围小丛壳Glomerella cingulata子囊孢子交配繁殖、生物学特性及致病性

炭疽叶枯病（Glomerella leaf spot）是我国苹果上新发现的一种病害。为了解围小丛壳Glomerella cingulata子囊孢子的交配方式、生物学特性和致病性,从安徽砀山、山东牟平等地采集病害样品,经分离培养和纯化获得单孢菌株。在适宜条件下单孢菌株可产生子囊和子囊孢子,经过毛细管破子囊壁后单孢分离,获得12个子囊,每个子囊有8个子囊孢子。其中10个子囊中有4个“正”孢子（+）和4个“负”孢子（-）,2个子囊中只有“负”孢子。子囊孢子单孢菌株培养72h,“正”菌株菌落白色,以营养生长为主;“负”菌株菌落灰白色,直径略小于正菌株,菌丝稀疏,边缘菌丝白色,中部有大量橙色的分生孢子堆。“正”、“负”菌株异宗配合后,可产生大量可育子囊壳;单独的“正”菌株有性生殖产生稀疏丛簇状的可育子囊壳;单个的“负”菌株只能产生分散且不育的子囊壳。“正”、“负”菌株菌落的生长速度没有差异,对温度、营养、光照和pH值的敏感性也没有差异,但“正”、“负”菌株的致病性存在差异。正菌株的有性生殖没有导致rDNA-ITS、β-tubulin基因碱基序列变异。     

Noradrenergic α₁ receptor antagonist treatment attenuates positive subjective effects of cocaine in humans: a randomized trial

Background

Preclinical research implicates dopaminergic and noradrenergic mechanisms in mediating the reinforcing effects of drugs of abuse, including cocaine. The objective of this study was to evaluate the impact of treatment with the noradrenergic α₁ receptor antagonist doxazosin on the positive subjective effects of cocaine.

Methods

Thirteen non-treatment seeking, cocaine-dependent volunteers completed this single-site, randomized, placebo-controlled, within-subjects study. In one study phase volunteers received placebo and in the other they received doxazosin, with the order counterbalanced across participants. Study medication was masked by over-encapsulating doxazosin tablets and matched placebo lactose served as the control. Study medication treatment was initiated at 1 mg doxazosin or equivalent number of placebo capsules PO/day and increased every three days by 1 mg. After receiving 4 mg doxazosin or equivalent number of placebo capsules participants received masked doses of 20 and 40 mg cocaine IV in that order with placebo saline randomly interspersed to maintain the blind.

Results

Doxazosin treatment was well tolerated and doxazosin alone produced minimal changes in heart rate and blood pressure. During treatment with placebo, cocaine produced dose-dependent increases in subjective effect ratings of “high”, “stimulated”, “like cocaine”, “desire cocaine”, “any drug effect”, and “likely to use cocaine if had access” (p<.001). Doxazosin treatment significantly attenuated the effects of 20 mg cocaine on ratings of “stimulated”, “like cocaine”, and “likely to use cocaine if had access” (p<.05). There were trends for doxazosin to reduce ratings of “stimulated”, “desire cocaine”, and “likely to use cocaine if had access” (p<.10).

Conclusions

Medications that block noradrenergic α₁ receptors, such as doxazosin, may be useful as treatments for cocaine dependence, and should be evaluated further.

Trial Registration

Clinicaltrials.gov {"type":"clinical-trial","attrs":{"text":"NCT01062945","term_id":"NCT01062945"}}NCT01062945     

Antigens in culture supernatant of Mycobacterium tuberculosis: epitopes defined by monoclonal and human antibodies.

Antigens of Mycobacterium tuberculosis found in the supernatant of heat-treated cultures were characterized in order to explore whether antigens from this source could be used for the development of a serological test. Culture supernatants and sonicates of 12, 25 and 39 d cultures were analysed by SDS-PAGE. In culture supernatant, major protein bands of 65, 24, and 12 kDa were visible after Coomassie brilliant blue staining. Using murine monoclonal antibodies in Western blots, a pattern of protein bands distinct from that of the corresponding M. tuberculosis sonicates was found in all the culture supernatants. Gel permeation chromatography, in the presence of SDS, was used to separate the major protein bands in the culture supernatant. In ELISA, sera from 20 of 26 patients with tuberculosis reacted with fractions containing mainly 24 kDa or 12 kDa proteins, whereas none of the control sera reacted. In Western blots, each patient serum had its own characteristic banding pattern with culture supernatant, but all the sera from tuberculosis patients and control subjects reacted with protein bands of 65, 61, 58, 30 and 24 kDa. The 12 kDa protein was recognized only by sera from patients with tuberculosis in both Western blots and ELISA. This suggests that different kinds of epitopes on proteins of M. tuberculosis are detected by human antibodies in Western blots and ELISA. We assume that epitopes recognized in Western blots by patients with tuberculosis and control subjects are ubiquitous and are also present on normal commensal bacteria. Epitopes recognized by only some patients with tuberculosis in Western blots may be linear and M. tuberculosis specific. Epitopes recognized by tuberculosis patients but by none of the control subjects in ELISA may be conformation related and M. tuberculosis specific. The major protein bands found in supernatants of heat-treated cultures, 24 and 12 kDa, possess epitopes that may be M. tuberculosis specific and are potentially valuable for the development of a serological test.     

