In planta chemical cross‐linking and mass spectrometry analysis of protein structure and interaction in Arabidopsis

Site‐specific chemical cross‐linking in combination with mass spectrometry analysis has emerged as a powerful proteomic approach for studying the three‐dimensional structure of protein complexes and in mapping protein–protein interactions (PPIs). Building on the success of MS analysis of in vitro cross‐linked proteins, which has been widely used to investigate specific interactions of bait proteins and their targets in various organisms, we report a workflow for in vivo chemical cross‐linking and MS analysis in a multicellular eukaryote. This approach optimizes the in vivo protein cross‐linking conditions in Arabidopsis thaliana, establishes a MudPIT procedure for the enrichment of cross‐linked peptides, and develops an integrated software program, exhaustive cross‐linked peptides identification tool (ECL), to identify the MS spectra of in planta chemical cross‐linked peptides. In total, two pairs of in vivo cross‐linked peptides of high confidence have been identified from two independent biological replicates. This work demarks the beginning of an alternative proteomic approach in the study of in vivo protein tertiary structure and PPIs in multicellular eukaryotes.     

Relationship between Optical Coherence Tomography and Electrophysiology of the Visual Pathway in Non-Optic Neuritis Eyes of Multiple Sclerosis Patients

Purpose

Loss of retinal ganglion cells in in non-optic neuritis eyes of Multiple Sclerosis patients (MS-NON) has recently been demonstrated. However, the pathological basis of this loss at present is not clear. Therefore, the aim of the current study was to investigate associations of clinical (high and low contrast visual acuity) and electrophysiological (electroretinogram and multifocal Visual Evoked Potentials) measures of the visual pathway with neuronal and axonal loss of RGC in order to better understand the nature of this loss.

Methods

Sixty-two patients with relapsing remitting multiple sclerosis with no previous history of optic neuritis in at least one eye were enrolled. All patients underwent a detailed ophthalmological examination in addition to low contrast visual acuity, Optical Coherence Tomography, full field electroretinogram (ERG) and multifocal visual evoked potentials (mfVEP).

Results

There was significant reduction of ganglion cell layer thickness, and total and temporal retinal nerve fibre layer (RNFL) thickness (p<0.0001, 0.002 and 0.0002 respectively). Multifocal VEP also demonstrated significant amplitude reduction and latency delay (p<0.0001 for both). Ganglion cell layer thickness, total and temporal RNFL thickness inversely correlated with mfVEP latency (r = −0.48, p<0.0001 respectively; r = −0.53, p<0.0001 and r = −0.59, p<0.0001 respectively). Ganglion cell layer thickness, total and temporal RNFL thickness also inversely correlated with the photopic b-wave latency (r = −0.35, p = 0.01; r = −0.33, p = 0.025; r = −0.36, p = 0.008 respectively). Multivariate linear regression model demonstrated that while both factors were significantly associated with RGC axonal and neuronal loss, the estimated predictive power of the posterior visual pathway damage was considerably larger compare to retinal dysfunction.

Conclusion

The results of our study demonstrated significant association of RGC axonal and neuronal loss in NON-eyes of MS patients with both retinal dysfunction and post-chiasmal damage of the visual pathway.     

<Emphasis Type="Italic">Streptomyces urticae</Emphasis> sp. nov., isolated from rhizosphere soil of <Emphasis Type="Italic">Urtica urens</Emphasis> L.

Two novel Gram-stain positive, spore-forming, aerobic actinomycetes, designated NEAU-PCY-1^T and NEAU-PCY-2, were isolated from rhizosphere soil of Urtica urens L. collected from Anshan, Liaoning Province, northeast China. The 16S rRNA gene sequence analysis showed that strains NEAU-PCY-1^T and NEAU-PCY-2 exhibited 99.8% similarity with each other and are closely related to Streptomyces abietis DSM 42080^T (98.2, 98.3%) and Streptomyces fildesensis DSM 41987^T (98.0, 98.1%). Phylogenetic analysis based on the 16S rRNA gene sequences indicated that the two strains formed a cluster with these two closely related species. Moreover, DNA–DNA hybridization results and some phenotypic, physiological and biochemical properties differentiated the two strains from their close relatives in the genus Streptomyces. Based on a polyphasic taxonomy study, strains NEAU-PCY-1^T and NEAU-PCY-2 are considered to represent a novel species of the genus Streptomyces, for which the name Streptomyces urticae sp. nov. is proposed, with NEAU-PCY-1^T (=?DSM 105115^T?=?CCTCC AA 2017015^T) as the type strain.     

