Spider silk as a resource for future biotechnologies

Insect silks have been used by mankind for millennia to produce textiles and in particular, the cocoon silk of Bombyx mori was the base of one of the most important industries in history. In fact, B. mori is probably the only domesticated insect if not invertebrate in its true and strict sense, comparable to cattle and other livestock that humans have known and bred since the Neolithic period. In contrast, reports regarding the use of spider silk throughout history have the character of travellers’ tales or anecdotes, and serious attempts to exploit these biomaterials on a large scale have not been undertaken until recently. Indeed, the cannibalism of these carnivores makes their farming difficult and the production of significant yields of spider silk virtually impossible. Only today, with recombinant technologies available, does this problem seem to have been overcome. But why use spider silk at all – if we have the infrastructure to produce significant yields of silk from Bombyx? In contrast to most insects, spiders do not spin from labial glands, and many spiders possess different types of gland, most of them active throughout the whole lifespan. Typical orb‐weavers (Araneoidea) for instance possess up to seven different types of silk gland to produce different silk fibers and glues. Each of these products has evolved for a particular use and the respective material properties are highly adapted to that use. As the group of Araneae is about 400 million years old, the oldest fossil orb‐weaver is dated about 150 million years, and the use of silk is crucial to a spider's survival, we can expect that evolution will have “squeezed out every iota” to achieve optimum performance at minimum cost. Indeed, some dragline silks such as the major ampullate silks of some Nephila species show amazing mechanical properties that, in terms of toughness, are far superior to Bombyx silk. Labels like “stronger than steel” or “even better than Kevlar” were attached to them, and the Canadian‐based biotech company Nexia created the trademark “bio‐steel” for their prospective product. The discovery of these exceptional mechanical properties of those protein fibers triggered intense research on spider silk, with the goal of their commercial exploitation. But there is more to Arachne's weave and science is beginning to pick up those threads.     

棒络新妇和悦目金蛛丝腺形态初步观察

研究比较了结网型蜘蛛棒络新妇Nephila clavata和悦目金蛛Argiope amoena的丝腺形态特征,为国内蜘蛛丝腺蛋白的研究提供原始的丝腺解剖图,同时结合对2种蜘蛛卵袋的解剖、网的特征和室内捕食黄粉虫Tenebrio molitor幼虫行为的观察比较,探讨了2种蜘蛛丝腺的生物学功能与其生存繁殖策略之间的关系。本文分别观察描述了棒络新妇和悦目金蛛的大壶状腺、小壶状腺、鞭状腺、柱状腺、葡萄状腺和梨状腺共6种丝腺。2种蜘蛛丝腺形态特征基本相似;部分丝腺在形态结构和颜色上有些差异;悦目金蛛的葡萄状腺比棒络新妇发达。观察表明2种蜘蛛的网和卵袋特征差异较大,两者捕食策略也不同,棒络新妇采用咬一捆缚（Bit—Wrapping）策略,悦目金蛛则采用捆缚一咬（Wrapping-Bit）策略。棒络新妇和悦目金蛛的网和卵袋特征与丝腺的颜色相一致。同时,其葡萄状腺数量和大小与其各自的捕食策略相关。     

Water extraction by the major ampullate duct during silk formation in the spider, Argiope aurantia Lucas

The water, K⁺ and Na⁺ content of naturally produced major ampullate silk as well as silk mechanically drawn from the spider Argiope aurantia have been compared to that of the major ampullate gland. It is demonstrated that water is extracted by the major ampullate duct and that this process is accompanied by an exchange of K⁺ for Na⁺. The significance of these observations is discussed.     

Indirect predator effects influence behaviour but not morphology of juvenile coral reef Ambon damselfish Pomacentrus amboinensis

A 6-week laboratory experiment exposed juvenile Ambon damselfish Pomacentrus amboinensis to visual and chemical cues of either a predator, a herbivore or a null control (sea water) and found no effect of predator cues on prey morphology (proportion of ocellus to eye diameter, body depth, standard length and fin area). Nonetheless, behaviour was significantly affected by predator presence, with prey less active and taking half as many feeding strikes when exposed to predators compared to fish from the null control. The presence of a herbivore also affected prey behaviour similar to that of the predator, suggesting that the presence of a non-predator may have important effects on development.     

Deciduous leaf drop reduces insect herbivory

Deciduous leaf fall is thought to be an adaptation that allows plants living in seasonal environments to reduce water loss and damage during unfavorable periods while increasing photosynthetic rates during favorable periods. Observations of natural variation in leaf shedding suggest that deciduous leaf fall may also allow plants to reduce herbivory. I tested this hypothesis by experimentally manipulating leaf retention for Quercus lobata and observing natural rates of herbivory. Quercus lobata is primarily deciduous although individuals show considerable natural variation in leaf retention. Oak saplings with no leaves through winter experienced reduced attack by cynipid gall makers the following spring. This pattern was consistent with the positive correlation between natural leaf persistence and gall numbers. These cynipids do not overwinter on the leaves that trees retain through winter, although they appear to use persistent leaves as oviposition cues. If these results are general for woody plants in continental temperate habitats, they suggest that an important and unrecognized consequence of deciduous leaf shedding may be a reduction in herbivore damage, and that this effect should be included in models of deciduous and evergreen behavior.     