Monoclonal antibodies against chicken type V collagen: production, specificity, and use for immunocytochemical localization in embryonic cornea and other organs

Two monoclonal antibodies have been produced against chick type V collagen and shown to be highly specific for separate, conformational dependent determinants within this molecule. When used for immunocytochemical tissue localization, these antibodies show that a major site for the in situ deposition of type V is within the extracellular matrices of many dense connective tissues. In these, however, it is largely in a form unavailable to the antibodies, thus requiring a specific “unmasking” treatment to obtain successful immunocytochemical staining. The specificity of these two IgG antibodies was determined by inhibition ELISA, in which only type V and no other known collagen shows inhibition. In ELISA, mixtures of the two antibodies give an additive binding reaction to the collagen, suggesting that each is against a different antigenic determinant. That both antigenic determinants are conformational dependent, being either in, or closely associated with, the collagen helix is demonstrated by the loss of antibody binding to molecules that have been thermally denatured. The temperature at which this occurs, as assayed by inhibition ELISA, is very similar to that at which the collagen helix melts, as determined by optical rotation. This gives strong additional evidence that the antibodies are directed against the collagen. The antibodies were used for indirect immunofluorescence analyses of cryostat sections of corneas and other organs from 17 to 18-day-old chick embryos. Of all tissues examined only Bowman’s membrane gave a strong staining reaction with cryostat sections of unfixed material. Staining in other areas of the cornea and in other tissues was very light or nonexistent. When, however, sections were pretreated with pepsin dissolved in dilute HAc or, surprisingly, with the dilute HAc itself dramatic new staining by the antibodies was observed in most tissues examined. The staining, which was specific for the anti-type V collagen antibodies, was largely confined to extracellular matrices of dense connective tissues. Experiments using protease inhibitors suggested that the “unmasking” did not involve proteolysis. We do not yet know the mechanism of this unmasking; however, one possibility is that the dilute acid causes swelling or conformational changes in a type-V collagen-containing supramolecular structure. Further studies should allow us to determine whether this is the case.     

A Flexible Mouse-On-Mouse Immunohistochemical Staining Technique Adaptable to Biotin-Free Reagents,Immunofluorescence, and Multiple Antibody Staining

Immunohistochemistry on mouse tissue utilizing mouse monoclonal antibodies presents a challenge. Secondary antibodies directed against the mouse monoclonal primary antibody of interest will also detect endogenous mouse immunoglobulin in the tissue. This can lead to significant spurious staining. Therefore, a “mouse-on-mouse” staining strategy is needed to yield credible data. This paper presents a method that is easy to use and highly flexible to accommodate both an avidin-biotin detection system as well as a biotin-free polymer detection system. The mouse primary antibody is first combined with an Fab fragment of an anti-mouse antibody in a tube and allowed sufficient time to form an antibody complex. Any non-complexed secondary antibody is bound up with mouse serum. The mixture is then applied to the tissue. The flexibility of this method is confirmed with the use of different anti-mouse antibodies followed by a variety of detection reagents. These techniques can be used for immunohistochemistry (IHC), immunofluorescence (IF), as well as staining with multiple primary antibodies. This method has also been adapted to other models, such as using human antibodies on human tissue and using multiple rabbit antibodies in dual immunofluorescence.     

Development of a quantitative diagnostic method of estrogen receptor expression levels by immunohistochemistry using organic fluorescent material-assembled nanoparticles

The detection of estrogen receptors (ERs) by immunohistochemistry (IHC) using 3,3′-diaminobenzidine (DAB) is slightly weak as a prognostic marker, but it is essential to the application of endocrine therapy, such as antiestrogen tamoxifen-based therapy. IHC using DAB is a poor quantitative method because horseradish peroxidase (HRP) activity depends on reaction time, temperature and substrate concentration. However, IHC using fluorescent material provides an effective method to quantitatively use IHC because the signal intensity is proportional to the intensity of the photon excitation energy. However, the high level of autofluorescence has impeded the development of quantitative IHC using fluorescence. We developed organic fluorescent material (tetramethylrhodamine)-assembled nanoparticles for IHC. Tissue autofluorescence is comparable to the fluorescence intensity of quantum dots, which are the most representative fluorescent nanoparticles. The fluorescent intensity of our novel nanoparticles was 10.2-fold greater than quantum dots, and they did not bind non-specifically to breast cancer tissues due to the polyethylene glycol chain that coated their surfaces. Therefore, the fluorescent intensity of our nanoparticles significantly exceeded autofluorescence, which produced a significantly higher signal-to-noise ratio on IHC-imaged cancer tissues than previous methods. Moreover, immunostaining data from our nanoparticle fluorescent IHC and IHC with DAB were compared in the same region of adjacent tissues sections to quantitatively examine the two methods. The results demonstrated that our nanoparticle staining analyzed a wide range of ER expression levels with higher accuracy and quantitative sensitivity than DAB staining. This enhancement in the diagnostic accuracy and sensitivity for ERs using our immunostaining method will improve the prediction of responses to therapies that target ERs and progesterone receptors that are induced by a downstream ER signal.     

The Application of the Tissue Microarray (TMA) Technology to Analyze Cerebral Organoids

“Multi-Omics” technologies have contributed greatly to the understanding of various diseases by enabling researchers to accurately and rapidly investigate the molecular circuitry that connects cellular systems. The tissue-engineered, three-dimensional (3D), in vitro disease model “organoid” integrates the “omics” results in a model system, elucidating the complex links between genotype and phenotype. These 3D structures have been used to model cancer, infectious disease, toxicity, and neurological disorders. Here, we describe the advantage of using the tissue microarray (TMA) technology to analyze human-induced pluripotent stem cell–derived cerebral organoids. Compared with the conventional processing of individual samples, sectioning and staining of TMA slides are faster and can be automated, decreasing labor and reagent costs. The TMA technology faithfully captures cell morphology variations and detects specific biomarkers. The use of this technology can scale up organoid research results in at least two ways: (1) in the number of specimens that can be analyzed simultaneously and (2) in the number of consecutive sections that can be produced for analysis with different probes and antibodies.     

