Saposin Lipid Nanoparticles: A Highly Versatile and Modular Tool for Membrane Protein Research

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A Golgi Lipid Signaling Pathway Controls Apical Golgi Distribution and Cell Polarity during Neurogenesis

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丹参非特异性脂质转移蛋白基因（SmLTP1）的克隆及其表达分析

对丹参EST序列进行Blast分析,获得一个新的非特异性脂质转移蛋白基因,命名为SmLTP1（GenBank注册号为EF187461）。该基因cDNA全长593bp,包含一个长为357bp的开放读码框,编码118个氨基酸。生物信息学结构分析表明,该蛋白具有植物nsLTP的典型结构,即4对二硫键,4个a-螺旋,1个可结合和容纳脂肪酸分子的类似口袋状的疏水结构。实时荧光定量PCR分析结果表明,SmLTP1基因在丹参不同组织器官中差异表达,其表达受病原菌和茉莉酸甲酯的诱导,显示SmLTP1基因在植物防御反应中发挥作用。     

Hydrogen Peroxide-Mediated Growth of the Root System Occurs via Auxin Signaling Modification and Variations in the Expression of Cell-Cycle Genes in Rice Seedlings Exposed to Cadmium Stress

The link between root growth, H₂O₂, auxin signaling, and the cell cycle in cadmium (Cd)‐stressed rice (Oryza sativa L. cv. Zhonghua No. 11) was analyzed in this study. Exposure to Cd induced a significant accumulation of Cd, but caused a decrease in zinc (Zn) content which resulted from the decreased expression of OsHMA9 and OsZIP. Analysis using a Cd‐specific probe showed that Cd was mainly localized in the meristematic zone and vascular tissues. Formation and elongation of the root system were significantly promoted by 3‐amino‐1,2,4‐triazole (AT), but were markedly inhibited by N,N’‐dimethylthiourea (DMTU) under Cd stress. The effect of H₂O₂ on Cd‐stressed root growth was further confirmed by examining a gain‐of‐function rice mutant (carrying catalase1 and glutathione‐S‐transferase) in the presence or absence of diphenylene iodonium. DR5‐GUS staining revealed close associations between H₂O₂ and the concentration and distribution of auxin. H₂O₂ affected the expression of key genes, including OsYUCCA, OsPIN, OsARF, and OsIAA, in the auxin signaling pathway in Cd‐treated plants. These results suggest that H₂O₂ functions upstream of the auxin signaling pathway. Furthermore, H₂O₂ modified the expression of cell‐cycle genes in Cd‐treated roots. The effects of H₂O₂ on root system growth are therefore linked to auxin signal modification and to variations in the expression of cell‐cycle genes in Cd‐stressed rice. A working model for the effects of H₂O₂ on Cd‐stressed root system growth is thus proposed and discussed in this paper.     

Control of Drosophila Growth and Survival by the Lipid Droplet-Associated Protein CG9186/Sturkopf

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Cellular senescence and chronological age in various human tissues: A systematic review and meta‐analysis

Senescent cells in tissues and organs are considered to be pivotal to not only the aging process but also the onset of chronic disease. Accumulating evidence from animal experiments indicates that the magnitude of senescence can vary within and between aged tissue samples from the same animal. However, whether this variation in senescence translates across to human tissue samples is unknown. To address this fundamental question, we have conducted a systematic review and meta‐analysis of all available literature investigating the magnitude of senescence and its association with chronological age in human tissue samples. While senescence is higher in aged tissue samples, the magnitude of senescence varies considerably depending upon tissue type, tissue section, and marker used to detect senescence. These findings echo animal experiments demonstrating that senescence levels may vary between organs within the same animal.     

钙调素对钙调素结合蛋白-10和玉米非特异性脂转移蛋白与脂质结合活性的影响

荧光标记的脂质结合实验表明,钙调素结合蛋白-10（CaMBP-10）具有典型的植物非特异性脂质转移蛋白与脂质结合的特性。进一步实验研究了钙调素（calmodulin,CaM）对CaMBP-10和玉米nsLTP与脂质结合的活性的影响,结果显示无论在有钙和无钙条件下,CaM对两者的影响均有不同之处,W-7和TFP能消除CaM的影响。提示CaM不仅与CaMBP-10和玉米nsLTP特异性相互作用,而且对2种脂转移蛋白可能具有不同的调节机制。     

