Distributed biological computation with multicellular engineered networks

Ongoing efforts within synthetic and systems biology have been directed towards the building of artificial computational devices using engineered biological units as basic building blocks. Such efforts, inspired in the standard design of electronic circuits, are limited by the difficulties arising from wiring the basic computational units (logic gates) through the appropriate connections, each one to be implemented by a different molecule. Here, we show that there is a logically different form of implementing complex Boolean logic computations that reduces wiring constraints thanks to a redundant distribution of the desired output among engineered cells. A practical implementation is presented using a library of engineered yeast cells, which can be combined in multiple ways. Each construct defines a logic function and combining cells and their connections allow building more complex synthetic devices. As a proof of principle, we have implemented many logic functions by using just a few engineered cells. Of note, small modifications and combination of those cells allowed for implementing more complex circuits such as a multiplexer or a 1-bit adder with carry, showing the great potential for re-utilization of small parts of the circuit. Our results support the approach of using cellular consortia as an efficient way of engineering complex tasks not easily solvable using single-cell implementations.     

Mal (MyD88-adapter-like) is required for Toll-like receptor-4 signal transduction.

The recognition of microbial pathogens by the innate immune system involves Toll-like receptors (TLRs), which recognize pathogen-associated molecular patterns. Different TLRs recognize different pathogen-associated molecular patterns, with TLR-4 mediating the response to lipopolysaccharide from Gram-negative bacteria. All TLRs have a Toll/IL-1 receptor (TIR) domain, which is responsible for signal transduction. MyD88 is one such protein that contains a TIR domain. It acts as an adapter, being involved in TLR-2, TLR-4 and TLR-9 signalling; however, our understanding of how TLR-4 signals is incomplete. Here we describe a protein, Mal (MyD88-adapter-like), which joins MyD88 as a cytoplasmic TIR-domain-containing protein in the human genome. Mal activates NF-kappaB, Jun amino-terminal kinase and extracellular signal-regulated kinase-1 and -2. Mal can form homodimers and can also form heterodimers with MyD88. Activation of NF-kappaB by Mal requires IRAK-2, but not IRAK, whereas MyD88 requires both IRAKs. Mal associates with IRAK-2 by means of its TIR domain. A dominant negative form of Mal inhibits NF-kappaB, which is activated by TLR-4 or lipopolysaccharide, but it does not inhibit NF-kappaB activation by IL-1RI or IL-18R. Mal associates with TLR-4. Mal is therefore an adapter in TLR-4 signal transduction.     

Altruism and social cheating in the social amoeba Dictyostelium discoideum

The social amoeba, Dictyostelium discoideum, is widely used as a simple model organism for multicellular development, but its multicellular fruiting stage is really a society. Most of the time, D. discoideum lives as haploid, free-living, amoeboid cells that divide asexually. When starved, 10(4)-10(5) of these cells aggregate into a slug. The anterior 20% of the slug altruistically differentiates into a non-viable stalk, supporting the remaining cells, most of which become viable spores. If aggregating cells come from multiple clones, there should be selection for clones to exploit other clones by contributing less than their proportional share to the sterile stalk. Here we use microsatellite markers to show that different clones collected from a field population readily mix to form chimaeras. Half of the chimaeric mixtures show a clear cheater and victim. Thus, unlike the clonal and highly cooperative development of most multicellular organisms, the development of D. discoideum is partly competitive, with conflicts of interests among cells. These conflicts complicate the use of D. discoideum as a model for some aspects of development, but they make it highly attractive as a model system for social evolution.     

采用短时傅里叶变换的铁路车载探地雷达数据解译方法

为了更好地实现探地雷达( GPR)在铁路路基检测中的应用,寻求一种用于识别路基不同病害特征的有效方法,克服传统人工解泽方法的低效率和主观性,选取大同一准格尔铁路线中典型的雷达实测图像,采用短时傅里叶变换(STFT)方法,通过对信号进行时域和频域分析,确定合适的窗函数类型和长度,得到一组可用于对不同铁路路基病害进行分类的时频域图像,分析表明,图中不同深度处的能量分布特性与实际路基情况相符.然后,通过对图像形态特征的分析,得到一组典型的特征参数值,研究了不同目标信号在该特征参数空间的聚类特征,并将STFT方法与Welch功率谱估计法、时域局部能量法的计算结果进行了对比分析,结果证明STFT方法具有优越性,可以作为GPR数据分类的基础.     