Synergistic Tissue Counterstaining and Image Segmentation Techniques for Accurate, Quantitative Immunohistochemistry

Quantitative analysis of digitized IHC-stained tissue sections is increasingly used in research studies and clinical practice. Accurate quantification of IHC staining, however, is often complicated by conventional tissue counterstains caused by the color convolution of the IHC chromogen and the counterstain. To overcome this issue, we implemented a new counterstain, Acid Blue 129, which provides homogeneous tissue background staining. Furthermore, we combined this counterstaining technique with a simple, robust, fully automated image segmentation algorithm, which takes advantage of the high degree of color separation between the 3-amino-9-ethyl-carbazole (AEC) chromogen and the Acid Blue 129 counterstain. Rigorous validation of the automated technique against manual segmentation data, using Ki-67 IHC sections from rat C6 glioma and β-amyloid IHC sections from transgenic mice with amyloid precursor protein (APP) mutations, has shown the automated method to produce highly accurate results compared with ground truth estimates based on the manually segmented images. The synergistic combination of the novel tissue counterstaining and image segmentation techniques described in this study will allow for accurate, reproducible, and efficient quantitative IHC studies for a wide range of antibodies and tissues. (J Histochem Cytochem 56:873–880, 2008)     

Pharmacogenomics of pediatric asthma

CONTEXT:

Asthma is a complex disease with multiple genetic and environmental factors contributing to it. A component of this complexity is a highly variable response to pharmacological therapy. Pharmacogenomics is the study of the role of genetic determinants in the variable response to therapy. A number of examples of possible pharmacogenomic approaches that may prove of value in the management of asthma are discussed below.

EVIDENCE ACQUISITION:

A search of PubMed, Google scholar, E-Medicine, BMJ and Mbase was done using the key words “pharmacogenomics of asthma”, “pharmacogenomics of β-agonist, glucocorticoids, leukotriene modifiers, theophylline, muscarinic antagonists in asthma”.

RESULTS:

Presently, there are limited examples of gene polymorphism that can influence response to asthma therapy. Polymorphisms that alter response to asthma therapy include Arg16Gly, Gln27Glu, Thr164Ile for β-agonist receptor, polymorphism of glucocorticoid receptor gene, CRHR1 variants and polymorphism of LTC4S, ALOX5. Polymorphic variants of muscarinic receptors, PDE4 and CYP450 gene variants.

CONCLUSION:

It was concluded that genetic variation can improve the response to asthma therapy. However, no gene polymorphism has been associated with consistent results in different populations. Therefore, asthma pharmacogenomic studies in different populations with a large number of subjects are required to make possible tailoring the asthma therapy according to the genetic characteristic of individual patient.     

Nodal Morphogens

Nodal signals belong to the TGF-β superfamily and are essential for the induction of mesoderm and endoderm and the determination of the left–right axis. Nodal signals can act as morphogens—they have concentration-dependent effects and can act at a distance from their source of production. Nodal and its feedback inhibitor Lefty form an activator/inhibitor pair that behaves similarly to postulated reaction–diffusion models of tissue patterning. Nodal morphogen activity is also regulated by microRNAs, convertases, TGF-β signals, coreceptors, and trafficking factors. This article describes how Nodal morphogens pattern embryonic fields and discusses how Nodal morphogen signaling is modulated.In his 1901 book “Regeneration,” Thomas Hunt Morgan speculated that “if we suppose the materials or structures that are characteristic of the vegetative half are gradually distributed from the vegetative to the animal half in decreasing amounts, then any piece of the egg will contain more of these things at one pole than the other” and “gastrulation depends on the relative amounts of the materials in the different parts of the blastula” (Morgan 1901). Although Morgan’s speculations referred to the sea urchin embryo, they foretold our current understanding of morphogen gradients in frog and fish development. Morgan’s “materials,” “structures,” and “things” are the Nodal signals that create a vegetal-to-animal activity gradient to regulate germ layer formation and patterning. This article discusses how Nodal signaling provides positional information to fields of cells. I first portray the components of the signaling pathway and describe the role of Nodal signals in mesendoderm induction and left–right axis specification. I then discuss how Nodal morphogen gradients are thought to be generated, modulated, and interpreted.     

A new era for plant biotechnology. A combination of pharmaceutical applications, technologies and safety measures could lead to new applications for genetically modified plants