小桐子非特异性脂质转移蛋白A的基因克隆及其表达和功能分析

基于我们最近获得的小桐子低温驯化转录组和数字基因表达谱数据,本工作研究了低温驯化条件下差异表达变化较大的非特异性脂质转移蛋白A基因JcnsLTPA。克隆到该基因的cDNA序列全长833 bp,开放阅读框长度513 bp,编码170个氨基酸,存在ATT_LTSS典型保守功能基序。其启动子区域中鉴定到了TATA框、CAAT框、CATA框、W框等顺式作用元件以及CRT/DRE低温响应元件。半定量RT-PCR分析表明,该基因在茎、根、叶中都有表达,以茎中表达量最高、且受低温诱导最显著。同时,酵母表达JcnsLTPA也提高了重组酵母菌的抗低温能力。这些结果充分说明了小桐子JcnsLTPA是与抗冷性密切相关的基因,可以用于小桐子的抗冷性遗传改良。     

Improvement in Lipid and Protein Trafficking in Niemann‐Pick C1 Cells by Correction of a Secondary Enzyme Defect

Different primary lysosomal trafficking defects lead to common alterations in lipid trafficking, suggesting cooperative interactions among lysosomal lipids. However, cellular analysis of the functional consequences of this phenomenon is lacking. As a test case, we studied cells with defective Niemann‐Pick C1 (NPC1) protein, a cholesterol trafficking protein whose defect gives rise to lysosomal accumulation of cholesterol and other lipids, leading to NPC disease. NPC1 cells also develop a secondary defect in acid sphingomyelinase (SMase) activity despite a normal acid SMase gene (SMPD1). When acid SMase activity was restored to normal levels in NPC1‐deficient CHO cells through SMPD1 transfection, there was a dramatic reduction in lysosomal cholesterol. Two other defects, excess lysosomal bis‐(monoacylglycerol) phosphate (BMP) and defective transferrin receptor (TfR) recycling, were also markedly improved. To test its relevance in human cells, the acid SMase activity defect in fibroblasts from NPC1 patients was corrected by SMPD1 transfection or acid SMase enzyme replacement. Both treatments resulted in a dramatic reduction in lysosomal cholesterol. These data show that correcting one aspect of a complex lysosomal lipid storage disease can reduce the cellular consequences even if the primary genetic defect is not corrected.     

血清中性粒细胞明胶酶相关脂质运载蛋白、胱抑素C联合红细胞分布宽度对脓毒症患儿急性肾损伤的预测价值

摘要 目的：研究血清中性粒细胞明胶酶相关脂质运载蛋白（NGAL）、胱抑素C联合红细胞分布宽度（RDW）对脓毒症患儿急性肾损伤（AKI）的预测价值。方法：选取我院从2016年1月～2019年1月收治的脓毒症患儿154例。将所有患儿根据是否合并AKI分为AKI组67例和非AKI组87例。分析两组临床资料的差异,检测并比较血清NGAL、胱抑素C以及RDW水平。采用多因素Logistic回归分析脓毒症患儿发生AKI的影响因素。采用受试者工作特征（ROC）曲线分析血清NGAL、胱抑素C以及RDW预测脓毒症患儿AKI的效能。结果：AKI组与非AKI组在年龄、小儿危重病例评分（PCIS）、儿童重症监护室（PICU）停留时间、第三代小儿死亡危险评分(PRISM-Ⅲ)、肌酐（Cr）方面比较差异均有统计学意义（P＜0.05）。AKI组血清NGAL、胱抑素C以及RDW水平均高于非AKI组（P＜0.05）。经多因素Logistic回归分析发现：脓毒症患儿发生AKI的危险因素有PICU停留时间≥15 d、PRISM-Ⅲ＞32分、Cr＞100 μmol/L、血清NGAL＞220 ng/mL、血清胱抑素C＞1.55 mg/L以及RDW＞14%（均OR＞1,P＜0.05）;而防止脓毒症患儿发生AKI的保护因素包括年龄＞50月、PCIS＞70分（均OR＜1,P＜0.05）。经ROC曲线分析发现：血清NGAL、胱抑素C以及RDW联合检测预测脓毒症患儿AKI的曲线下面积、灵敏度、特异度以及约登指数均高于上述三项指标单独检测。结论：脓毒症合并AKI的患儿血清NGAL、胱抑素C以及RDW水平异常升高,且联合检测以上指标对脓毒症患儿AKI的预测价值较高,具有一定临床应用价值。     

