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The treatment of herpes zoster (HZ) by the traditional Chinese medicine of acupuncture is attracting attention. However, there is still a controversy about the effectiveness and safety of acupuncture treatment of HZ.Articles on randomized controlled trials examining acupuncture and Western medicine treatments of HZ published since the establishment of the PubMed, Embase, Medline, and Chinese Biomedical Literature (CBM) databases to March 2021 were electronically retrieved. The Cochrane System Evaluation Manual was used for the data analysis with Review Manager 5.3 software, and the Cochrane Handbook version 5.3 systematic review writing manual was adopted to evaluate the risk of bias.In total, 11 articles, comprising 1,156 patients (585 in the experimental group and 571 in the control group) were included in the meta-analysis, and the results showed that the treatments used in the experimental and control groups were significant differences of total treatment efficiency [odds ratio (OR) =6.76; 95% confidence interval (CI): 3.46 to 13.21; P0.05] in terms of the incidence of posterior neuralgia (OR =0.07; 95% CI: 0.02 to 0.21; P0.05), pain-relief time [mean difference (MD) =-2.17; 95% CI: -2.90 to -1.44; P0.05], shingles time (MD =-1.61; 95% CI: -2.84 to -0.38; P0.05), and scabbing time (MD =-1.62; 95% CI: -2.64 to -0.61; P0.05), and patients' visual analogue scale (VAS) pain scores improved [standard MD (SMD) =0.87; 95% CI: 0.01 to 1.73; P=0.05] was no significant difference.Compared to Western medicine treatments, acupuncture had a better effect on HZ, reduced the posterior neuralgia rate of patients, and shortened the course of treatment, but had no obvious effect on the relief of pain.

Objective Fatty acids are important for biological function; however, in excess, they can cause metabolic dysregulation. Methods to image and detect fatty acids in real time are lacking. Therefore, the current study examined the dynamics of fatty acid trafficking and signaling utilizing novel fluorescent and luminescent approaches. Methods We generated fluorescent and luminescent-based genetically-encoded sensors based upon the ligand-dependent interaction between PPARα and SRC-1 to image and detect cellular dynamics of fatty acid trafficking. Results The use of a fluorescent sensor demonstrates that fatty acids traffic rapidly from lipid droplets to the nucleus. Both major lipases ATGL and HSL contribute to fatty acid signaling from lipid droplet to nucleus, however, their dynamics differ. Furthermore, direct activation of lipolysis, independent of receptor-mediated signaling is sufficient to promote lipid droplet to nuclear trafficking of fatty acids. A luminescent-based sensor that reports intracellular fatty acid levels is amenable to high-throughput analysis. Conclusions Fatty acids traffic from lipid droplets to the nucleus within minutes of stimulated lipolysis. Genetically-encoded fluorescent and luminescent based sensors can be used to probe the dynamics of fatty acid trafficking and signaling., Graphical abstract Image 1, Highlights • Genetically-encoded fatty acid sensors report on the temporal and spatial regulation of lipolysis. • A fluorescence-based fatty acid sensor can detect fatty acids within minutes of lipolysis. • Fatty acids dynamically traffic from lipid droplets to the nucleus to modulate gene expression. • A luminescence-based fatty acid sensor is amenable to high-throughput analysis.