桉树种植对林地土壤丛枝菌根真菌群落结构及多样性的影响

桉树(Eucalyptus spp.)种植所产生的生态争议已受到广泛关注。丛枝菌根真菌(Arbuscular Mycorrhizal Fungi, AM真菌)是土壤微生物的重要组成部分,与根系共生后可促进植物的养分运输、提高植物逆境生存能力等。然而,桉树种植对土壤AM真菌群落结构和功能的影响尚不清楚。研究比较了次生林改种桉树后不同年限(2年/5年/10年)的土壤理化性质、AM真菌种类、群落组成及多样性的变化。结果表明,桉树种植显著改变了土壤理化性质,具体表现为2年生和10年生桉树林中土壤pH和有机碳显著降低(P0.05);AM真菌的孢子密度在桉树林土壤中显著低于次生林土壤(其中在5年生土壤中最低);AM真菌的种丰度随种植年份的增加逐渐下降,在10年生桉树林土壤中有所恢复; 5年生桉树的菌根侵染率最高,10年生桉树侵染率最低。AM真菌群落组成和结构发生显著变化,优势种Funneliforms geosporus的丰度在2年生和10年生林地中显著降低;而Septoglomus deserticola的丰度在2年生和10年生林地中显著增加。另外,冗余分析(Redundancy analysis, RDA)结果表明,土壤AM真菌群落结构主要受土壤pH值(解释率:89.88%)变化的影响。研究揭示了次生林改种桉树林后土壤AM真菌的群落变化特征,为桉树林的栽培管理和环境修复提供了数据支撑。     

A wheat biorefining strategy based on solid-state fermentation for fermentative production of succinic acid

In this study, a novel generic feedstock production strategy based on solid-state fermentation (SSF) has been developed and applied to the fermentative production of succinic acid. Wheat was fractionated into bran, gluten and gluten-free flour by milling and gluten extraction processes. The bran, which would normally be a waste product of the wheat milling industry, was used to produce glucoamylase and protease enzymes via SSF using Aspergillus awamori and Aspergillus oryzae, respectively. The resulting solutions were separately utilised for the hydrolysis of gluten-free flour and gluten to generate a glucose-rich stream of over 140gl(-1) glucose and a nitrogen-rich stream of more than 3.5gl(-1) free amino nitrogen. A microbial feedstock consisting of these two streams contained all the essential nutrients required for succinic acid fermentations using Actinobacillus succinogenes. In a fermentation using only the combined hydrolysate streams, around 22gl(-1) succinic acid was produced. The addition of MgCO(3) into the wheat-derived medium improved the succinic acid production further to more than 64gl(-1). These results demonstrate the SSF-based strategy is a successful approach for the production of a generic feedstock from wheat, and that this feedstock can be efficiently utilised for succinic acid production.     

Integrating multiple references for single-cell assignment

Efficient single-cell assignment is essential for single-cell sequencing data analysis. With the explosive growth of single-cell sequencing data, multiple single-cell sequencing data sources are available for the same kind of tissue, which can be integrated to further improve single-cell assignment; however, an efficient integration strategy is still lacking due to the great challenges of data heterogeneity existing in multiple references. To this end, we present mtSC, a flexible single-cell assignment framework that integrates multiple references based on multitask deep metric learning designed specifically for cell type identification within tissues with multiple single-cell sequencing data as references. We evaluated mtSC on a comprehensive set of publicly available benchmark datasets and demonstrated its state-of-the-art effectiveness for integrative single-cell assignment with multiple references.     