Quenching quorum-sensing-dependent bacterial infection by an N-acyl homoserine lactonase.

Bacterial cells sense their population density through a sophisticated cell-cell communication system and trigger expression of particular genes when the density reaches a threshold. This type of gene regulation, which controls diverse biological functions including virulence, is known as quorum sensing. Quorum-sensing signals, such as acyl-homoserine lactones (AHLs), are the essential components of the communication system. AHLs regulate virulence gene expression in a range of plant and animal (including human) bacterial pathogens. AHL-producing tobacco restored the pathogenicity of an AHL-negative mutant of Erwinia carotovora. Different bacterial species may produce different AHLs, which vary in the length and substitution of the acyl chain but contain the same homoserine lactone moiety. Here we show that the acyl-homoserine lactonase (AHL-lactonase), a new enzyme from Bacillus sp., inactivates AHL activity by hydrolysing the lactone bond of AHLs. Plants expressing AHL-lactonase quenched pathogen quorum-sensing signalling and showed significantly enhanced resistance to E. carotovora infection. Our results highlight a promising potential to use quorum-sensing signals as molecular targets for disease control, thereby broadening current approaches for prevention of bacterial infections.     

细胞编程性死亡及其生物学意义

细胞编程性死亡(英文编写为PCD)是多细胞生物的自然属性,它按照一定的基因编码在时空上顺次表达使死亡细胞呈现一定的表观现象及生理生化特征。PCD受来自机体内其它细胞信号等内源性因素和药物、物理等外源性因素的调节。PCD是一种主动死亡方式且表现为单细胞性,与病理因素作用下的坏死不同,但又有一定联系。PCD在动、植物体的生长发育和成体新陈代谢中有重要的调节作用,PCD异常是某些疾病发病的重要原因     

Evolution of novel cooperative swarming in the bacterium Myxococcus xanthus

Cooperation among individuals is necessary for evolutionary transitions to higher levels of biological organization. In such transitions, groups of individuals at one level (such as single cells) cooperate to form selective units at a higher level (such as multicellular organisms). Though the evolution of cooperation is difficult to observe directly in higher eukaryotes, microorganisms do offer such an opportunity. Here we report the evolution of novel cooperative behaviour in experimental lineages of the bacterium Myxococcus xanthus. Wild-type strains of M. xanthus exhibit socially dependent swarming across soft surfaces by a mechanism known as 'S-motility' that requires the presence of extracellular type IV pili. In lineages of M. xanthus unable to make pili, a new mechanistic basis for cooperative swarming evolved. Evolved swarming is mediated, at least in part, by enhanced production of an extracellular fibril matrix that binds cells-and their evolutionary interests-together. Though costly to individuals, fibril production greatly enhanced population expansion in groups of interconnected cells. These results show that fundamental transitions to primitive cooperation can readily occur in bacteria.     

人工合成非肽类Caspase-3抑制剂的研究进展

半胱氨酸蛋白酶（Caspase）家族是代表着一类细胞内的蛋白酶系统,在介导细胞凋亡扮演着重要的角色[1].细胞凋亡的发生是一个复杂Caspase家族引导的蛋白酶级联反应过程,尽管对于不同的细胞或不同信号传导途径诱发的凋亡过程中参与的Caspase有所不同,但Caspase-3是细胞凋亡蛋白酶级联反应的必经之路,也是凋亡的关键酶和执行者.而由Caspase调控的细胞凋亡的不正常激活是引起人体机能紊乱的一些疾病的主要根源,例如肿瘤、自身免疫性疾病、病毒性感染以及各种神经退行性疾病等.所以针对Caspases-3的抑制剂将可能是上述疾病的一种非常有效的治疗药物.由于天然的Caspase抑制剂和人工合成肽类Caspase抑制剂在特异性,透膜性,体内稳定性和活性等方面的不足,人们便开始了对人工合成非肽类抑制剂的研究.本文对近年来人工合成非肽类Caspase-3抑制剂的研究进展情况作一综述.     