New applications and technologies, and more rigorous safety measures could herald a new era for genetically modified crops with improved traits, for use in agriculture and the pharmaceutical industry.The imminent prospect of the first approval of a plant-made pharmaceutical (PMP) for human use could herald a new era for applied plant science, after a decade of public backlash against genetically modified crops, particularly in Europe. Yet, the general resistance to genetically modified organisms might have done plant biotechnology a favour in the long run, by forcing it to adopt more-rigorous procedures for efficacy and safety in line with the pharmaceutical industry. This could, in turn, lead to renewed vigour for plant science, with the promise of developing not only food crops that deliver benefits to consumers and producers, but also a wide range of new pharmaceuticals.This is certainly the view of David Aviezer, CEO of Protalix, an Israeli company that has developed what could become the first recombinant therapeutic protein from plants to treat Gaucher disease. The protein is called taliglucerase alpha; it is a recombinant human form of the enzyme glucocerebrosidase that is produced in genetically engineered carrot cells. This enzyme has a crucial role in the breakdown of glycolipids in the cell membrane and is either used to provide energy or for cellular recognition. Deficiency of this enzyme causes accumulation of lipids with a variety of effects including premature death.“My feeling is that there is a dramatic change in this area with a shift away from the direction where a decade ago biotech companies like Monsanto and Dow went with growing transgenic plants in an open field, and instead moving this process into a more regulatory well-defined process inside a clean room,” Aviezer said. “Now the process is taking place in confined conditions and is very highly regulated as in the pharmaceutical industry.”…resistance to genetically modified organisms might have done plant biotechnology a favour […] forcing it to adopt more-rigorous procedures for efficacy and safety…He argues that this is ushering in a new era for plant biotechnology that could lead to greater public acceptance, although he denies that the move to clean-room development has been driven purely by the environmental backlash against genetically modified organisms in the late 1990s and early 2000s. “That was one aspect, but I think the move has been coming more from an appreciation that biopharmaceuticals require a more regulatory defined setting than is achieved at the moment with transgenic plants.”Interest in deriving pharmaceuticals from plants, known colloquially as ‘pharming'', first took off in the 1990s after researchers showed that monoclonal antibodies could be made in tobacco plants (Hiatt et al, 1989). This led to genetic engineering of plants to produce vaccines, antibodies and proteins for therapeutics, but none gained regulatory approval, mostly because of safety concerns. Moreover, the plants were grown in open fields, therefore attracting the same criticisms as transgenic food crops. In fact, a recent study showed that the views of the public on pharming depended on the product and the means to produce it; the researchers found increasing acceptance if the plants were used to produce therapeutics against severe diseases and grown in containment (Pardo et al, 2009).However, it was the technical challenges involved in purification and the associated regulatory issues that really delayed the PMP field, according to George Lomonossoff, project leader in biological chemistry at the John Innes Centre for plant research in Norwich in the UK, part of the Biotechnology and Biological Sciences Research Council (BBSRC). “Extraction from plants required the development of systems which are not clogged by the large amounts of fibrous material, mainly cellulose, and the development of GMP [good manufacturing practice; quality and testing guidelines for pharmaceutical manufacture] compliant methods of purification which are distinct from those required from, say, mammalian cells,” said Lomonossoff. “All this is very time consuming.”“Secondly there was no regulatory framework in place to assess the risks associated with proteins produced in plants, and determining how equivalent they are to mammalian-cell-produced material and what kind of contaminants you might have to guard against,” Lomonossoff added. “Again, attempting to address all possible concerns is a lengthy and expensive process.” Yet recent work by Protalix and a few other companies, such as Dow Agrosciences, has given grounds for optimism that purification and GMP-compliant methods of production have finally been established, Lomonossoff added.…a recent study showed that the views of the public on pharming depended on the product and the means to produce it…The first important breakthrough for PMPs came in 2006, when Dow Agrosciences gained regulatory approval from the US Department of Agriculture for a vaccine against Newcastle disease, a contagious bird infection caused by paramyxovirus PMV-1. “Though the vaccine, produced in tobacco-suspension culture cells, was never deployed commercially, it showed that regulatory approval for a plant-made pharmaceutical can be obtained, albeit for veterinary use in this case,” Lomonossoff said.As approval is imminent for taliglucerase alpha for human use, it is natural to ask why plants, as opposed to micro-organisms and animals, are worth the effort as sources of vaccines, antibiotics or hormones. There are three reasons: first, plants can manufacture some existing drugs more cheaply; second, they can do it more quickly; and third, and perhaps most significantly, they will be able to manufacture more complex proteins that cannot be produced with sufficient yield in any other way.An important example in the first category is insulin, which is being manufactured in increasing quantities to treat type 1 diabetes and some cases of type 2 diabetes. Until the arrival of recombinant DNA technology, replacement insulin was derived from the pancreases of animals in abattoirs, mostly cattle and pigs, but it is now more often produced from transgenic Escherichia coli, or sometimes yeast. Recently, there has been growing interest in using plants rather than bacteria as sources of insulin (Davidson, 2004; Molony et al, 2005). SemBioSys, a plant biotechnology company based in Calgary, Canada, is now developing systems to produce insulin and other therapeutic proteins in the seeds of safflower, an oilseed crop (Boothe et al, 2009).…plants can in principle be engineered to produce any protein, including animal ones…“We have developed technology that combines the high-capacity, low-cost production of therapeutic proteins in seeds with a novel technology that simplifies downstream purification,” said Joseph Boothe, vice president of research and development at SemBioSys. “The target proteins are engineered to associate with small, oil-containing structures within the seed known as oilbodies,” Boothe explained. “When extracted from the seed these oilbodies and associated proteins can be separated from other components by simple centrifugation. As a result, much of the heavy lifting around the initial purification is accomplished without chromatography, providing for substantial cost savings.”The second potential advantage of PMPs is their speed to market, which could prove most significant for the production of vaccines, either against emerging diseases or seasonal influenza, for which immunological changes in the virus mean that newly formulated vaccines are required each year. “In terms of a vaccine, I think influenza is very promising particularly as speed is of the essence in combating new strains,” Lomonossoff said. “Using transient expression methods, you can go from sequence to expressed protein in two weeks.” Transient gene expression involves injection of genes into a cell to produce a target protein, rather than permanently incorporating the gene into a host genome. This is emerging as a less technically difficult and faster alternative to developing stable cell lines for expressing bioengineered proteins. The process of injecting the desired gene into the target genome, known as transfection, can be effected not only by viruses, but also by non-viral agents including various lipids, polyethylenine and calcium phosphate.The last of the three advantages of plants for pharmaceutical production—the ability to manufacture proteins not available by other means—is creating perhaps the greatest excitement. The Protalix taliglucerase alpha protein falls into this category, and is likely to be followed by other candidates for treating disorders that require enzymes or complex molecules beyond the scope of bacteria, according to Aviezer. “I would say that for simpler proteins, bacteria will still be the method of choice for a while,” Aviezer said. “But for more complex proteins currently made via mammalian cells, I think we can offer a very attractive alternative using plant cells.”Indeed, plants can in principle be engineered to produce any protein, including animal ones, as Boothe pointed out. “In some cases this may require additional genetic engineering to enable the plant to perform certain types of protein modification that differ between plants and animals,” he said. “The classic example of this is glycosylation. With recent advances in the field it is now possible to engineer plants to glycosylate proteins in a manner similar to that of mammalian cells.” Glycosylation is a site-directed process that adds mono- or polysaccharides to organic molecules, and plays a vital role in folding and conferring stability on the finished molecule or macromolecule. Although plants can be engineered to perform it, bacteria generally cannot, which is a major advantage of plant systems over micro-organisms for pharmaceutical manufacture, according to Aviezer. “This enables plant systems to do complex folding and so make proteins for enzyme replacement or antibodies,” Aviezer said.Genomic-assisted breeding is being used either as a substitute for, or a complement to, genetic-modification techniques…In addition to plants themselves, their viruses also have therapeutic potential, either to display epitopes—the protein, sugar or lipid components of antigens on the surface of an infectious agent—so as to trigger an immune response or, alternatively, to deliver a drug directly into a cell. However, as Lomonossoff pointed out, regulatory authorities remain reluctant to approve any compound containing foreign nucleic acids for human use because of the risk of infection as a side effect. “I hope the empty particle technology [viruses without DNA] we have recently developed will revive this aspect,” Lomonossoff said. “The empty particles can also be used as nano-containers for targeted drug delivery and we are actively exploring this.”As pharmaceutical production is emerging as a new field for plant biology, there is a small revolution going on in plant breeding, with the emergence of genomic techniques that allow simultaneous selection across several traits. Although genetic modification can, by importing a foreign gene, provide instant expression of a desired trait, such as drought tolerance, protein content or pesticide resistance, the new field of genomics-assisted breeding has just as great potential through selection of unique variants within the existing gene pool of a plant, according to Douwe de Boer, managing director of the Netherlands biotech group Genetwister. “With this technology it will be possible to breed faster and more efficiently, especially for complex traits that involve multiple genes,” he said. “By using markers it is possible to combine many different traits in one cultivar, variety, or line in a pre-planned manner and as such breed superior crops.”“The application of genomics technologies and next generation sequencing will surely revolutionize plant breeding and will eventually allow this to be achieved with clinical precision”Genomic-assisted breeding is being used either as a substitute for, or a complement to, genetic-modification techniques, both for food crops to bolt on traits such as nutrient value or drought resistance, and for pharmaceutical products, for example to increase the yield of a desired compound or reduce unwanted side effects. Yet, there is more research required to make genomic-assisted breeding as widely used as established genetic-modification techniques. “The challenge in our research is to find markers for each trait and as such we extensively make use of bio-informatics for data storage, analysis and visualization,” de Boer said.The rewards are potentially enormous, according to Alisdair Fernie, a group leader from the Max-Planck-Institute for Molecular Plant Physiology in Potsdam, Germany. “Smart breeding will certainly have a massive impact in the future,” Fernie said. “The application of genomics technologies and next generation sequencing will surely revolutionize plant breeding and will eventually allow this to be achieved with clinical precision.” The promise of such genomic technologies in plants extends beyond food and pharmaceuticals to energy and new materials or products such as lubricants; the potential of plants is that they are not just able to produce the desired compound, but can often do so more quickly, efficiently and cheaply than competing biotechnological methods.     