Towards the Molecular Mechanism of Pulmonary Surfactant Protein SP-B: At the Crossroad of Membrane Permeability and Interfacial Lipid Transfer

Pulmonary surfactant is a lipid-protein complex that coats the alveolar air-liquid interface, enabling the proper functioning of lung mechanics. The hydrophobic surfactant protein SP-B, in particular, plays an indispensable role in promoting the rapid adsorption of phospholipids into the interface. For this, formation of SP-B ring-shaped assemblies seems to be important, as oligomerization could be required for the ability of the protein to generate membrane contacts and to mediate lipid transfer among surfactant structures. SP-B, together with the other hydrophobic surfactant protein SP-C, also promotes permeability of surfactant membranes to polar molecules although the molecular mechanisms underlying this property, as well as its relevance for the surface activity of the protein, remain undefined. In this work, the contribution of SP-B and SP-C to surfactant membrane permeability has been further investigated, by evaluation of the ability of differently-sized fluorescent polar probes to permeate through giant vesicles with different lipid/protein composition. Our results are consistent with the generation by SP-B of pores with defined size in surfactant membranes. Furthermore, incubation of surfactant with an anti-SP-B antibody not only blocked membrane permeability but also affected lipid transfer into the air-water interface, as observed in a captive bubble surfactometer device. Our findings include the identification of SP-C and anionic phospholipids as modulators required for maintaining native-like permeability features in pulmonary surfactant membranes. Proper permeability through membrane assemblies could be crucial to complement the overall role of surfactant in maintaining alveolar equilibrium, beyond its biophysical function in stabilizing the respiratory air-liquid interface.     

大气CO2与O3浓度升高对毛竹叶片膜脂过氧化和抗氧化系统的影响

以盆栽一年生毛竹为材料,利用开顶式气室模拟大气O3和CO2浓度升高情景,分析毛竹叶片细胞膜脂过氧化、抗氧化酶(SOD、CAT、POD和APX)活性和渗透调节物质(可溶性糖和可溶性蛋白)含量的响应规律,为气候变化背景下的竹林适应性管理提供理论支撑.结果表明:(1)短期(30 d)高浓度O3(92～106 nL-L-1)胁迫能刺激毛竹叶片抗氧化酶活性和渗透调节物质含量显著提高,清除活性氧能力增强,并未出现细胞膜脂过氧化现象,即毛竹对短期高浓度O3具有较强的适应性;但长期(90 d)O3胁迫条件下,毛竹叶片抗氧化酶活性显著降低,叶片细胞膜脂过氧化程度加剧,膜结构破坏,发生严重的氧化伤害.(2)短期高浓度CO2(685～730 μmol·mol-1)处理总体上对毛竹叶片细胞膜脂过氧化和抗氧化酶系统活性影响并不明显;而长期高浓度CO2处理能一定程度上增强毛竹的抗氧化能力和渗透调节功能,减轻氧化损伤,体现了对毛竹的保护效应.(3)高浓度O3和CO2复合作用下,相对于单一高浓度O3胁迫,毛竹叶片能够维持较高的抗氧化酶活性和渗透调节物质含量,有效地调节活性氧产生与清除间的平衡,细胞膜脂过氧化程度变化不明显,说明高浓度CO2可在一定程度上缓解高浓度O3对毛竹所造成的生理伤害.     

不同浓度糖氮对离体穗培养小麦粒重和叶片膜脂过氧化作用的影响

在相同浓度配比的蔗糖和谷氨酰胺培养液中培养小麦品种扬麦9号和徐州26号的离体穗,检测不同水平的糖氮对小麦粒重及叶片膜脂过氧化作用的影响.结果表明,在一定浓度范围内,提高培养液中蔗糖和谷氨酰胺供应水平后,单粒重、叶片的超氧化物歧化酶(sOD)、过氧化氢酶(CAT)活性提高,膜脂过氧化产物丙二醛(MDA)含量降低;培养液中糖氮水平过高,叶片的sOD和CAT活性则下降,MDA含量升高,籽粒单粒重下降.     