Rewiring cellular morphology pathways with synthetic guanine nucleotide exchange factors

Eukaryotic cells mobilize the actin cytoskeleton to generate a remarkable diversity of morphological behaviours, including motility, phagocytosis and cytokinesis. Much of this diversity is mediated by guanine nucleotide exchange factors (GEFs) that activate Rho family GTPases-the master regulators of the actin cytoskeleton. There are over 80 Rho GEFs in the human genome (compared to only 22 genes for the Rho GTPases themselves), and the evolution of new and diverse GEFs is thought to provide a mechanism for linking the core cytoskeletal machinery to a wide range of new control inputs. Here we test this hypothesis and ask if we can systematically reprogramme cellular morphology by engineering synthetic GEF proteins. We focused on Dbl family Rho GEFs, which have a highly modular structure common to many signalling proteins: they contain a catalytic Dbl homology (DH) domain linked to diverse regulatory domains, many of which autoinhibit GEF activity. Here we show that by recombining catalytic GEF domains with new regulatory modules, we can generate synthetic GEFs that are activated by non-native inputs. We have used these synthetic GEFs to reprogramme cellular behaviour in diverse ways. The GEFs can be used to link specific cytoskeletal responses to normally unrelated upstream signalling pathways. In addition, multiple synthetic GEFs can be linked as components in series to form an artificial cascade with improved signal processing behaviour. These results show the high degree of evolutionary plasticity of this important family of modular signalling proteins, and indicate that it may be possible to use synthetic biology approaches to manipulate the complex spatio-temporal control of cell morphology.     

Transformation of cell adhesion properties by exogenously introduced E-cadherin cDNA

E-cadherin is a cell surface glycoprotein responsible for Ca2+-dependent intercellular adhesion between epithelial cells; it is also called uvomorulin, L-CAM (ref. 3), cell-CAM 120/80 (ref.4) or Arc-1 (ref. 5). Because blocking the action of E-cadherin by monoclonal antibodies causes dispersion of compact cell colonies, this molecule is thought to be an important factor for maintenance of multicellular systems. To demonstrate directly that E-cadherin is involved in cell-cell adhesion, we cloned full-length cDNA encoding E-cadherin from F9 cells and introduced it into L fibroblasts deficient in E-cadherin. These L cells acquire strong Ca2+-dependent aggregating activity by expressing the E-cadherin derived from the introduced cDNA and were morphologically transformed so as to form colonies in which cells were tightly connected to each other.     

Terrain imaging using a SAR system based on reflected GPS signals

This paper describes a 3D multi-static synthetic aperture radar (SAR) imaging system which utilises reflected GPS signals from moving objects on the Earth's surface. The principle of bi-static radar is used to model the reflected GPS signals. The movement of a visible GPS satellite serves as a base for a synthetic aperture over an observation time period. As an example, a MATLAB simulation has been carried out in order to detect the movement of imaged object sunder the assumption of one static GPS receiver with two targets which move with different speeds. The influence of the visible satellite'sposition and velocity on the spatial resolution of such a SAR system isdiscussed. Simulation results show that by measuring the cross-correlation of the reflected GPS signal from the terrain and objects on it,the detection of the objects can enjoy a good spatial resolution for thecase of moving objects and a moving GPS receiver. Furthermore, thespatial resolution is also related to the selection of visible GPS satelliteswith respect to their azimuths, elevations and velocities. This systemhas the following useful features: (a) no dedicated signal transmitter is required; (b) the GPS signal frequency is reused; (c) GPS operates round-the-clock and its signals cover the entire Earth's surface; (d) low power consumption; and (e) known GPS signal structure.     

新生儿坏死性小肠结肠炎中细胞死亡机制的研究进展

坏死性小肠结肠炎(Necrotizing Enterocolitis, NEC)是由多因素作用导致的肠道急性炎症性疾病,是早产儿主要的死亡原因之一。以往的研究认为,细胞凋亡是NEC中肠上皮细胞最主要的死亡形式。但近年来的研究发现,程序性坏死(Necroptosis)、细胞焦亡(Pyroptosis)及铁死亡(Ferroptosis)等非凋亡形式的程序性细胞死亡(Programmed Cell Death, PCD)也可能参与到NEC的发病机制中,不同形式的细胞死亡的信号调节通路不同,可能会相互影响或存在共同的调节机制如细胞广泛凋亡小体(PANoptosome)等。本文综述了非凋亡形式的不同类型程序性细胞死亡方式及其信号调节通路,以及其在NEC中的作用机制,并提出NEC诊断生物标志物或防治的新靶点,以期为临床NEC的预防和管理提供思路。     