Will we wake up to biodiversity?: The International Year of Biodiversity has failed to raise awareness or halt decline as economic crises and political interests have sidelined the environment

Looking back on the International Year of Biodiversity, some conservationists hope that it has raised awareness, if nothing else. Even so, many scientists remain pessimistic about our efforts to halt biodiversity decline.The United Nations'' (UN) International Year of Biodiversity in 2010 was supposed to see the adoption of measures that would slow global environmental decline and the continuing loss of endangered species and habitats. Even before, in 2002, most UN members had committed to halting the decline in biodiversity, which is a measure of the health of ecosystems. But the results of these international efforts have been funereal. Moreover, the current global economic crisis, coupled with growing anti-science attitudes in the USA, are adding to the concern of scientists about whether there is the political will to address the loss of biodiversity and whether habitat loss and extinction rates are reaching a point of no return.“There is not a single report received last year that claimed to have stopped or reduced the loss of biodiversity”Ahmed Djoghlaf, Executive Secretary of the Convention on Biological Diversity under the UN Environment Programme based in Montreal, Canada, said that of the 175 national reports submitted as part of the International Year of Biodiversity to his agency last year, none reported any progress. “There is not a single report received last year that claimed to have stopped or reduced the loss of biodiversity,” he said. “These reports confirm that the rate of loss of biodiversity today is unprecedented and the rate is 1,000 higher than the rate of natural extinction on species, and [his agency''s Global Biodiversity Outlook 2010; UN, 2010a] predicts that if business is allowed to continue then major ecosystems, the ocean, the fish, the forests, will reach the tipping point, meaning that there will be irreversible and irreparable damage done to the ecosystems.”The UN campaign traces its roots to the European Union (EU) commitment in 2001 to halt the loss of biodiversity by 2010. The 2010 goal was incorporated into the UN Millennium Development Goals because of the severe impact of biodiversity loss on human well-being. However, the EU last year conceded in a report that it missed its 2010 target, too. The EU''s Biodiversity Action Plan, launched in 2006, shows that Europe''s biodiversity “remains under severe threat from the excessive demands we are making on our environment, such as changes in land use, pollution, invasive species and climate change.” Yet, EU Environment Commissioner Janez Potočnik has seen some positive signs: “We have learned some very important lessons and managed to raise biodiversity to the top of the political agenda. But we need everyone on board and not just in Europe. The threat around the world is even greater than in the EU,” he wrote last year (EC, 2010).Despite the initiative''s poor report card, Djoghlaf was upbeat about the International Year of Biodiversity. “It was a success because it was celebrated everywhere,” he said. “In Switzerland, they conducted a survey before and after the International Year of Biodiversity and they concluded that at the end of the year, 67% of all the Swiss people are now aware of biodiversity. When the year started it was 40%. People are more and more aware. In addition, biodiversity has entered the top of the political agenda.”In October 2010, delegates from 193 countries attended the UN Convention on Biodiversity in Nagoya, Japan, and adopted 20 strategic goals to be achieved by 2020 (UN, 2010b). The so-called Aichi Biodiversity Targets include increased public awareness of the values of biodiversity and the steps that individuals can take to conserve and act sustainably; the halving or halting of the rate of loss of all natural habitats, including forests; and the conservation of 17% of terrestrial and inland water, and 10% of coastal and marine areas through effective and equitable management, resulting in ecologically representative and well-connected systems. By contrast, 13% of land areas and 1% of marine areas were protected in 2010.However, the Convention on Biological Diversity is not enforceable. Anne Larigauderie, Executive Director of DIVERSITAS (Paris, France), which promotes research on biodiversity science, said that it is up to the individual countries to adopt enforceable legislation. “In principle, countries have committed. Now it depends on what individual countries are going to do with the agreement,” she said. “I would say that things are generally going in the right direction and it''s too early to tell whether or not it''s going to have an impact in terms of responding and in terms of the biodiversity itself.”Researchers, however, have been disappointed by The International Year of Biodiversity. Conservation biologist Stuart Butchart, of Birdlife International in Cambridge, UK—a partnership of non-governmental environmental organizations and colleagues from other environmental groups—compiled a list of 31 indicators to measure progress towards the 2010 goal of the International Year of Biodiversity. He and his collaborators reported in Science (Butchart et al, 2010) that these indicators, including species population trends, extinction risks and habitat conditions, showed declines with no significant rate reductions. At the same time, indicators of pressure on biodiversity, such as resource consumption, invasive alien species, nitrogen pollution, over-exploitation and climate change impacts showed increases. “Despite some local successes and increasing responses (including extent and biodiversity coverage of protected areas, sustainable forest management, policy responses to invasive alien species and biodiversity-related aid), the rate of biodiversity loss does not appear to be slowing,” the researchers wrote.​wrote.Open in a separate window© Thomas Kitchin & Victoria Hurst/Wave/CorbisButchart pointed out that even if the International Year of Biodiversity had an impact on raising awareness and reducing biodiversity loss, detecting the change would take time. He said that the International Year of Biodiversity fell short of increasing awareness in parts of government not dealing with the environment, including ministries of transport, tourism, treasury and finance. It also seems probable that the campaign had little impact on the business sector, which affects development projects with a direct impact on biodiversity. “People can''t even seem to get together on global climate change, which is a whole lot more obvious and right there,” Peter Raven, president emeritus of the Missouri Botanical Gardens in St Louis, USA, explained. “Biodiversity always seems to be a sort of mysterious background thing that isn''t quite there.”“People can''t even seem to get together on global climate change, which is a whole lot more obvious and right there…”Illka Hanski, a professor in the Department of Ecology and Evolutionary Biology at the University of Helsinki in Finland, said that studies such as Butchart''s “indicate that nothing really happened in 2010. Biodiversity decline continued and has been declining over the past 10 years.”Other researchers are more positive, although with reservations. Conservation biologist Thomas Eugene Lovejoy III, Heinz Center Biodiversity Chair and former president of the Center in Washington, DC, USA—a non-partisan, non-profit organization dedicated to advancing sound environmental policy—said that economic trends affect biodiversity and that biodiversity efforts might actually be benefiting from the current global economic crisis. For example, the decline in the housing markets in the USA and Europe has reduced the demand on lumber for new construction and has led to a reduction in deforestation. “Generally speaking, when there is an economic downturn, some of the things that are pressuring biodiversity actually abate somewhat. That''s the good news. The bad news is that the ability to marshal resources to do some things proactively gets harder,” he said.Chris Thomas, a conservation biologist at the University of York in the UK, who studies ecosystems and species in the context of climate change, said that economic depressions do slow the rate of damage to the environment. “But it also takes eyes off the ball of environmental issues. It''s not clear whether these downturns, when you look over a period of a decade, make much difference or not.” Hanski agreed: “[B]ecause there is less economic activity, there may be less use of resources and such. But I don''t think this is a way to solve our problems. It won''t lead to any stable situation. It just leads to a situation where economic policies become more and more dependent on measures that try actually just to increase the growth as soon as possible.”