Semen Cuscutae, called Tu-si-zi in Chinese, is a kind of dried mature seed in the Convolvulaceae family. It mainly distributes in China, Korea, Pakistan, Vietnam, India and Thailand. It is used as a kidney-tonifying drug for treatment of aging related diseases such as osteoporosis in traditional Chinese medicine. However, the exact mechanisms on bone resorption are poorly studied.The aim of this study was to investigate the potential effect of Semen Cuscutae on ovariectomy (OVX)-induced osteoporosis in mice and clarify the exact mechanisms by which Semen Cuscutae exert the anti-osteoporosis effect.Qualitative and quantitative analyses of Semen Cuscutae were performed by UPLC-Q-TOF-MS and HPLC-MS/MS, respectively. Changes in bone mineral density (BMD) induced by OVX in mice were measured by dual-energy X-ray absorptiometry and micro-computed tomography (μCT). Tartrate-resistant acid phosphatase (TRAP) staining as well as hematoxylin and eosin (HE) staining were used to observe bone microarchitectural changes. ELISA kits were used to assess the therapeutic effects of Semen Cuscutae on the serum levels of osteoprotegerin (OPG), tartrate-resistant acid phosphatase 5b (TRACP-5b), and receptor activator of nuclear factor-κB (RANKL). The effect of Semen Cuscutae on primary cell viability was assessed using CCK-8 and anti-tartrate phosphatase assays. TRAP staining and actin ring staining were used to observe the effect of Semen Cuscutae on osteoclast differentiation. Western blotting was used to measure the effects of Semen Cuscutae on expressions of NFATC1, c-Src kinase, and c-fos.Results from UPLC-Q-TOF-MS showed that the main components of Semen Cuscutae were flavonoid compounds that included quercitrin, quercetin, hyperoside, caffeic acid, rutin, chlorogenic acid, luteolin, apigenin, kaempferol, isoquercetin, cryptochlorogenic acid, isorhamnetin-3-O-glucoside, and astragalin. After the Semen Cuscutae extract was orally administered to OVX mice, bone density increased (P 0.01) and bone microstructure was significantly improved (P 0.01 or 0.05). Additionally, Semen Cuscutae exhibited a significant descending effect in the levels of serum TRACP-5b and RANKL, while there was a significant increase in OPG in the Semen Cuscutae group compared with the OVX group, especially at high doses. Moreover, we found that increasing of c-fos, c-Src kinase, and NFATC1 protein expressions were reversed by Semen Cuscutae in vitro and in vivo.Our results showed that Semen Cuscutae exhibited anti-osteoporosis effects through the c-fos/c-Src kinase/NFATC1 signaling pathway.

Background Mechanisms underlying oral immunotherapy (OIT) are unclear and the effects on immune cells at varying maintenance doses are unknown. Objective We aimed to determine the immunologic changes caused by peanut OIT in preschool aged children and determine the effect on these immune responses in groups ingesting low or high-dose peanut OIT (300 mg or 3000 mg, respectively) as maintenance therapy. Methods Blood was drawn at several time-points throughout the OIT protocol and PBMCs isolated and cultured with peanut antigens. Secreted cytokines were quantified via multiplex assay, whereas Treg and peanut-responsive CD4 T cells were studied with flow cytometry. Basophil activation assays were also conducted. Results Th2-, Th1-, Th9- and Tr1-type cytokines decreased over the course of OIT in groups on high- and low-dose OIT. There were no significant differences detected in cytokine changes between the high- and low-dose groups. The initial increase in both the number of peanut-responsive CD4 T cells and the number of Tregs was transient and no significant differences were found between groups. Basophil activation following peanut stimulation was decreased over the course of OIT and associated with increased peanut-IgG4/IgE ratios. No differences were found between high- and low-dose groups in basophil activation at the time of desensitization or sustained unresponsiveness oral food challenges. Conclusions and clinical relevance Peanut OIT leads to decreases in pro-allergic cytokines, including IL-5, IL-13, and IL-9 and decreased basophil activation. No differences in T cell or basophil responses were found between subjects on low or high-dose maintenance OIT, which has implications for clinical dosing strategies.

In through-wall radar imaging, the wall parameters, including permittivity, wall thickness and conductivity, are of crucial importance for locating targets precisely and obtaining a well-focused image. There are usually ambiguities in practice. To obtain a quick and accurate estimation of wall parameters, a robust approach based on machine learning is introduced. The measurement results demonstrate that the estimation approach has the advantages of high precision and low computational time.

This study developed a method for simultaneous determination of 13 elements of Semen Cuscutae (quercitrin, quercetin, hyperoside, caffeic acid, chlorogenic acid, luteolin, apigenin, kaempferol, isoquercitrin, cryptochlorogenic acid, isorhamnetin-3-O-glucoside, astragalin, and rutin) in rat plasma using high-performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS) in the negative MRM mode. The analytes were analyzed with CORTECS®C18 column (4.6 × 150 mm, 2.7 μm) with mobile phases consisting of 0.1% formic acid in water (A) and acetonitrile (B). The intra- and interday precision of the target compounds were expressed as relative standard deviation (RSD) in the range of 0.5%–10.4%, and the accuracy of the target compounds was expressed as relative error (RE) not exceeding ±14.5% for all analytes. In the meantime, the extraction recovery of the target compounds in plasma samples ranged from 87.4% to 106.2% and matrix effect from 81.0% to 115.5%. The established method was successfully accomplished for the pharmacokinetic study of the analytes in rat plasma samples following oral administration of Semen Cuscutae extract, and the pharmacokinetic parameters of seven compounds were obtained.

