Assessing the Conformational Changes of pb5, the Receptor-binding Protein of Phage T5, upon Binding to Its Escherichia coli Receptor FhuA

Within tailed bacteriophages, interaction of the receptor-binding protein (RBP) with the target cell triggers viral DNA ejection into the host cytoplasm. In the case of phage T5, the RBP pb5 and the receptor FhuA, an outer membrane protein of Escherichia coli, have been identified. Here, we use small angle neutron scattering and electron microscopy to investigate the FhuA-pb5 complex. Specific deuteration of one of the partners allows the complete masking in small angle neutron scattering of the surfactant and unlabeled proteins when the complex is solubilized in the fluorinated surfactant F₆-DigluM. Thus, individual structures within a membrane protein complex can be described. The solution structure of FhuA agrees with its crystal structure; that of pb5 shows an elongated shape. Neither displays significant conformational changes upon interaction. The mechanism of signal transduction within phage T5 thus appears different from that of phages binding cell wall saccharides, for which structural information is available.     

Latitude drives diversification in Madagascar's endemic dry forest rodent Eliurus myoxinus (subfamily Nesomyinae)

Numerous hypotheses have been proposed for the historical processes governing the rich endemism of Madagascar's biodiversity. The ‘watershed model’ suggests that drier climates in the recent geological past have resulted in the contraction of forests around major watersheds, thereby defining areas of endemism. We test whether this hypothesis explains phylogeographical patterns in a dry forest‐dependent rodent, Eliurus myoxinus, an endemic species widely distributed through western Madagascar. We sequenced the mitochondrial cytochrome b locus and nuclear introns of the β‐fibrinogen and the growth hormone receptor genes for E. myoxinus. Using a parametric bootstrapping approach, we tested whether the mitochondrial gene tree data fit expectations of local differentiation given the watershed model. We additionally estimated population differentiation and historical demographic parameters, and reconstructed the spatial history of E. myoxinus to highlight spatial and temporal patterns of differentiation. The data do not support the watershed model as a clear explanation for the genetic patterns of diversity within extant E. myoxinus populations. We find striking patterns of latitudinal genetic structure within western Madagascar, and indicate possible roles for environmental and ecological gradients along this axis in generating phylogeographical diversity. © 2013 The Linnean Society of London, Biological Journal of the Linnean Society, 2013, 110 , 500–517.     

Effect of cooling rate and cryoprotectant concentration on intracellular ice formation of small abalone (Haliotis diversicolor) eggs

The intracellular ice formation (IIF) behavior of Haliotis diversicolor (small abalone) eggs is investigated in this study, in relation to controlling the cooling rate and the concentration of dimethyl sulfoxide (DMSO). The IIF phenomena are monitored under a self-developed thermoelectric cooling (TEC) cryomicroscope system which can achieve accurate temperature control without the use of liquid nitrogen. The accuracy of the isothermal and ramp control is within ±0.5 °C. The IIF results indicate that the IIF of small abalone eggs is well suppressed at cooling rates of 1.5, 3, 7 and 12 °C/min with 2.0, 2.5, 3.0 and 4.0 M DMSO in sea water. As 2.0 M DMSO in sea water is the minimum concentration that has sufficient IIF suppression, it is selected as the suspension solution for the cryopreservation of small abalone eggs in order to consider the solution’s toxicity effect. Moreover, IIF characteristics of the cumulative probability of IIF temperature distribution are shown to be well fitted by the Weibull probabilistic distribution. According to our IIF results and the Weibull distribution parameters, we conclude that cooling at 1.5 °C/min from 20 to −50 °C with 2.0 M DMSO in sea water is more feasible than other combinations of cooling rates and DMSO concentrations in our experiments. Applying this protocol and observing the subsequent osmotic activity, 48.8% of small abalone eggs are osmotically active after thawing. In addition, the higher the cooling rate, the less chance of osmotically active eggs. A separate fertility test experiment, with a cryopreservation protocol of 1.5 °C/min cooling rate and 2.0 M DMSO in sea water, achieves a hatching rate of 23.7%. This study is the first to characterize the IIF behavior of small abalone eggs in regard to the cooling rate and the DMSO concentration. The Weibull probabilistic model fitting in this study is an approach that can be applied by other researchers for effective cryopreservation variability estimation and analysis.     

