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51.
MyD88是IL-1R/TLR受体超家族向细胞内转导胞外信号时募集到受体胞浆尾部的重要接头蛋白.由TIR结构域介导的MyD88分子同源二聚化是它招募到受体胞浆尾部的前提,然后二聚化的MyD88再募集下游信号分子,传递信号,引发促炎基因的表达.本研究旨在建立一种模型,以实现活细胞原位的、基于荧光信号变化的MyD88二聚化抑制物的高通量筛选.我们分别构建了MyD88 TIR与GFP和RFP的融合蛋白表达质粒,瞬时转染HeLa细胞,在488 nm激发光下,转染GFP-MyD88 TIR和RFP-MyD88 TIR细胞,检测到绿色荧光与红色荧光间的共振能量转移(FRET).而当细胞转染GFP-MyD88 TIR和RFP或RFP-MyD88 TIR和GFP,因TIR二聚化不能实现,FRET效率受到严重影响.实验结果提示,依赖双阳性表达GFP-MyD88 TIR和RFP-MyD88 TIR的细胞株,检测不同化合物对于荧光FRET效率的影响,可以建立MyD88 TIR二聚化抑制药物的筛选模型.此外,我们构建了原核表达质粒,利用纯化的His-MyD88 TIR分别与GST或GST-MyD88 TIR蛋白进行体外结合实验,发现GST-MyD88 TIR(而非GST)可以与His-MyD88 TIR相互结合.结果的差异性提示,利用His-MyD88 TIR和GST-MyD88 TIR体外结合实验分析,可以进一步确定抑制物是否直接阻断了TIR的相互作用.结合真核细胞的荧光FRET阻断结果和原核表达的重组蛋白相互作用分析,可确定MyD88 TIR二聚化的抑制物.利用这一模型可以对商品化的小分子库、自行制备的天然产物组分进行广泛的筛选,从中获得有效抑制MyD88二聚化的化合物,参与对MyD88信号通路依赖的慢性炎症、自身免疫性疾病的药物治疗. 相似文献
52.
报道中国鳖甲族1新纪录属及1新种:异颚弗鳖甲Freudeia heteromaxillaria sp.nov.,描述了新种的形态特征并附鉴别特征图和整体照片。新种模式标本保存于中国科学院西北高原生物研究所和河北大学博物馆。 相似文献
53.
Kirby TW Harvey S DeRose EF Chalov S Chikova AK Perrino FW Schaaper RM London RE Pedersen LC 《The Journal of biological chemistry》2006,281(50):38466-38471
The epsilon subunit of Escherichia coli DNA polymerase III possesses 3'-exonucleolytic proofreading activity. Within the Pol III core, epsilon is tightly bound between the alpha subunit (DNA polymerase) and subunit. Here, we present the crystal structure of epsilon in complex with HOT, the bacteriophage P1-encoded homolog of , at 2.1 A resolution. The epsilon-HOT interface is defined by two areas of contact: an interaction of the previously unstructured N terminus of HOT with an edge of the epsilon central beta-sheet as well as interactions between HOT and the catalytically important helix alpha1-loop-helix alpha2 motif of epsilon. This structure provides insight into how HOT and, by implication, may stabilize the epsilon subunit, thus promoting efficient proofreading during chromosomal replication. 相似文献
54.
Curcio A Noma T Naga Prasad SV Wolf MJ Lemaire A Perrino C Mao L Rockman HA 《American journal of physiology. Heart and circulatory physiology》2006,291(4):H1754-H1760
Adverse remodeling after myocardial infarction (MI) determines the progression of heart failure. Failing hearts are characterized by downregulation of beta-adrenergic receptor (beta-AR) signaling in part because of increased beta-AR kinase 1 activity. Our previous studies have shown that overexpression of the phosphoinositide kinase (PIK) domain of phosphoinositide 3-kinase (PI3K), prevents beta-AR downregulation and enhances adrenergic agonist responsiveness by inhibiting the targeting of PI3K to the beta-AR complex. To investigate whether preventing beta-AR downregulation in the heart ameliorates cardiac function post-MI, transgenic mice with cardiac-specific overexpression of the PIK domain peptide (TgPIK) underwent left coronary artery ligation and were subsequently followed by serial echocardiography at 4, 8, 12, 16, and 20 wk. Despite having similar infarction sizes, TgPIK mice showed better systolic function, less cardiac dilatation, and improved hemodynamic response to dobutamine compared with littermate controls after MI. To test that displacement of PI3K from the beta-AR complex, but not the total loss of PI3K-gamma, is critical for amelioration of cardiac function, mice lacking the PI3K-gamma (PI3K-gamma-KO) underwent MI, and their cardiac function was assessed 20 wk post-MI. Serial echocardiographic measurements showed severe reduction in contractile performance in PI3K-gamma-KO compared with TgPIK mice. Furthermore, significant beta-AR downregulation and desensitization were only seen in infarcted wild-type and PI3K-gamma-KO mice and not in TgPIK mice. Together, these results demonstrate that adverse remodeling of the ventricle after MI can be attenuated by a strategy that prevents recruitment of PI3K to the plasma membrane and restores normal beta-AR function. 相似文献
55.