The estimation of the wall parameters is important in through-the-wall radar imaging TWRI. Ambiguities in the wall characteristics, including wall thickness, permittivity, and conductivity, will distort the imaging and shift the target position. To obtain a quick and accurate estimation of wall parameters, an efficient method based on machine learning is proposed. The estimation problem is converted to a regression problem. A map between wall parameters and the received signals is established and is regressed as a linear formulation after machine learning; in this manner, the wall parameters can be estimated in few seconds. The measurement results demonstrate that the estimated approach has the advantages of high precision and low computational time. The influence of the size, the location, the number of the targets and the length of the wall, the sampling interval, and noise on the estimation problems is discussed, and the image entropy is given to verify the effectiveness of the estimation values. The results based on support vector machines and least-square support vector machines LS-SVMs, which are both machine-learning approaches, are compared. The comparison results reveal that the LS-SVM-based method can provide comparable performances in terms of accuracy and convenience but poor performances in terms of generalization and robustness.

In this article, a double-metallic-layered Huygens' surface Huygens' surface that efficiently refracts normally incident electromagnetic waves at the telecommunication frequency of 25 GHz is reported. The phase gradient along the interface is controlled by a series of isotropic, nonresonant, spatial phase shifters with dimensions of 0.5λ0 × 0.5λ0, where λ0 is the free-space wavelength at 25 GHz. The electric and magnetic responses of the unit cells are controlled by metallic patterns etched on both sides of an ultrathin dielectric substrate with thickness of 0.07λ0. These surface microstructures are shown to be able to fabricate high-efficiency refract-arrays, ultrathin, wideband focusing lenses. The full-wave simulations demonstrate their excellent performances in manipulating electromagnetic wavefronts. The measured results for the fabricated refract-array further show that it can refract normally incident waves to the predefined angle, and the peak value of the transmission efficiency approaches 76%. © 2016 Wiley Periodicals, Inc. Int J RF and Microwave CAE 26:449-455, 2016.

Epimedium herb is one of the most vital traditional Chinese medicines (TCMs), which is used for &ldquo, nourishing the kidney and reinforcing the Yang&rdquo, In the guidance of TCM theory, Epimedium herb is usually processed with lamb oil to increase its efficacy. The contents of active ingredients in different Epimedium are significantly varied, which may derive from their different species, regions and processing methods. In this research, 13 batches of raw Epimedium collected from 6 provinces were identified. After optimization of the processing method of Epimedium, a liquid chromatography&ndash, mass spectrometry (LC&ndash, MS/MS) method for simultaneous determination of 16 compounds was established to evaluate the quality of raw and processed. Then the multivariate statistical technique was applied to compare different batches of Epimedium based on the LC&ndash, MS/MS data. As a conclusion, the herbs collected from 6 areas were ascribed to 5 species by microscopic and appearance features. Meanwhile, all of the raw and processed samples were classified by partial least squares discriminant analysis (PLS-DA) based on the 16 analyzed compounds. The comparison results indicate that processing and species both have important influences on Epimedium compositions contents.

Background Epstein–Barr virus (EBV) infection is causatively associated with a variety of human cancers, including gastric cancer (GC), which has one of the highest mortality rates of all human cancers. Long non-coding RNAs (lncRNAs) show important regulatory roles in human GC. SNHG8 is a recently identified lncRNA that was reported to show abnormal expression pattern in GC. However, little is known of its biological function in EBV-associated GC. Methods We used cell viability, colony formation and cell cycle assays to investigate the roles of lncRNA SNHG8 in the cell growth of EBV-associated GC. Results The transcript levels of SNHG8 in the cultured EBV-associated GC cells were significantly higher in the cultured EBV-associated GC cells compared with the levels in normal human gastric mucosal cells and EBV-negative GC cells. Knockdown of SNHG8 with specific shRNAs inhibited cell proliferation and colony formation and arrested the cell cycle in the G0/G1 phase in vitro. We also found that knockdown of SNHG8 suppressed tumor growth in vivo. Conclusions These data indicate the pro-oncogenic potential of SNHG8 in EBV-associated GC, meaning it is a latent therapeutic target for the treatment of this type of cancer.