Synthesis and SAR studies of 5-(pyridin-4-yl)-1,3,4-thiadiazol-2-amine derivatives as potent inhibitors of Bloom helicase

Human cells utilize a variety of complex DNA repair mechanisms in order to combat constant mutagenic and cytotoxic threats from both exogenous and endogenous sources. The RecQ family of DNA helicases, which includes Bloom helicase (BLM), plays an important function in DNA repair by unwinding complementary strands of duplex DNA as well as atypical DNA structures such as Holliday junctions. Mutations of the BLM gene can result in Bloom syndrome, an autosomal recessive disorder associated with cancer predisposition. BLM-deficient cells exhibit increased sensitivity to DNA damaging agents indicating that a selective BLM inhibitor could be useful in potentiating the anticancer activity of these agents. In this work, we describe the medicinal chemistry optimization of the hit molecule following a quantitative high-throughput screen of >355,000 compounds. These efforts lead to the identification of ML216 and related analogs, which possess potent BLM inhibition and exhibit selectivity over related helicases. Moreover, these compounds demonstrated cellular activity by inducing sister chromatid exchanges, a hallmark of Bloom syndrome.     

二郎山小型兽类区系及分布格局

动物种类和数量在不同的山地或同一山地的不同垂直高度有很大的差异,正因如此,国际上和国内的研究者们一直在努力探讨动物在山地的分布规律和产生差异的原因(Merriam et al.,1890;鲍毅新等,1984;Heaney,2001;Rickart,2001;SánchezCordero,2001;张云智等,2002;李义明等,2003;马俊等,2010).横断山是中国最长、最宽和最典型的南北向山系,位于青藏高原东南部,通常为四川、云南两省西部和西藏自治区东部南北向山脉的总称.横断山是全球25个生物多样性热点地区之一,其生物多样性资源极其丰富(Myers et al.,2000).     

单气囊小肠镜在诊断胶囊内镜检查阴性的可疑小肠疾病患者中的应用价值

目的：探讨单气囊小肠镜（single-balloonenteroscop,SBE）在胶囊内镜检查阴性的可疑小肠疾病患者中的应用价值。方法：选取在我院行胶囊内镜检查无异常发现,后行单气囊小肠镜检查的可疑小肠疾病患者24例,分析后者对胶囊内镜检查阴性可疑小肠疾病患者的阳性发现率和病因的分布特点。结果：24例行SBE检查者有22例（91．7％）获得成功,2例失败,10例被检出阳性病变,其中间质瘤5例,小肠息肉2例,过敏性紫癜1例,血管畸形2例,SBE对胶囊内镜检查阴性的患者小肠疾病的再检出率为41．7％。结论：胶囊内镜和SBE在小肠疾病的诊断上有着各自的优缺点,对于胶囊内镜检查阴性的可疑小肠疾病患者进一步行SBE检查有助于确诊。     

Proteomic analysis of Rac1 signaling regulation by guanine nucleotide exchange factors

The small GTPase Rac1 is implicated in various cellular processes that are essential for normal cell function. Deregulation of Rac1 signaling has also been linked to a number of diseases, including cancer. The diversity of Rac1 functioning in cells is mainly attributed to its ability to bind to a multitude of downstream effectors following activation by Guanine nucleotide Exchange Factors (GEFs). Despite the identification of a large number of Rac1 binding partners, factors influencing downstream specificity are poorly defined, thus hindering the detailed understanding of both Rac1's normal and pathological functions. In a recent study, we demonstrated a role for 2 Rac-specific GEFs, Tiam1 and P-Rex1, in mediating Rac1 anti- versus pro-migratory effects, respectively. Importantly, via conducting a quantitative proteomic screen, we identified distinct changes in the Rac1 interactome following activation by either GEF, indicating that these opposing effects are mediated through GEF modulation of the Rac1 interactome. Here, we present the full list of identified Rac1 interactors together with functional annotation of the differentially regulated Rac1 binding partners. In light of this data, we also provide additional insights into known and novel signaling cascades that might account for the GEF-mediated Rac1-driven cellular effects.     