Spider dragline silk: correlated and mosaic evolution in high-performance biological materials

The evolution of biological materials is a critical, yet poorly understood, component in the generation of biodiversity. For example, the diversification of spiders is correlated with evolutionary changes in the way they use silk, and the material properties of these fibers, such as strength, toughness, extensibility, and stiffness, have profound effects on ecological function. Here, we examine the evolution of the material properties of dragline silk across a phylogenetically diverse sample of species in the Araneomorphae (true spiders). The silks we studied are generally stronger than other biological materials and tougher than most biological or man-made fibers, but their material properties are highly variable; for example, strength and toughness vary more than fourfold among the 21 species we investigated. Furthermore, associations between different properties are complex. Some traits, such as strength and extensibility, seem to evolve independently and show no evidence of correlation or trade-off across species, even though trade-offs between these properties are observed within species. Material properties retain different levels of phylogenetic signal, suggesting that traits such as extensibility and toughness may be subject to different types or intensities of selection in several spider lineages. The picture that emerges is complex, with a mosaic pattern of trait evolution producing a diverse set of materials across spider species. These results show that the properties of biological materials are the target of selection, and that these changes can produce evolutionarily and ecologically important diversity.     

Chimeric spider silk proteins mediated by intein result in artificial hybrid silks

Hybrid silks hold a great potential as specific biomaterials due to its controlled mechanical properties. To produce fibers with tunable properties, here we firstly made chimeric proteins in vitro, called W2C4CT and W2C8CT, with ligation of MaSp repetitive modules (C) with AcSp modules (W) by intein trans splicing technology from smaller precursors without final yield reduction. Intein mediated chimeric proteins form fibers at a low concentration of 0.4 mg/mL in 50 mM K₃PO₄ pH 7.5 just drawn by hand. Hybrid fibers show smoother surface, and also have stronger chemical resistance as compared with fibers from W2CT (W fibers) and mixture of W2CT/C8CT (MHF8 fibers). Fibers from chimeric protein W2C4CT (HFH4) have improved mechanical properties than W fibers; however, with more C modules W2C8CT fibers (HFH8) properties decreased, indicates the length proportion of various modules is very important and should be optimized for fibers with specific properties. Generally, hybrid silks generated via chimeric proteins, which can be simplified by intein trans splicing, has greater potential to produce fibers with tunable properties. Our research shows that intein mediated directional protein ligation is a novel way to make large chimeric spider silk proteins and hybrid silks. © 2016 Wiley Periodicals, Inc. Biopolymers 105: 385–392, 2016.     

Tree diversity reduces herbivory by forest insects

Biodiversity loss from plant communities is often acknowledged to affect primary production but little is known about effects on herbivores. We conducted a meta-analysis of a worldwide data set of 119 studies to compare herbivory in single-species and mixed forests. This showed a significant reduction of herbivory in more diverse forests but this varied with the host specificity of insects. In diverse forests, herbivory by oligophagous species was virtually always reduced, whereas the response of polyphagous species was variable. Further analyses revealed that the composition of tree mixtures may be more important than species richness per se because diversity effects on herbivory were greater when mixed forests comprised taxonomically more distant tree species, and when the proportion of non-host trees was greater than that of host trees. These findings provide new support for the role of biodiversity in ecosystem functioning across trophic levels.     

悦目金蛛拖丝的超微结构研究

采用非固定的抽丝方法,从一只未麻醉的悦目金蛛(Argiope amoena)的纺器将拖丝抽出,然后用扫描电镜(SEM)对拖丝进行超微结构的观察,结果表明:悦目金蛛拖丝至少具有2根、3根、4根及多根单丝纤维构成的4种不同结构,其中有一种类似弹簧的结构;另外,丝的表面还出现一种小环结构,这两种结构可能是拖丝纤维具有优良机械性能的原因之一。"束状结构"和"小环结构"在文献中未见报道。拖丝的直径范围为0.25~10.77μm;悦目金蛛似乎能调节拖丝的结构和直径,以适应其所面临的即时环境。本文基于上述观察结果并结合前人的研究,提出了蜘蛛拖丝结构-生物学功能多样性假说,对蜘蛛丝的结构与生物学功能之间的关系作了探讨。     