Perilipin (PLIN) proteins constitute an ancient family important in lipid droplet (LD) formation and triglyceride metabolism. We identified an additional PLIN clade (plin6) that is unique to teleosts and can be traced to the two whole genome duplications that occurred early in vertebrate evolution. Plin6 is highly expressed in skin xanthophores, which mediate red/yellow pigmentation and trafficking, but not in tissues associated with lipid metabolism. Biochemical and immunochemical analyses demonstrate that zebrafish Plin6 protein targets the surface of pigment-containing carotenoid droplets (CD). Protein kinase A (PKA) activation, which mediates CD dispersion in xanthophores, phosphorylates Plin6 on conserved residues. Knockout of plin6 in zebrafish severely impairs the ability of CD to concentrate carotenoids and prevents tight clustering of CD within carotenoid bodies. Ultrastructural and functional analyses indicate that LD and CD are homologous structures, and that Plin6 was functionalized early in vertebrate evolution for concentrating and trafficking pigment. DOI: http://dx.doi.org/10.7554/eLife.21771.001

Alpha-beta hydrolase domain-containing 5 (ABHD5), the defective gene in human Chanarin-Dorfman syndrome, is a highly conserved regulator of adipose triglyceride lipase (ATGL)-mediated lipolysis that plays important roles in metabolism, tumor progression, viral replication, and skin barrier formation. The structural determinants of ABHD5 lipolysis activation, however, are unknown. We performed comparative evolutionary analysis and structural modeling of ABHD5 and ABHD4, a functionally distinct paralog that diverged from ABHD5 ~500 million years ago, to identify determinants of ABHD5 lipolysis activation. Two highly conserved ABHD5 amino acids (R299 and G328) enabled ABHD4 (ABHD4 N303R/S332G) to activate ATGL in Cos7 cells, brown adipocytes, and artificial lipid droplets. The corresponding ABHD5 mutations (ABHD5 R299N and ABHD5 G328S) selectively disrupted lipolysis without affecting ATGL lipid droplet translocation or ABHD5 interactions with perilipin proteins and ABHD5 ligands, demonstrating that ABHD5 lipase activation could be dissociated from its other functions. Structural modeling placed ABHD5 R299/G328 and R303/G332 from gain-of-function ABHD4 in close proximity on the ABHD protein surface, indicating they form part of a novel functional surface required for lipase activation. These data demonstrate distinct ABHD5 functional properties and provide new insights into the functional evolution of ABHD family members and the structural basis of lipase regulation.

In order to solve the undetected probability of multiple targets in ultra-wideband (UWB) through-the-wall radar imaging (TWRI), a time-delay and amplitude modified back projection (BP) algorithm is proposed. The refraction point is found by Fermat`s principle in the presence of a wall, and the time-delay is correctly compensated. On this basis, transmission loss of the electromagnetic wave, the absorption loss of the refraction wave, and the diffusion loss of the spherical wave are analyzed in detail. Amplitude compensation is deduced and tested on a model with a single-layer wall. The simulating results by finite difference time domain (FDTD) show that it is effective in increasing the scattering intensity of the targets behind the wall. Compensation for the diffusion loss in the spherical wave also plays a main role. Additionally, the two-layer wall model is simulated. Then, the calculating time and the imaging quality are compared between a singlelayer wall model and a two-layer wall model. The results illustrate the performance of the time-delay and amplitude-modified BP algorithm with multiple targets and multiple-layer walls of UWB TWRI.