Effective aerial monitoring of cyanobacterial harmful algal blooms is dependent on understanding cellular migration

Cyanobacterial harmful algal blooms (CHABs) degrade water quality and may produce toxins. The distribution of CHABs can change rapidly due to variations in population dynamics and environmental conditions. Biological and ecological aspects of CHABs were studied in order to better understand CHABs dynamics. Field experiments were conducted near Hartington, Ontario, Canada in ponds dominated by Microcystis aeruginosa and CHABs floating experiments were conducted at Lake Taihu during the summers of 2015 and 2016. Single colonies composed of hundreds to thousands of cells with an average median of 0.2–0.5 mm in diameter were the basic form assumed by the Microcystis, and this remained the same throughout the growing season. Thorough mixing of the water column followed by calm conditions resulted in over 90% of the cyanobacteria floating after 1 h. Multiple colonies floated on the water surface in four types of assemblages: aggregates, ribbons, patches, and mats. It is the mats that are conventionally considered the blooming stage of cyanobacteria.Presence of CHABs on open water surfaces also depends on environmental influences such as direct and indirect wind effects. For example, field tests revealed that the surface coverage of CHABs can be reduced to half within an hour at wind speeds of 0.5 m/s.Because our findings indicated that blooming involves surface display of cyanobacteria essentially presenting as a two-dimensional plane under defined conditions, the use of surface imagery to quantify CHABs was justified. This is particularly important in light of the fact that traditional detection methods do not provide accurate distribution information. Nor do they portray CHABs events in a real-time manner due to limitations in on-demand surveillance and delays between sample time and analyzed results. Therefore, a new CHAB detection method using small unmanned aerial systems with consumer-grade cameras was developed at a maximum detection altitude of 80 m. When cyanobacteria were floating on the surface, CHABs detection through RGB band cameras and spectral enhancement techniques was efficient and accurate. Small unmanned aerial systems were capable of providing coverage up to 1 km² per mission and the short intervals between sampling and results (approx. 2 h) allowed for the rapid analysis of data and for implementing follow-up monitoring or treatments. This method is very cost-effective at an estimate of as low as $100 CAD per mission with an average cyanobacterial detection accuracy of 86%. Thus, it is a good candidate method to fill the urgent need for CHABs detection, providing cost effective, rapid, and accurate information to improve risk management at a local level as well as to help quickly allocate resources for purposes of mitigation.     

Identification of ortho-hydroxy anilide as a novel scaffold for lysine demethylase 5 inhibitors

Fe(II)/α-ketoglutarate-dependent lysine demethylases (KDMs) are attractive drug targets for several diseases including cancer. In this study, we designed and screened ortho-substituted anilides that are expected to function as Fe(II) chelators, and identified ortho-hydroxy anilide as a novel scaffold for KDM5A inhibitors. Treatment of human lung cancer A549 cells with a prodrug form of 4-carboxy-2-hydroxy-formanilide (9c) increased trimethylated lysine 4 on histone H3 level, suggesting KDM5 inhibition in the cells.     

HSP60/10 chaperonin systems are inhibited by a variety of approved drugs,natural products,and known bioactive molecules

All living organisms contain a unique class of molecular chaperones called 60?kDa heat shock proteins (HSP60 – also known as GroEL in bacteria). While some organisms contain more than one HSP60 or GroEL isoform, at least one isoform has always proven to be essential. Because of this, we have been investigating targeting HSP60 and GroEL chaperonin systems as an antibiotic strategy. Our initial studies focused on applying this antibiotic strategy for treating African sleeping sickness (caused by Trypanosoma brucei parasites) and drug-resistant bacterial infections (in particular Methicillin-resistant Staphylococcus aureus – MRSA). Intriguingly, during our studies we found that three known antibiotics – suramin, closantel, and rafoxanide – were potent inhibitors of bacterial GroEL and human HSP60 chaperonin systems. These findings prompted us to explore what other approved drugs, natural products, and known bioactive molecules might also inhibit HSP60 and GroEL chaperonin systems. Initial high-throughput screening of 3680 approved drugs, natural products, and known bioactives identified 161 hit inhibitors of the Escherichia coli GroEL chaperonin system (4.3% hit rate). From a purchased subset of 60 hits, 29 compounds (48%) re-confirmed as selective GroEL inhibitors in our assays, all of which were nearly equipotent against human HSP60. These findings illuminate the notion that targeting chaperonin systems might be a more common occurrence than we previously appreciated. Future studies are needed to determine if the in vivo modes of action of these approved drugs, natural products, and known bioactive molecules are related to GroEL and HSP60 inhibition.