蜘蛛丝蛋白研究进展

由于蜘蛛丝蛋白分子高度重复的一级结构、特殊的溶解特性和分子折叠行为以及具有形成非凡力学特性丝纤维的能力而引人注目。本文从蛛丝蛋白基因、天然蛛丝形成过程、蛛丝蛋白的基因工程生产及蛛丝蛋白的应用前景等几个方面着重介绍了近20年来对蛛丝蛋白的研究进展。围绕蛛丝蛋白展开的研究将有助于揭示蛋白质一级结构、蛋白质分子折叠与蛋白质大分子特性之间的内在联系。     

Urbanization drives unique latitudinal patterns of insect herbivory and tree condition

Urban landscapes are characterized by high proportions of impervious surface resulting in higher temperatures than adjacent natural landscapes. In some cities, like those at cooler latitudes, trees may benefit from warmer urban temperatures, but trees in many cities are beset with problems like drought stress and increased herbivory. What drives patterns of urban tree health across urbanization and latitudinal temperature gradients? In natural systems, latitude–herbivory relationships are well‐studied, and recent temperate studies have shown that herbivory generally increases with decreasing latitudes (warmer temperatures). However, the applicability of this latitude–herbivory theory in already‐warmed urban systems is unknown. In this study, we investigated how the interaction of urbanization, latitudinal warming and scale insect abundance affected urban tree health. We predicted that trees in warmer, lower latitude cities would be in poorer health at lower levels of urbanization than trees at cooler, higher latitudes due to the interaction of urbanization, latitudinal temperature and herbivory. To evaluate our predictions, we surveyed the abundance of scale insect herbivores on a single, common tree species Acer rubrum in eight US cities spanning 10° of latitude. We estimated urbanization at two extents, a local one that accounted for the direct effects on an individual tree, and a larger one that captured the surrounding urban landscape. We found that urban tree health did not vary with latitudinal temperature but was best predicted by local urbanization and herbivore abundance. We did not observe increased herbivore abundance in warmer, lower latitudes cities, but instead herbivore abundance peaked in the mid latitudes of our study. This study demonstrates that urban landscapes may deviate from classical theory developed in natural systems and reinforces the need for research reconciling ecological patterns in urban landscapes.     

A novel property of spider silk: chemical defence against ants

Spider webs are made of silk, the properties of which ensure remarkable efficiency at capturing prey. However, remaining on, or near, the web exposes the resident spiders to many potential predators, such as ants. Surprisingly, ants are rarely reported foraging on the webs of orb-weaving spiders, despite the formidable capacity of ants to subdue prey and repel enemies, the diversity and abundance of orb-web spiders, and the nutritional value of the web and resident spider. We explain this paradox by reporting a novel property of the silk produced by the orb-web spider Nephila antipodiana (Walckenaer). These spiders deposit on the silk a pyrrolidine alkaloid (2-pyrrolidinone) that provides protection from ant invasion. Furthermore, the ontogenetic change in the production of 2-pyrrolidinone suggests that this compound represents an adaptive response to the threat of natural enemies, rather than a simple by-product of silk synthesis: while 2-pyrrolidinone occurs on the silk threads produced by adult and large juvenile spiders, it is absent on threads produced by small juvenile spiders, whose threads are sufficiently thin to be inaccessible to ants.     

两种不同种植方式对龙葵叶片虫食状的影响研究

昆虫与植被之间相互关系的深入研究,有助于更好的理解生态系统结构与功能之间的关系以及生态系统生物多样性维持机理。本文通过对集中种植和分块种植下两种不同种植方式龙葵叶片虫食状进行调查分析,以期对龙葵的栽培和养护提供理论基础和技术建议。研究结果表明:龙葵叶片中共识别出11种虫食状类型,其中,集中种植方式有10种,分块种植方式有11种,各种虫食状类型出现频率在0.4%-24.7%;集中种植样地的龙葵叶片虫食状种类数、Shannon-Wiener指数和Pielou均匀度指数均低于分块种植;分块种植的龙葵叶片受到植食性昆虫的伤害频率较高,但是集中种植的龙葵叶片受到的伤害程度却明显高于分块种植方式。因此,种植龙葵应尽量避免大规模集中种植方式,不同生态系统边界之间边缘效应的尺度和强弱的关系是未来研究的重点和核心问题。     