Cyclin J (CycJ) is a poorly characterized member of the Cyclin superfamily of cyclin-dependent kinase regulators, many of which regulate the cell cycle or transcription. Although CycJ is conserved in metazoans its cellular function has not been identified and no mutant defects have been described. In Drosophila, CycJ transcript is present primarily in ovaries and very early embryos, suggesting a role in one or both of these tissues. The CycJ gene (CycJ) lies immediately downstream of armitage (armi), a gene involved in the Piwi-associated RNA (piRNA) pathways that are required for silencing transposons in the germline and adjacent somatic cells. Mutations in armi result in oogenesis defects but a role for CycJ in oogenesis has not been defined. Here we assessed oogenesis in CycJ mutants in the presence or absence of mutations in armi or other piRNA pathway genes. CycJ null ovaries appeared normal, indicating that CycJ is not essential for oogenesis under normal conditions. In contrast, armi null ovaries produced only two egg chambers per ovariole and the eggs had severe axis specification defects, as observed previously for armi and other piRNA pathway mutants. Surprisingly, the CycJ armi double mutant failed to produce any mature eggs. The double null ovaries generally had only one egg chamber per ovariole and the egg chambers frequently contained an overabundance of differentiated germline cells. Production of these compound egg chambers could be suppressed with CycJ transgenes but not with mutations in the checkpoint gene mnk, which suppress oogenesis defects in armi mutants. The CycJ null showed similar genetic interactions with the germline and somatic piRNA pathway gene piwi, and to a lesser extent with aubergine (aub), a member of the germline-specific piRNA pathway. The strong genetic interactions between CycJ and piRNA pathway genes reveal a role for CycJ in early oogenesis. Our results suggest that CycJ is required to regulate egg chamber production or maturation when piRNA pathways are compromised.

An electromagnetic model is simulated using finite difference time domain for through-the-wall radar imaging. In the numerical simulation narrow-, wide-, and ultra-wide-band signals are used as emission sources. The back projection algorithm is applied to image square and rectangular ideal metal bodies based on a refraction model. The results demonstrate that there is obvious refraction in the through-the-wall propagation of electromagnetic waves. For the calculation of propagation delay, the variation in propagation path and time must be taken into account. Then the actual refractive point is found accurately and rapidly according to Fermat's principle. For imaging, all three signal sources can be used to acquire the image. The image result is best for the ultra-wide band and worst for the narrow- wide band. For the azimuth, synthetic aperture technique is adopted to achieve high resolution. For the range, there is a positive proportional relationship between the resolution and signal bandwidth. The wide band signal significantly increases the range resolution. Therefore, ultra-wide band SAR radar has more precise imaging and positioning capabilities.

Frequency selective surfaces (FSSs) are planar structures composed of a periodic array of unit cells that filter electromagnetic (EM) waves impinging on the surfaces. They have been widely used in stealth technology, electromagnetic countermeasure, and communication systems [1, 2]. Typical FSSs are composed of metallic grids or dipole arrays, which exploit the strong interactions between EM waves and metals. However, metals are problematic at very high incident power. Strong currents in the conductors will incur severe heating due to ohmic losses; arcing will occur at field concentration points [3]. Besides, metal materials are not quite favorable for high-temperature resistance and corrosion-resistant requirements. Consider that dielectrics exhibit a much weaker interaction with the field than metals do, all-dielectric FSSs (ADFSSs) make better alternative. Recently, J. H. Barton et al. [4] developed an ADFSS based on guided-mode resonance (GMR), which extended the fractional bandwidth of conventional GMR devices from 1% to 16%, and could survive with no damage at a high pulsed microwave power of 1.7GW/m2. In 2015, they presented another novel ADFSS using genetic algorithms and fast Fourier transforms [5], which exhibited a larger fractional bandwidth. Both of the designs have a total height about λ 0 /3 (λ 0 is the center wavelength of the operating EM waves). In optical region, subwavelength dielectric granules that possess strong Mie resonances are often used to fabricate metamaterials. Recently, a single-negative all-dielectric metamaterial, which is comprised of a single layer of cylindrical silicon resonators on a silicon-on-insulator substrate, is experimentally proved to possess broadband perfect reflection in the infrared region [6].

