEMENTARY INFORMATION: Additional files, code and a program for carrying out model discrimination are available at http://www.engr.uconn.edu/~srivasta/modisc.html.     

Rapid and Robust PCR-Based All-Recombinant Cloning Methodology

We report here a PCR-based cloning methodology that requires no post-PCR modifications such as restriction digestion and phosphorylation of the amplified DNA. The advantage of the present method is that it yields only recombinant clones thus eliminating the need for screening. Two DNA amplification reactions by PCR are performed wherein the first reaction amplifies the gene of interest from a source template, and the second reaction fuses it with the designed expression vector fragments. These vector fragments carry the essential elements that are required for the fusion product selection. The entire process can be completed in less than 8 hours. Furthermore, ligation of the amplified DNA by a DNA ligase is not required before transformation, although the procedure yields more number of colonies upon transformation if ligation is carried out. As a proof-of-concept, we show the cloning and expression of GFP, adh, and rho genes. Using GFP production as an example, we further demonstrate that the E. coli T7 express strain can directly be used in our methodology for the protein expression immediately after PCR. The expressed protein is without or with 6xHistidine tag at either terminus, depending upon the chosen vector fragments. We believe that our method will find tremendous use in molecular and structural biology.     

Antibody Profiling by Proteome Microarray with Multiplex Isotype Detection Reveals Overlap between Human and Aotus nancymaae Controlled Malaria Infections

The development of vaccines against malaria and serodiagnostic tests for detecting recent exposure requires tools for antigen discovery and suitable animal models. The protein microarray is a high‐throughput, sample sparing technique, with applications in infectious disease research, clinical diagnostics, epidemiology, and vaccine development. We recently demonstrated Qdot‐based indirect immunofluorescence together with portable optical imager ArrayCAM using single isotype detection could replicate data using the conventional laser confocal scanner system. We developed a multiplexing protocol for simultaneous detection of IgG, IgA, and IgM and compared samples from a controlled human malaria infection model with those from controlled malaria infections of Aotus nancymaae, a widely used non‐human primate model of human malaria. IgG profiles showed the highest concordance in number of reactive antigens; thus, of the 139 antigens recognized by human IgG antibody, 111 were also recognized by Aotus monkeys. Interestingly, IgA profiles were largely non‐overlapping. Finally, on the path toward wider deployment of the portable platform, we show excellent correlations between array data obtained in five independent laboratories around the United States using the multiplexing protocol (R²: 0.60–0.92). This study supports the use of this platform for wider deployment, particularly in endemic areas where such a tool will have the greatest impact on global human health.     

若尔盖人-狼冲突现状与管理研究

大中型食肉动物肇事事件导致人类与野生动物关系恶化,给生物多样性保护工作带来巨大的挑战。若尔盖湿地是我国三大湿地之一,湿地、草原分布广泛,生物多样性丰富,畜牧业发达,但近年来狼(Canis lupus)捕杀牲畜的肇事事件时有发生。为了解若尔盖野生狼肇事件的空间分布以及牧民对人-狼冲突管理的看法,本研究于2022年对若尔盖县13个乡镇83个行政村进行走访调查。结果表明：(1)多数受访者(66.0%)认为在过去5年内,若尔盖县野生狼数量有所增加;(2)狼肇事事件具有明显的空间分异性,最严重的是包座乡。包座乡临近山区,该区域牧场面积广阔、牧民饲养牲畜数量多等原因导致该镇发生狼肇事事件较多;(3)对于狼肇事,绝大多数牧民(85.0%)更希望采取经济补偿或者驱赶措施,只有少数牧民(9.4%)希望采取捕杀的措施;(4)影响牧民对狼肇事管理措施的偏好因子中,受教育程度、年龄、民族以及被杀牲畜数量有显著影响。建议加强狼种群监测管理,采取措施减少狼捕杀牲畜,优化补偿机制,缓解当地牧民与狼之间的矛盾。本研究为当前若尔盖县野生动物保护和管理决策提供了依据,对其他地区大型食肉动物与当地居民冲突管理具有借鉴意义。     

利用恒河婴猴模型对肠道病毒71型传播感染特征的生物学分析

本文在前期工作基础上,进一步对肠道病毒71型(EV71)从恒河婴猴的感染个体向其他未感染个体传播的可能性及相关生物学特性做了初步分析.通过喷雾形式经呼吸道感染1~2月龄恒河婴猴(A组);在观察临床症状同时,于感染后第7天,取该组动物粪便处理后,将上清液以喷雾形式经呼吸道感染新的婴猴个体(B组),随后对该次代感染个体进行...