Bispecific antibody and its clinical applications in cancer

Bispecific antibody (BsAb) usually consists of two different antigen-binding arms, by which it is capable of simultaneously binding to target cells and effector cells, and can directly mediate the killing of target cells by retargeting and activating effector cells. The development of BsAb research goes through three main stages: chemical cross- linking of murine-derived monoclonal antibody, hybrid hybridomas and engineered BsAb. Among them, engineered BsAb has more formats than the other two, such as diabody, ScdHLX, ScZip, ScCH3, ScFab and BsIgG, etc. Compared with former murine-derived BsAbs, engineered BsAb has lower immunogenicity and stronger penetrating capacity, and currently, some of them appear suitable for clinical application in yields and qualities. Up to now, several phase Ⅰand phase Ⅱ clinical studies of BsAb, for instance, some (Fab’)₂ and Diabodies, have been performed. Among those BsAbs, anti-CD3/anti-tumor BsAbs is most common, they not only can activate T cell and induce CD3AK cytotoxic activity in in vitro experiment, and inhibit the growth of tumor on tumor-bearing mouse by retargeting T cells to lyse tumor cells, but also offer great promise in the therapy of some malignancies in clinic, especially of some advanced cancers as well as elimination of minimal residual tumors, indicated by increasing the tumor/blood ratio of antibody in patients and improving the natural killer cell (NK) anti-tumor activity in tumor sites, and also presenting of an increase level in TNF-α,INF-γ, IL-6, IL-8, IL-10 and soluble CD25, etc. The responses are also shown via improving the quality of life and prolonging the survival of partial patients. The “Knobs into Holes” technology is a new strategy emerging during research on engineered BsAb, it is likely to be useful for heterodimerization and can improve the quantity, purity and stability of BsAb, it is also anticipated to increase the clinical potential of BsAb in the future.     

基于小波的AWJ表面拓扑结构特征分析

为了详细地研究水切割-AWJ（abrasive water jet）表面的主要特征,本文分别研究了由传统接触式测量仪和光学测量仪获得的信号。通过对信号的小波分解和重构,提取了形态误差、波纹度以及粗糙度等表面特征。另外,本文还详细地论述了小波基函数的选择以及小波分解层次确定等基本的应用问题。实验结果表明小波能够有效地应用于AWJ表面拓扑结构特征的分析和描述。     

基因芯片研究小檗碱对胰岛素抵抗卵巢颗粒细胞基因表达的影响

应用基因芯片技术研究小檗碱对胰岛素抵抗卵巢颗粒细胞基因表达谱的影响。方法采用胰岛素信号转导通路调节糖代谢的关键分子——PI-3K的特异性阻断剂沃漫青霉素作用于猪卵巢颗粒细胞,人工诱导胰岛素抵抗(IR)的细胞模型;同时应用小檗碱作用在细胞模型上,作用48h后提取细胞总RNA,应用猪全基因组单通道芯片对样本基因进行筛查,得出差异表达基因数据并进行分析。结果显示,中药小檗碱作用模型细胞后差异表达基因42个。差异表达基因主要涉及的功能有:物质代谢、炎症与免疫反应、信号转导等。由此可得出结论:小檗碱可通过多种途径在基因水平上对胰岛素抵抗卵巢颗粒细胞起作用。     

Two distinct modes of guidance signalling during collective migration of border cells

Although directed migration is a feature of both individual cells and cell groups, guided migration has been studied most extensively for single cells in simple environments. Collective guidance of cell groups remains poorly understood, despite its relevance for development and metastasis. Neural crest cells and neuronal precursors migrate as loosely organized streams of individual cells, whereas cells of the fish lateral line, Drosophila tracheal tubes and border-cell clusters migrate as more coherent groups. Here we use Drosophila border cells to examine how collective guidance is performed. We report that border cells migrate in two phases using distinct mechanisms. Genetic analysis combined with live imaging shows that polarized cell behaviour is critical for the initial phase of migration, whereas dynamic collective behaviour dominates later. PDGF- and VEGF-related receptor and epidermal growth factor receptor act in both phases, but use different effector pathways in each. The myoblast city (Mbc, also known as DOCK180) and engulfment and cell motility (ELMO, also known as Ced-12) pathway is required for the early phase, in which guidance depends on subcellular localization of signalling within a leading cell. During the later phase, mitogen-activated protein kinase and phospholipase Cgamma are used redundantly, and we find that the cluster makes use of the difference in signal levels between cells to guide migration. Thus, information processing at the multicellular level is used to guide collective behaviour of a cell group.     