Maps representing the binary interactions among proteins have become valuable tools for understanding how proteins work together to mediate biological processes. One of the most effective methods for detecting biologically important protein interactions has been the yeast two-hybrid system. Here we present an efficient two-hybrid strategy to facilitate construction of protein interaction maps on a genome-wide scale. The strategy begins with two arrays of yeast expressing known proteins fused to either a DNA binding domain (BD), or a transcription activation domain (AD). The fusion proteins are conditionally expressed using regulated promoters that can be repressed during construction and amplification of the yeast arrays. Interaction assays are conducted in two phases. In the first phase, small pools of AD strains are mated with the array of BD strains. In the second phase, individual BD strains are mated with appropriate subsets of the AD array corresponding to positive pools in the first phase. This strategy has several advantages over previously described approaches, including the ability to detect interactions with proteins that inhibit yeast growth or that activate transcription as BD fusions. Moreover, by minimizing the number of mating operations and sequencing reactions needed to test large sets of binary interactions, this strategy is more efficient than either matrix or library screening approaches. We also present a three-dimensional pooling scheme to further increase the efficiency of large-scale two-hybrid analyses.

SummaryFat and muscle lipolysis involves functional interactions of adipose triglyceride lipase (ATGL), α-β hydrolase domain-containing protein 5 (ABHD5), and tissue-specific perilipins 1 and 5 (PLIN1 and PLIN5). ABHD5 potently activates ATGL, but this lipase-promoting activity is suppressed when ABHD5 is bound to PLIN proteins on lipid droplets. In adipocytes, protein kinase A (PKA) phosphorylation of PLIN1 rapidly releases ABHD5 to activate ATGL, but mechanisms for rapid regulation of PLIN5-ABHD5 interaction in muscle are unknown. Here, we identify synthetic ligands that release ABHD5 from PLIN1 or PLIN5 without PKA activation and rapidly activate adipocyte and muscle lipolysis. Molecular imaging and affinity probe labeling demonstrated that ABHD5 is directly targeted by these synthetic ligands and additionally revealed that ABHD5-PLIN interactions are regulated by endogenous ligands, including long-chain acyl-CoA. Our results reveal a new locus of lipolysis control and suggest ABHD5 ligands might be developed into novel therapeutics that directly promote fat catabolism.

We investigated if complement component 1 inhibitor-like (C1-INH-L) protein found on human spermatozoa exists on mouse spermatozoa and is relevant to reproduction. We used Western blot analysis and immunofluorescence assays to detect C1-INH on murine sperm and tested the effects of the antibodies to C1-INH and albumin (negative control) on in vitro mouse sperm motility and fertilization. C1-INH-L, with molecular weight similar to human C1-INH (100 kDa), was present on the surface of spermatozoan head and midpiece. Treating mouse sperm with anti-C1-INH reduced the mouse sperm motility (P < 0.01), in vitro fertilization (P < 0.01) and embryo development rates (P < 0.01). Anti-albumin failed to do so. We conclude that C1-inhibitor-like protein is present on mouse sperm surface and appears to be relevant to reproduction.

Cytosolic caspase-3-like proteases, such as caspase-3 and caspase-7, have a central role in mediating the progress of apoptosis. Here to conveniently monitor caspase-3-like activity in the multicellular environment, we have developed genetically encoded switch-on fluorescence-base indicators that are cyclized chimeras containing a caspase-3 cleavage site as a switch. When cleaved by caspase-3-like proteases, the non-fluorescent indicator rapidly becomes fluorescent, and thus detects in real-time the activation of such caspases. We generate cultured cells constitutively expressing these chimeras, and all the healthy cells are non-fluorescent. When these cells are exposed to apoptotic stimuli, dead cells show strong fluorescence depending on caspase activation. With these tools, we monitor in real-time caspase-3-like activity in each cell under various conditions, and show for the first time that the environment of cancer cells affects their sensitivity to chemotherapeutic drugs in a modified soft agar assay. These biosensors should enable better understanding of the biological relevance of caspase-3-like proteases.

Hereditary angioedema afflicts patients with unpredictable episodes of swelling that can be life threatening. Treatments approved by the Food and Drug Administration for routine prophylaxis include danazol given orally and the nanofiltered human C1 esterase inhibitor, CINRYZE™, which is approved for intravenous administration. Approved for the treatment of acute attacks are the C1 esterase inhibitor, Berinert®, given intravenously, and the kallikrein inhibitor, KALBITOR®, given subcutaneously. C1 inhibitor has generally been non-toxic and neither pro-inflammatory nor pro-fibrotic, suggesting that it may be suitable for subcutaneous infusion. The current study used a swine model to compare blood levels of human C1 inhibitor following intravenous and subcutaneous infusion, and the effect of infusion route on heart and skin pathology. Levels of C1 inhibitor achieved with SC infusion compared favorably with levels achieved after IV infusion and were relatively more stable than those after IV infusion. Neither cardiac nor skin toxicity was observed.