GRAHAM E. ROTHERAY MARIA‐ANGELES MARCOS‐GARCÍA GEOFF HANCOCK CELESTE PÉREZ‐BAÑÓN CHRIS T. MAIER 《Zoological Journal of the Linnean Society》2009,156(4):697-749
We studied 42 species of saprophagous, Neotropical Copestylum (Diptera, Syrphidae) reared from decaying Cactaceae and Agavaceae. Thirty‐three species were reared during fieldwork in Bolivia, Costa Rica, Ecuador, Mexico, Peru, and Trinidad from 1998–2007. Nine species came from museum and private collections. Seven were new species. We describe these new species and the third stage larva and/or puparium and breeding sites of 40 species. Not described are two apparent species related to Copestylum apicale (Loew, 1866) reared from Cactaceae. Resolution of their status was beyond the scope of this paper but reference is made to their distinctive larval morphology. Based on early stage characters all reared species can be placed in ten species groups, all but three of which have been recognized previously on the basis of adult characters. A high level of congruence was found between adult and larval characters in terms of these species groups. Eight of the groups appear to be related closely and may represent a monophyletic lineage within Copestylum that has diversified in xeric habitats. Early stage morphology varied within and amongst groups but two trends in functional morphology are recognizable. One trend is towards feeding in watery decay and the other towards feeding in firmer decay. The latter trend is characterized by species that scoop food and use grinding mills in their head skeletons to break it up. They also have armoured thoraces with varying arrangements of sclerotized spicules or stiffened setae for gripping and protection during tunnelling, a short anal segment, and a short posterior breathing tube for protecting the openings. The former trend is characterized by species with opposite and contrasting features. They filter food and have well‐developed pre‐oral setal filters but they lack grinding mills or only have poorly developed grinding mills. They have reduced thoracic armature, elongate anal segments, and posterior breathing tubes which facilitates simultaneous feeding and respiration. Comparison with 23 Copestylum species reared from bromeliads (Bromeliaceae) suggests a common pattern of diversification in that species groups with the largest body sizes are more specialized. 相似文献
56.
目的探讨应用BR-TRG-I型体腔热灌注治疗系统进行持续循环腹腔热灌注治疗(continuous circulatory hyperthermic intraperitoneal perfusion treatment,CCHIP)对实验动物的生命体征、肝肾功能及内脏器官病理形态的影响。方法选用家猪作为实验动物,应用BR-TRG-I型体腔热灌注治疗系统建立CCHIP动物模型,分别用44℃、45℃的设定温度进行CCHIP。在CCHIP期间记录实验动物的生命体征变化;CCHIP前及治疗后1d、3d、7d和14d抽取外周血液保存备检;CCHIP结束后即刻及2周后处死动物,观察内脏器官的大体形态学改变,切取肝、肾、小肠等器官进行病理检查。结果44℃CCHIP持续1.5h对猪的生命体征、肝肾功能无明显影响,肝、肾、小肠等脏器损伤较轻,2周后恢复正常;45℃CCHIP持续1.5h对猪的生命体征影响严重,肝肾功能损害持续2周尚不能恢复正常,肝、肾、小肠等脏器病理损伤明显。结论44℃温度CCHIP1.5h安全可行,可以作为CCHIP的安全温度;45℃温度CCHIP1.5h可对实验动物肝肾功能造成严重损害,不适合作为CCHIP的治疗温度。 相似文献
57.