蛋白质内含子介导蛛丝功能化平台的建立

为建立高效快捷的蛛丝功能化修饰平台,蛋白质内含子的反式剪接技术被首次应用于重组蛛丝的功能化修饰。在体外通过Ssp Dna B的反式剪接作用,在蛋白质水平上将12 k Da泛素相关修饰蛋白(SUMO)与蛛丝蛋白(W2CT)连接形成功能化蛛丝蛋白SUMOW2CT。修饰后SUMOW2CT与W2CT均能形成纳米至微米级的丝纤维,但SUMOW2CT自动成丝速度明显下降且产量约为W2CT的一半。与W2CT丝纤维(W)相似,SUMOW2CT丝纤维(UW)不具有超收缩能力和对2%SDS不耐受,但机械性能低于W2CT丝纤维。功能化蛋白SUMOW2CT形成的丝纤维中SUMO蛋白仍保持着正确三维结构,可被SUMO蛋白酶酶切。外源功能化蛋白质虽在一定程度上降低了丝的形成速度和机械性能,但修饰上的功能化蛋白仍保持着生物活性,表明断裂蛋白质内含子介导的蛛丝修饰平台成功建立,也为蛛丝的功能化修饰和应用奠定了坚实的技术基础。     

昆虫植食对植物有性生殖的影响

植物在个体发育的各个阶段都与不同的群落成员相互作用,如竞争的植物、有益的传粉者和敌对的植食动物。昆虫植食在各类生态系统中普遍存在,并可能对植物有性生殖产生各种影响。植食昆虫可通过对植物有性生殖结构的消耗直接对植物生殖产生影响,也可通过影响植物资源分配和花性状等改变传粉者服务,从而间接对植物有性生殖带来正面、负面或中性的影响。同一植物的植食昆虫和传粉者往往对植物的吸引性状 （如花大小、气味、颜色等）有相同的偏好,因此植食者与传粉者均能对植物有性生殖性状施加选择压力。本文从昆虫植食对植物有性生殖的直接影响、间接影响以及植食昆虫对植物有性生殖性状选择的影响三个方面进行综述,以期为昆虫植食和生物资源多样性保护相关研究提供参考。     

Purification of Spider Silk-elastin from Transgenic Plants and Application for Human Chondrocyte Proliferation

Research on spider silk proteins has led to the possibility of designing genetically engineered silks according to defined material properties. Here we show the efficient and stable production of spider silk-elastin fusion proteins in transgenic tobacco and potato plants by retention in the ER. The proteins were purified by a simple method, using heat treatment and 'inverse transition cycling'. Laboratory scale extraction of 1 kg tobacco leaf material leads to a yield of 80 mg pure recombinant spider silk-elastin protein. As a possible application, as well as to demonstrate biocompatibility, the growth of anchorage-dependent mammalian cells on spider silk-elastin coated culture plates was compared with conventional coatings such as collagen, fibronectin and poly-D-lysine. The anchorage-dependent chondrocytes showed similar growth behaviour and a rounded phenotype on collagen and on spider silk-elastin coated plates and the proliferation was remarkably superior to untreated polystyrene plates.     

Predator modulation of plant pathogen spread through induced changes in vector development rates

1. Predation on vectors of pathogens can indirectly influence infection spread. In addition to the consumptive aspect of predation, non-consumptive, predator-induced changes in various vector traits can lead to trait-mediated indirect effects on pathogen spread, potentially operating in various directions and magnitudes. 2. A widespread non-consumptive effect of predation is the alteration of individual prey development rates. Yet, the implications of this phenomenon for the spread of vector-borne plant pathogens have not been studied. It is hypothesized that the epidemiological effects of predator-induced changes in vector development rate depend on the pattern in which the transmission biology of the vector changes along its ontogeny. 3. A general epidemiological model was developed that considers the role of predation in the infection dynamics of a plant pathogen, while incorporating vector stage structure to allow for variation in its development rate. 4. By contrasting scenarios that represent typical plant disease systems, this study confirms that the magnitude of the effect of altered development rate on infection prevalence depends on the disparity between juvenile and adult vectors in their pathogen transmission potential. 5. The model also reveals that the effect of predator-induced change in development rate can impact pathogen spread counterintuitively. Specifically, slowing down vector development can result in increased pathogen prevalence due to apparent competition between infected and uninfected vector populations. 6. More detailed, stage-specific studies of non-consumptive predator effects on vectors are likely to advance our understanding of plant disease ecology, and the development of more effective biological control practices in agriculture.     

蜘蛛拖丝蛋白基因的构建及在大肠杆菌中的表达

蜘蛛大壶腹线产生的拖丝是非常优良的纤维蛋白, 具有独特的强度和弹性。基于拖丝蛋白高度重复序列和部分cDNA序列, 合成蜘蛛拖丝蛋白基因单体, 通过头尾相连的构建策略, 得到拖丝蛋白多聚体, 与原核高效表达载体pET30a(+)连接, 转化大肠杆菌BLR(DE3), 用IPTG诱导表达。 表达产物经His.Bind树脂金属螯合亲和层析一步纯化, 纯度达90%以上, 表达量为20mg/L。SDS-PAGE和蛋白质印迹图谱显示表达产物分子量为37kD, 其值与氨基酸组分分析结果与理论推算值基本符合。 