An autoregulatory circuit for long-range self-organization in Dictyostelium cell populations

Nutrient-deprived Dictyostelium amoebae aggregate to form a multicellular structure by chemotaxis, moving towards propagating waves of cyclic AMP that are relayed from cell to cell. Organizing centres are not formed by founder cells, but are dynamic entities consisting of cores of outwardly rotating spiral waves that self-organize in a homogeneous cell population. Spiral waves are ubiquitously observed in chemical reactions as well as in biological systems. Although feedback control of spiral waves in spatially extended chemical reactions has been demonstrated in recent years, the mechanism by which control is achieved in living systems is unknown. Here we show that mutants of the cyclic AMP/protein kinase A pathway show periodic signalling, but fail to organize coherent long-range wave territories, owing to the appearance of numerous spiral cores. A theoretical model suggests that autoregulation of cell excitability mediated by protein kinase A acts to optimize the number of signalling centres.     

Membrane currents that govern smooth muscle contraction in a ctenophore

Ctenophores are transparent marine organisms that swim by means of beating cilia; they are the simplest animals with individual muscle fibres. Predatory species, such as Beroe ovata, have particularly well-developed muscles and are capable of an elaborate feeding response. When Beroe contacts its prey, the mouth opens, the body shortens, the pharynx expands, the prey is engulfed and the lips then close tightly. How this sequence, which lasts 1 s, is accomplished is unclear. The muscles concerned are structurally uniform and are innervated at each end by a neuronal nerve net with no centre for coordination. Isolated muscle cells studied under voltage-clamp provide a solution to this puzzle. We find that different groups of muscle cells have different time-dependent membrane currents. Because muscle contraction depends upon calcium entry during each action potential, these different currents produce different patterns of contraction. We conclude that in a simple animal such as a ctenophore, a sophisticated set of membrane conductances can compensate for the absence of an elaborate system of effectors.     

Autophagy in immunity and inflammation

Autophagy is an essential, homeostatic process by which cells break down their own components. Perhaps the most primordial function of this lysosomal degradation pathway is adaptation to nutrient deprivation. However, in complex multicellular organisms, the core molecular machinery of autophagy - the 'autophagy proteins' - orchestrates diverse aspects of cellular and organismal responses to other dangerous stimuli such as infection. Recent developments reveal a crucial role for the autophagy pathway and proteins in immunity and inflammation. They balance the beneficial and detrimental effects of immunity and inflammation, and thereby may protect against infectious, autoimmune and inflammatory diseases.     

Revealing structure and assembly cues for Arabidopsis root-inhabiting bacterial microbiota

The plant root defines the interface between a multicellular eukaryote and soil, one of the richest microbial ecosystems on Earth. Notably, soil bacteria are able to multiply inside roots as benign endophytes and modulate plant growth and development, with implications ranging from enhanced crop productivity to phytoremediation. Endophytic colonization represents an apparent paradox of plant innate immunity because plant cells can detect an array of microbe-associated molecular patterns (also known as MAMPs) to initiate immune responses to terminate microbial multiplication. Several studies attempted to describe the structure of bacterial root endophytes; however, different sampling protocols and low-resolution profiling methods make it difficult to infer general principles. Here we describe methodology to characterize and compare soil- and root-inhabiting bacterial communities, which reveals not only a function for metabolically active plant cells but also for inert cell-wall features in the selection of soil bacteria for host colonization. We show that the roots of Arabidopsis thaliana, grown in different natural soils under controlled environmental conditions, are preferentially colonized by Proteobacteria, Bacteroidetes and Actinobacteria, and each bacterial phylum is represented by a dominating class or family. Soil type defines the composition of root-inhabiting bacterial communities and host genotype determines their ribotype profiles to a limited extent. The identification of soil-type-specific members within the root-inhabiting assemblies supports our conclusion that these represent soil-derived root endophytes. Surprisingly, plant cell-wall features of other tested plant species seem to provide a sufficient cue for the assembly of approximately 40% of the Arabidopsis bacterial root-inhabiting microbiota, with a bias for Betaproteobacteria. Thus, this root sub-community may not be Arabidopsis-specific but saprophytic bacteria that would naturally be found on any plant root or plant debris in the tested soils. By contrast, colonization of Arabidopsis roots by members of the Actinobacteria depends on other cues from metabolically active host cells.