Bioimaging nitric oxide in activated macrophages in vitro and hepatic inflammation in vivo based on a copper–naphthoimidazol coordination compound

Nitric oxide (NO) serves as a messenger for cellular signaling and physiological reactions such as inflammatory responses in vivo. Fluorescent bioimaging of nitric oxide is a very useful tool in NO functional research. Although many encouraging results have been achieved in the field of NO fluorescent detection, there is rarely satisfying result in inflammatory NO imaging in vivo. Here we report that fluorescent 5′-chloro-2-(2′-hydroxyphenyl)-1H-naphtho[2,3-d]imidazol can coordinate with Cu(II) to form a non-fluorescent coordination compound, which is able to directly and quickly image NO in cellular system or in vivo inflammation system with a turn-on fluorescence, based on a redox action of Cu(II). It was used to image NO produced by inducible nitric oxide synthase (iNOS) in lipopolysaccharide (LPS) activated murine macrophages. More importantly, it could image the NO production in an acute severe hepatic injury (ASHI) model of BALB/c mice induced by integrative LPS and d-galactosamine (GalN) treatment. The results prove that the 5′-chloro-2-(2′-hydroxyphenyl)-1H-naphtho[2,3-d]imidazol coordinated with cupric ions can serve as an excellent NO bioimaging agent in different biological systems especially in inflammation related systems, and it may be valuable for diagnostic and pathological studies of NO related diseases.     

Fast bilayer-micelle fusion mediated by hydrophobic dipeptides

To understand the transition from inanimate matter to life, we studied a process that directly couples simple metabolism to evolution via natural selection, demonstrated experimentally by Adamala and Szostak. In this process, dipeptides synthesized inside precursors of cells promote absorption of fatty acid micelles to vesicles, inducing their preferential growth and division at the expense of other vesicles. The process is explained on the basis of coarse-grained molecular dynamics simulations, each extending for tens of microseconds, carried out to model fusion between a micelle and a membrane, both made of fatty acids in the absence and presence of hydrophobic dipeptides. In all systems with dipeptides, but not in their absence, fusion events were observed. They involve the formation of a stalk made by hydrophobic chains from the micelle and the membrane, similar to that postulated for vesicle-vesicle fusion. The emergence of a stalk is facilitated by transient clusters of dipeptides, side chains of which form hydrophobic patches at the membrane surface. Committor probability calculations indicate that the size of a patch is a suitable reaction coordinate and allows for identifying the transition state for fusion. Free-energy barrier to fusion is greatly reduced in the presence of dipeptides to only 4–5 kcal/mol, depending on the hydrophobicity of side chains. The mechanism of mediated fusion, which is expected to apply to other small peptides and hydrophobic molecules, provides a robust means by which a nascent metabolism can confer evolutionary advantage to precursors of cells.     

藏东南巴松错200年沉积过程及其对硅藻记录的影响

青藏高原东南部高寒山区广泛发育冰川湖泊,湖泊沉积过程同时受控于区域气候、流域水文、地质条件及湖泊形态特征。基于放射性²¹⁰Pb/¹³⁷Cs和¹⁴C定年法,对巴松错沉积物理(粒度、分选系数)和化学指标(TOC、TN、C/N)进行分析,发现18世纪末到19世纪末湖泊沉积过程显著变化,表现为迅速变缓趋势。通过分析区域树轮重建的气候序列(温度、降水及相对湿度)及冰川地貌调查资料,认为气候变化及流域冰川分布位置是影响该湖泊沉积过程的重要因素。小冰期末期冰川前缘靠近湖区,随后温度上升导致冰川融水激增、水动力加强,从而引起湖泊沉积粒度的粗化。随着冰川前缘不断后退,径流输送距离增长、沉积分选变好、粒度细化。此外,该地区活跃的地质活动也可能是湖泊沉积过程明显变化的重要诱因。湖泊沉积硅藻是研究气候环境变化的有力指标。过去200多年巴松错硅藻组合变化不明显(DCCA=0.47 SD),说明该地区气候环境变化未超过其生态阈值。通过与其他沉积指标进行对比分析发现,巴松错硅藻记录受到流域水文和湖泊沉积过程影响,主要表现为外源输入和/或湖岸浅水区来源...