…biodiversity efforts might actually be benefiting from the current global economic crisisRaven said that in bad times, major interests such as those involved in raising cattle, growing soybeans and clearing habitat for oil palms have reduced political clout because there is less money available for investment. But he said economic downturns do not slow poor people scrounging for sustenance in natural habitats.To overcome this attitude of neglect, Lovejoy thinks there ought to be a new type of ‘economics'' that demonstrates the benefits of biodiversity and brings the “natural world into the normal calculus.” Researchers are already making progress in this direction. Thomas said that the valuation of nature is one of the most active areas of research. “People have very different opinions as to how much of it can be truly valued. But it is a rapidly developing field,” he said. “Once you''ve decided how much something is worth, then you''ve got to ask what are the financial or other mechanisms by which the true value of this resource can be appreciated.”Hanski said that the main problem is the short-term view of economic forecasts. “Rapid use of natural resources because of short-term calculation may actually lead to a sort of exploitation rather than conservation or preservation.” He added that the emphasis on economic growth in rich societies in North America and Europe is frustrating. “We have become much richer than in 1970 when there actually was talk of zero growth in serious terms. So now we are richer and we are becoming more and more dependent on continued growth, the opposite of what we should be aiming at. It''s a problem with our society and economics clearly, but I can''t be very optimistic about the biodiversity or other environmental issues in this kind of situation.” He added that biodiversity is still taking a backseat to economics: “There is a very long way to go right now with the economic situation in Europe, it''s clear that these sorts of [biodiversity] issues are not the ones which are currently being debated by the heads of states.”The economic downturn, which has led to reduced government and private funding and declines in endowments, has also hurt organizations dedicated to preserving biodiversity. Butchart said that some of the main US conservation organizations, including the Nature Conservancy and the World Wildlife Federation, have experienced staff cuts up to 30%. “Organizations have had to tighten their belts and reign in programmes just to stay afloat, so it''s definitely impacted the degree to which we could work effectively,” he said. “Most of the big international conservation organizations have had to lay off large numbers of staff.”…a new type of ‘economics'' that demonstrates the benefits of biodiversity and brings the “natural world into the normal calculus”Cary Fowler, Executive Director of the Global Crop Diversity Trust in Rome, Italy, a public–private partnership to fund key crop collections for food security, also feels the extra challenges of the global economic crisis. “We invest our money conservatively like a foundation would in order to generate income that can reliably pay the bills in these seed banks year after year. So I''m always nervous and I have the computer on at the moment looking at what''s happening with the sovereign debt crisis here in Europe. It''s not good,” he said. “Governments are not being very generous with contributions to this area. Donors will rarely give a reason [for cutting funding].”The political situation in the USA, the world''s largest economy, is also not boding well for conservation of and research into biodiversity. The political extremism of the Republican Party during the run up to the 2012 presidential election has worried many involved in biodiversity issues. Republican contender Texas Governor Rick Perry has been described as ‘anti-science'' for his denial of man-made climate change, a switch from the position of 2008 Republican candidate John McCain. Perry was also reported to describe evolution as a “theory that''s out there, and it''s got some gaps in it” at a campaign event in New Hampshire earlier in the year.“Most of the big international conservation organizations have had to lay off large numbers of staff”Raven said this attitude is putting the USA at a disadvantage. “It drives us to an anti-intellectualism and a lack of real verification for anything which is really serious in terms of our general level of scientific education and our ability to act intelligently,” he said.Still, Larigauderie said that although the USA has not signed the conventions on biodiversity, she has seen US observers attend the meetings, especially under the Obama administration. “They just can''t speak,” she said. Meanwhile, Lovejoy said that biodiversity could get lost in the “unbelievable polarisation affecting US politics. I have worked out of Washington for 36 years now—I''ve never seen anything like this: an unwillingness to actually listen to the other side.”Raven said it is vital for the USA to commit to preserving biodiversity nationally and internationally. “It''s extremely important because our progress towards sustainability for the future will depend on our ability to handle biodiversity in large part. We''re already using about half of all the total photosynthetic productivity on land worldwide and that in turn means we''re cutting our options back badly. The US is syphoning money by selling debt and of course promoting instability all over the world,” he explained. “It''s clear that there is no solution to it other than a level population, more moderate consumption levels and new technologies altogether.”The EU and the UN have also changed the time horizon for halting the decline in biodiversity. As part of the Nagoya meeting, the UN announced the UN Decade for Biodiversity. The strategic objectives include a supporting framework for the implementation of the Biodiversity Strategic Plan 2011–2020 and the Aichi Biodiversity Targets, as well as guidance to regional and international organizations, and more public awareness of biodiversity issues.But Butchart remains sceptical. “I suspect ‘decades of whatever'' have even less impact than years,” he said. “2008 was the International Year of the Potato. I don''t know how much impact that had on your life and awareness. I think there is greater awareness and greater potential to make significant progress in addressing biodiversity loss now than there was 10 years ago, but the scale of the challenge is still immense.”“…our progress towards sustainability for the future will depend on our ability to handle biodiversity in large part”Hanski has similar doubts. “I believe it''s inevitable that a very large fraction of the species on Earth will go extinct in the next hundred years. I can''t see any change to that.” But he is optimistic that some positive change can be made. “Being pessimistic doesn''t help. The nations still can make a difference.” He said he has observed ecotourism playing a role in saving some species in Madagascar, where he does some of his research.“We''re not going to fundamentally be able to wipe life off the planet,” Thomas said. “We will wipe ourselves off the planet virtually certainly before we wipe life out on Earth. However, from the point of view of humanity as a culture, and in terms of the resources we might be able to get from biodiversity indirectly or directly, if we start losing things then it takes things millions of years to ‘re-evolve'' something that does an equivalent job. From a human perspective, when we wipe lots of things out, they''re effectively permanently lost. Of course it would be fascinating and I would love to be able to come back to the planet in 10 million years and see what it looks like, assuming humans are not here and other stuff will be.”Djoghlaf, by contrast, is more optimistic about our chances: “I believe in the human survival aspect. When humankind realises that the current pattern of production and consumption and the current way that it is dealing with nature is unsustainable, we will wake up.”     