Analysis of Apoptosis Induced by HIV-1 Vpr and Examination of the Possible Role of the hHR23A Protein

The HIV-1 Vpr protein induces apoptosis of cells, the mechanism of which is unknown. To clarify how this function may be related to other Vpr functions, we simultaneously assessed the effects of multiple point mutations upon various Vpr properties. Our data suggest that induction of arrest by Vpr may be unnecessary for induction of apoptosis. This is exemplified by a C-terminal mutant, R80A, that does not arrest cells, yet induces low but significant levels of apoptosis. We also show that mutation of Vpr at both of its nuclear localization sequences (within its alpha-helices and the overlapping leucine zipper-like domain) does not affect induction of either apoptosis or cell cycle arrest. This indicates that neither sequence is essential for these two functions of Vpr. It further suggests that multimerization of Vpr, which maps to residues 60 and 67 within the leucine-rich region, is unnecessary for initiation of apoptosis and arrest. We previously found that the Vpr-binding protein, hHR23A, can partially alleviate induction of arrest. We now show that overexpression of hHR23A itself causes apoptosis of cells. Mutation of its C-terminal UBA( 2 ) domain that is responsible for binding Vpr disrupts the apoptotic effect. This suggests that Vpr may induce apoptosis through a pathway involving hHR23A.     

A Molecular Model of Inducible Costimulator Protein and Three-Dimensional Analysis of its Relation to the CD28 Family of T Cell-Specific Costimulatory Receptors

Inducible costimulator protein (ICOS) has recently been identified as a new member of the CD28 family of T cell costimulatory molecules. A molecular model of the extracellular immunoglobulin-like domain of ICOS was built based on the structure of CD152, another member of the CD28 family. Despite low sequence identity, ICOS shares consensus residues characteristic of immunoglobulin variable-type domains with CD152 and CD28 and also some unique features, suggesting that their three-dimensional structures are more similar to each other than to other proteins belonging to the immunoglobulin superfamily. The ICOS model was used to study sequence conservation in three dimensions and to compare the distribution of N-linked glycosylation sites in the extended CD28 family. The limited number of residues outside consensus/core positions that are conserved in ICOS and CD28 and/or CD152 are widely distributed over the extracellular domain. A few residues in CD152 and CD28 that are critical for binding of CD80/CD86 are also conserved in ICOS. However, the region in ICOS that corresponds to the CD80/CD86 binding site is masked by N-linked glycosylation. This suggests that this site is not available for binding of CD80/CD86 or other ligands. ICOS has probably diverged early from CD28 and CD152 and developed the capacity to recognize ligand(s) other than CD80/CD86, very likely utilizing a different molecular region and mechanism for binding.Supplementary material to this paper is available in electronic form at http://dx.doi.org/10.1007/s008940050116     

H9724, a Monoclonal Antibody to Borrelia burgdorferi's Flagellin, Binds to Heat Shock Protein 60 (HSP60) Within Live Neuroblastoma Cells: A Potential Role for HSP60 in Peptide Hormone Signaling and in an Autoimmune Pathogenesis of the Neuropathy of Lyme Disease

Although Borrelia burgdorferi, the causative agent of Lyme disease, is found at the site of many disease manifestations, local infection may not explain all its features. B. burgdorferi's flagellin cross-reacts with a component of human peripheral nerve axon, previously identified as heat shock protein 60 (HSP60). The cross-reacting epitopes are bound by a monoclonal antibody to B. burgdorferi's flagellin, H9724. Addition of H9724 to neuroblastoma cell cultures blocks in vitro spontaneous and peptide growth-factor–stimulated neuritogenesis. Withdrawal of H9724 allows return to normal growth and differentiation. Using electron microscopy, immunoprecipitation and immunoblotting, and FACS analysis we sought to identify the site of binding of H9724, with the starting hypotheses that the binding was intracellular and not identical to the binding site of II-13, a monoclonal anti-HSP60 antibody. The current studies show that H9724 binds to an intracellular target in cultured cells with negligible, if any, surface binding. We previously showed that sera from patients with neurological manifestations of Lyme disease bound to human axons in a pattern identical to H9724's binding; these same sera also bind to an intracellular neuroblastoma cell target. II-13 binds to a different HSP60 epitope than H9724; II-13 does not modify cellular function in vitro. As predicted, II-13 bound to mitochondria, in a pattern of cellular binding very different from H9724, which bound in a scattered cytoplasmic, nonorganelle-related pattern. H9724's effect is the first evidence that HSP60 may play a role in peptide-hormone–receptor function and demonstrates the modulatory potential of a monoclonal antibody on living cells.