'Systematic identification of protein interactions for the bacterium Campylobacter jejuni using high-throughput yeast two-hybrid screens detected interactions for 80% of the organism's proteins., Background Data from large-scale protein interaction screens for humans and model eukaryotes have been invaluable for developing systems-level models of biological processes. Despite this value, only a limited amount of interaction data is available for prokaryotes. Here we report the systematic identification of protein interactions for the bacterium Campylobacter jejuni, a food-borne pathogen and a major cause of gastroenteritis worldwide. Results Using high-throughput yeast two-hybrid screens we detected and reproduced 11,687 interactions. The resulting interaction map includes 80% of the predicted C. jejuni NCTC11168 proteins and places a large number of poorly characterized proteins into networks that provide initial clues about their functions. We used the map to identify a number of conserved subnetworks by comparison to protein networks from Escherichia coli and Saccharomyces cerevisiae. We also demonstrate the value of the interactome data for mapping biological pathways by identifying the C. jejuni chemotaxis pathway. Finally, the interaction map also includes a large subnetwork of putative essential genes that may be used to identify potential new antimicrobial drug targets for C. jejuni and related organisms. Conclusion The C. jejuni protein interaction map is one of the most comprehensive yet determined for a free-living organism and nearly doubles the binary interactions available for the prokaryotic kingdom. This high level of coverage facilitates pathway mapping and function prediction for a large number of C. jejuni proteins as well as orthologous proteins from other organisms. The broad coverage also facilitates cross-species comparisons for the identification of evolutionarily conserved subnetworks of protein interactions.

The ability to express heterologous genes in yeast has become indispensable for many biological research techniques. Expression systems that can be regulated are particularly useful because they allow an experimenter to control the timing and levels of gene expression. Despite their many advantages, however, surprisingly few conditional expression systems are available for yeast. Moreover, of those that have been described, many are not ideal either because they have high background expression levels, low induced levels, or because they require restrictive growth conditions. Here we describe a conditional expression system that takes advantage of the yeast MAL62 promoter (MAL62p), which can be controlled by adding maltose or glucose to the growth medium to induce or repress transcription, respectively. In addition, we use a mating scheme to dramatically increase the dynamic range of expression levels possible. We show that MAL62p background activity can be effectively eliminated by maintaining expression constructs in a mal(-) yeast strain. High-level expression can be induced in diploids formed by mating the mal(-) strain with a MAL(+) strain. A similar mating scheme may be useful for other conditional expression systems as well. Among other uses, this approach should aid high throughput yeast two-hybrid assays, which rely on maintaining large libraries of expression strains, which are eventually mated to conduct assays for protein interactions. We demonstrate a two-hybrid system in which MAL62p is used in conjunction with the yeast GAL1 promoter to independently regulate expression of both hybrid proteins, and to allow detection of interactions involving toxic proteins.

We studied complement 1 inhibitor (C1-INH) as an inhibitor of the alternative complement pathway. C1-INH prevented lysis, induced by the alternative complement pathway, of paroxysmal nocturnal hemoglobinuria (PNH) erythrocytes in human serum. It inhibited the binding of both factors B and C3 to PNH and rabbit erythrocytes and blocked the ability of factor B to restore alternative-pathway function in factor B–depleted serum. C1-INH did not bind to factors B or D but did bind to immobilized C3b and cobra venom factor (CVF), a C3b analogue. C1-INH prevented factor B from binding to CVF-coated beads and dissociated bound factor B from such beads. Factor B and C1-INH showed cross competition in binding to CVF-coated beads. Factor D cleaved factor B into Bb and Ba in the presence of C3b. Cleavage was markedly inhibited when C3b was preincubated with C1-INH. C1-INH inhibited the formation of CVFBb and decreased the C3 cleavage. Removal of C1-INH from serum, in the presence of Mg-EGTA with an anti–C1-INH immunoabsorbant, markedly increased alternative-pathway lysis. C1-INH interacts with C3b to inhibit binding of factor B to C3b. At physiologic concentrations, it is a downregulator of the alternative pathway convertase.