Classification and analysis of regulatory pathways using graph property, biochemical and physicochemical property, and functional property

Given a regulatory pathway system consisting of a set of proteins, can we predict which pathway class it belongs to? Such a problem is closely related to the biological function of the pathway in cells and hence is quite fundamental and essential in systems biology and proteomics. This is also an extremely difficult and challenging problem due to its complexity. To address this problem, a novel approach was developed that can be used to predict query pathways among the following six functional categories: (i) “Metabolism”, (ii) “Genetic Information Processing”, (iii) “Environmental Information Processing”, (iv) “Cellular Processes”, (v) “Organismal Systems”, and (vi) “Human Diseases”. The prediction method was established trough the following procedures: (i) according to the general form of pseudo amino acid composition (PseAAC), each of the pathways concerned is formulated as a 5570-D (dimensional) vector; (ii) each of components in the 5570-D vector was derived by a series of feature extractions from the pathway system according to its graphic property, biochemical and physicochemical property, as well as functional property; (iii) the minimum redundancy maximum relevance (mRMR) method was adopted to operate the prediction. A cross-validation by the jackknife test on a benchmark dataset consisting of 146 regulatory pathways indicated that an overall success rate of 78.8% was achieved by our method in identifying query pathways among the above six classes, indicating the outcome is quite promising and encouraging. To the best of our knowledge, the current study represents the first effort in attempting to identity the type of a pathway system or its biological function. It is anticipated that our report may stimulate a series of follow-up investigations in this new and challenging area.     

The Electrophysiological Underpinnings of Processing Gender Stereotypes in Language

Despite the widely documented influence of gender stereotypes on social behaviour, little is known about the electrophysiological substrates engaged in the processing of such information when conveyed by language. Using event-related brain potentials (ERPs), we examined the brain response to third-person pronouns (lei “she” and lui “he”) that were implicitly primed by definitional (passeggera _FEM “passenger”, pensionato _MASC “pensioner”), or stereotypical antecedents (insegnante “teacher”, conducente “driver”). An N400-like effect on the pronoun emerged when it was preceded by a definitionally incongruent prime (passeggera _FEM – lui; pensionato _MASC – lei), and a stereotypically incongruent prime for masculine pronouns only (insegnante – lui). In addition, a P300-like effect was found when the pronoun was preceded by definitionally incongruent primes. However, this effect was observed for female, but not male participants. Overall, these results provide further evidence for on-line effects of stereotypical gender in language comprehension. Importantly, our results also suggest a gender stereotype asymmetry in that male and female stereotypes affected the processing of pronouns differently.     