58.
It has been proposed that N-terminal myristoylation of calcineurin B is necessary for the membrane association of calcineurin. We tested the effects of Ca(2+) and myristoylation on the binding of calcineurin B alone or heterodimeric calcineurin to phosphatidylserine or phosphatidylcholine vesicles. In the presence of excess phosphatidylserine, 50-60% of total calcineurin associated with phosphatidylserine in a Ca(2+)-sensitive manner. Calcineurin did not associate with phosphatidylcholine. Calcineurin containing both the alpha and beta catalytic subunit isoforms bound to phosphatidylserine. Calmodulin interfered with the association of calcineurin with phosphatidylserine. In the presence of Ca(2+), myristoylated calcineurin B alone did not bind to phosphatidylcholine but did bind to phosphatidylserine, although to a lesser extent than the calcineurin heterodimer. Non-myristoylated calcineurin B alone, or calcineurin containing non-myristoylated calcineurin B did not associate with phosphatidylserine in the presence of Ca(2+). These results indicate: (i) Both isoforms of calcineurin bind to phosphatidylserine. (ii) A phospholipid binding site is located on the calcineurin B subunit. (iii) Calcineurin B myristoylation is required for the Ca(2+)-sensitive binding of calcineurin to phosphatidylserine vesicles in vitro. 相似文献
59.
棉蚜体色变化的生态遗传学研究 总被引:15,自引:0,他引:15
调查了不同寄主上棉蚜刀Aphis gossypll自受精卵孵化出的自然种群、室内混合饲养以及单个饲养蚜虫的体色变化。结果表明:不论是自然还是实验种群,是群体还是个体饲养,不论寄主、栽培条件、生育期营养相同与否,棉蚜体色在世代内稳定不变,即出生时是什么颜色保持终生不变;在世代间则随温度升高体色渐变为黄色,温度降低体色逐渐转绿。伏蚜由苗蚜而来。X2检验证实:棉蚜体色变化与营养、寄主种类、光照、光质、栽培条件等无关,仅与温度密切相关,属于同一基因型在不同环境条件下的反应规范。但在太槿上还发现有个别深黄色棉蚜,从卵孵化到迁飞体色不随温度变化,表明棉蚜体色变化中还存在遗传多态现象。胚胎学观察与染色体校型分析结果证实了上述结论与观点。 相似文献
60.
In situ hybridization at the electron microscope level: hybrid detection by autoradiography and colloidal gold 总被引:14,自引:4,他引:10 下载免费PDF全文
In situ hybridization has become a standard method for localizing DNA or RNA sequences in cytological preparations. We developed two methods to extend this technique to the transmission electron microscope level using mouse satellite DNA hybridization to whole mount metaphase chromosomes as the test system. The first method devised is a direct extension of standard light microscope level using mouse satellite DNA hybridization to whole mount metaphase chromosomes as the test system. The first method devised is a direct extension of standard light microscope in situ hybridization. Radioactively labeled complementary RNA (cRNA) is hybridized to metaphase chromosomes deposited on electron microscope grids and fixed in 70 percent ethanol vapor; hybridixation site are detected by autoradiography. Specific and intense labeling of chromosomal centromeric regions is observed even after relatively short exposure times. Inerphase nuclei present in some of the metaphase chromosome preparations also show defined paatterms of satellite DNA labeling which suggests that satellite-containing regions are associate with each other during interphase. The sensitivity of this method is estimated to at least as good as that at the light microscope level while the resolution is improved at least threefold. The second method, which circumvents the use of autoradiogrphic detection, uses biotin-labeled polynucleotide probes. After hybridization of these probes, either DNA or RNA, to fixed chromosomes on grids, hybrids are detected via reaction is improved at least threefold. The second method, which circumvents the use of autoradiographic detection, uses biotin-labeled polynucleotide probes. After hybridization of these probes, either DNA or RNA, to fixed chromosomes on grids, hybrids are detected via reaction with an antibody against biotin and secondary antibody adsorbed to the surface of over centromeric heterochromatin and along the associated peripheral fibers. Labeling is on average ten times that of background binding. This method is rapid and possesses the potential to allow precise ultrastructual localization of DNA sequences in chromosomes and chromatin. 相似文献