Monoclonal antibodies against plant proteins recognise animal intermediate filaments

Four monoclonal antibodies were raised against polypeptides present in a high-salt detergent-insoluble fraction from cells of Chlamydomonas reinhardtii. Indirect immunofluorescence microscopy of fibroblasts and epithelial cells grown in culture using these plant antibodies revealed staining arrays identical to those obtained with well characterised antibodies to animal intermediate filaments. Immunofluorescence microscopy of Chlamydomonas with these monoclonal antibodies and a monoclonal antibody that recognises all animal intermediate filaments (anti-IFA) gave a diffuse, patchy cytoplasmic staining pattern. Both the plant antibodies and anti-IFA stained interphase onion root tip cells in a diffuse perinuclear pattern. In metaphase through to telophase, the labelling patterns colocalised with those of microtubules. Labelling of the phragmoplast was also detected but not staining of the preprophase band. On Western blots of various animal cell lines and tissues, all the antibodies labelled known intermediate filament proteins. On Western blots of whole Chlamydomonas proteins, all the antibodies labelled a broad band in the 57,000 Mr range, and three antibodies labelled bands around 66,000 and 140,000 Mr but with variable intensities. On Western blots of whole onion root tip proteins, all the antibodies labelled 50,000 Mr (two to three bands) polypeptides and a diffuse band around 60,000 Mr and three of the antibodies also labelled several polypeptides in the 90,000-200,000 Mr range. The consistent labelling of these different bands by several different monoclonal antibodies recognising animal intermediate filaments makes these polypeptides putative plant intermediate filament proteins.     

Plasticity in interactions of fibroblast growth factor 1 (FGF1) N terminus with FGF receptors underlies promiscuity of FGF1

Tissue-specific alternative splicing in the second half of Ig-like domain 3 (D3) of fibroblast growth factor receptors 1–3 (FGFR1 to -3) generates epithelial FGFR1b-FGFR3b and mesenchymal FGFR1c-FGFR3c splice isoforms. This splicing event establishes a selectivity filter to restrict the ligand binding specificity of FGFRb and FGFRc isoforms to mesenchymally and epithelially derived fibroblast growth factors (FGFs), respectively. FGF1 is termed the “universal FGFR ligand” because it overrides this specificity barrier. To elucidate the molecular basis for FGF1 cross-reactivity with the “b” and “c” splice isoforms of FGFRs, we determined the first crystal structure of FGF1 in complex with an FGFRb isoform, FGFR2b, at 2.1 Å resolution. Comparison of the FGF1-FGFR2b structure with the three previously published FGF1-FGFRc structures reveals that plasticity in the interactions of the N-terminal region of FGF1 with FGFR D3 is the main determinant of FGF1 cross-reactivity with both isoforms of FGFRs. In support of our structural data, we demonstrate that substitution of three N-terminal residues (Gly-19, His-25, and Phe-26) of FGF2 (a ligand that does not bind FGFR2b) for the corresponding residues of FGF1 (Phe-16, Asn-22, and Tyr-23) enables the FGF2 triple mutant to bind and activate FGFR2b. These findings taken together with our previous structural data on receptor binding specificity of FGF2, FGF8, and FGF10 conclusively show that sequence divergence at the N termini of FGFs is the primary regulator of the receptor binding specificity and promiscuity of FGFs.     

Let's make Golgi

Does the Golgi self-organize or does it form around an instructive template? Evidence on both sides is piling up, but a definitive conclusion is proving elusive.In the battle to define the Golgi, discussions easily spiral into what can appear like nitpicking. In a contentious poster session, an entire worldview rests on whether you think a particular mutant is arrested with vesicles that are close to but distinct from the ER or almost budded from but still attached to the ER.Sometimes obscured by these details are the larger issues. This debate “gets to the fundamental issue of how you think of the Golgi,” says Ben Glick of the University of Chicago (Chicago, IL). “The dogma has been that you need a template to build an organelle. But in the secretory system it''s possible in principle that you could get de novo organization of structure. That''s the issue that stirs people emotionally and intellectually.”Then there are the collateral issues. There is an ongoing controversy about the nature of forward transport through the Golgi—it may occur via forward movement of small vesicles, or by gradual maturation of one cisterna to form the next. The cisternal maturation model “argues for a Golgi that can be made and consumed,” says Graham Warren (Yale University, New Haven, CT)—a situation that is more difficult to reconcile with Warren''s template-determined universe.Even more confusing is the situation in mitosis. Accounts vary wildly on how much of the Golgi disappears into the ER during mitosis. The answer would determine to what extent the cell has to rebuild the Golgi after mitosis, and what method it might use to do so.Several laboratories have made major contributions to address these issues. But none define them so clearly as those of Warren and Jennifer Lippincott-Schwartz (National Institutes of Health, Bethesda, MD). At almost every turn, on almost every issue, it seems that Warren and Lippincott-Schwartz reach opposite conclusions, sometimes based on similar or identical data.And yet, at least in public, there is a remarkable lack of rancor. “These are not easy experiments for us to do,” says Warren. “It''s all cutting-edge research and we are pushing the technology to the limit. Part of that is that you push your own interpretation.” For her part, Lippincott-Schwartz approaches a lengthy poster-session debate with Warren with something approaching glee. This is not triumphal glee, however. Rather, Lippincott-Schwartz seems to relish the opportunity to exchange ideas, and on this point Warren agrees. “Complacency is the worst thing to have in a field,” he says. The debate “has made all of us think a lot harder.”