Design and application of highly responsive fluorescence resonance energy transfer biosensors for detection of sugar in living Saccharomyces cerevisiae cells

A protein sensor with a highly responsive fluorescence resonance energy transfer (FRET) signal for sensing sugars in living Saccharomyces cerevisiae cells was developed by combinatorial engineering of the domain linker and the binding protein moiety. Although FRET sensors based on microbial binding proteins have previously been created for visualizing various sugars in vivo, such sensors are limited due to a weak signal intensity and a narrow dynamic range. In the present study, the length and composition of the linker moiety of a FRET-based sensor consisting of CFP-linker(1)-maltose-binding protein-linker(2)-YFP were redesigned, which resulted in a 10-fold-higher signal intensity. Molecular modeling of the composite linker moieties, including the connecting peptide and terminal regions of the flanking proteins, suggested that an ordered helical structure was preferable for tighter coupling of the conformational change of the binding proteins to the FRET response. When the binding site residue Trp62 of the maltose-binding protein was diversified by saturation mutagenesis, the Leu mutant exhibited an increased binding constant (82 microM) accompanied by further improvement in the signal intensity. Finally, the maltose sensor with optimized linkers was redesigned to create a sugar sensor with a new specificity and a wide dynamic range. When the optimized maltose sensors were employed as in vivo sensors, highly responsive FRET images were generated from real-time analysis of maltose uptake of Saccharomyces cerevisiae (baker's yeast).     

Mycobacterium kansasii-induced death of murine macrophages involves endoplasmic reticulum stress responses mediated by reactive oxygen species generation or calpain activation

Although pathogenic mechanisms of tuberculosis have been extensively studied, little is known about the pathogenic mechanisms of Mycobacterium kansasii. In this work the influence of virulence and ER-stress mediated apoptosis of macrophages during two different strains of M. kansasii infection was investigated. We show that M. kansasii infection is associated with ER stress-mediated apoptosis in the murine macrophage cell line RAW 264.7. Infection of RAW 264.7 cells in vitro with apoptosis-inducing a clinical isolate of M. kansasii SM-1 (SM-1) resulted in strong induction of ER stress responses compared with M. kansasii type strain (ATCC 12478)-infected RAW 264.7 cells. Interestingly, inhibition of calpain prevented the induction of CHOP and Bip in ATCC 12478-infected RAW 264.7 cells but not in RAW 264.7 cells infected with SM-1. In contrast, reactive oxygen species (ROS) were significantly increased only in RAW 264.7 cells infected with SM-1. We propose that ROS generation is important for triggering ER stress-mediated apoptosis during SM-1 infection, whereas ATCC 12478-induced, ER stress-mediated apoptosis is associated with calpain activation. Our results demonstrate that the ER stress pathway plays important roles in the pathogenesis of M. kansasii infections, and that different strains of M. kansasii induce different patterns of ER stress-mediated apoptosis.     

三种几丁质合成抑制剂对意大利蝗的防治研究

本文用3种几丁质合成抑制剂卡死克、噻嗪酮和灭幼脲对意大利蝗Calliptatmnus italicus(L.)卵和3龄蝗蝻进行药剂试验.实验结果显示卡死克对意大利蝗药效最高:LC50、LC90分别为1.34、14.17 mg/L.灭幼脲次之LC50、LC90分别为2.09、45.22 mg/L.灭幼脲和卡死克在50 m...     

Intentionally introduced species: more easily invited than removed

Alien species are brought into countries world wide on a massive scale for agricultural production, ex situ conservation, landscape aesthetics, gardens, and ecosystem restoration. Unfortunately, some of these species have escaped and adversely impacted on regional as well as global biodiversity conservation and agricultural production. To reduce such risks, it is necessary to implement specific and effective measures. Since various government departments and institutions are involved in the management of alien species, it is difficult to prevent native and agroecosystems from being invaded by invited species. We propose the establishment of a supervision and inspection continuum over intentional species introduction, similar to that which exists in some countries over unintentional species introductions. Namely, a justification of the necessity to import, a risk assessment, assurances as to provision of an adequate containment facility assessment, and a damage-limitation protocol should that need to be invoked. These requirements should be satisfied before an alien species is knowingly imported, and the necessary follow-up supervision is important post- importation.     

Iron metabolism in diabetes-induced Alzheimer’s disease: a focus on insulin resistance in the brain

Alzheimer’s disease (AD) is characterized by an excessive accumulation of toxic amyloid beta (Aβ) plaques and memory dysfunction. The onset of AD is influenced by age, genetic background, and impaired glucose metabolism in the brain. Several studies have demonstrated that diabetes involving insulin resistance and glucose tolerance could lead to AD, ultimately resulting in cognitive dysfunction. Even though the relationship between diabetes and AD was indicated by significant evidences, the critical mechanisms and metabolic alterations in diabetes induced AD are not clear until now. Recently, iron metabolism has been shown to play multiple roles in the central nervous system (CNS). Iron deficiency and overload are associated with neurodegenerative diseases. Iron binds to Aβ and subsequently regulates Aβ toxicity in the CNS. In addition, previous studies have shown that iron is involved in the aggravation of insulin resistance. Considering these effects of iron metabolism in CNS, we expect that iron metabolism may play crucial roles in diabetic AD brain. Thus, we review the recent evidence regarding the relationship between diabetes-induced AD and iron metabolism.     

几个生态因子对黄盖鲽的影响

黄盖鲽(Psuedopleuronectes yokohameGünther)是鲽科中的冷温性鱼类,广泛分布在日本、朝鲜及我国的东海、黄海和渤海。地方种群多,洄游范尉小,适应能力强,既是底栖生物食性,又可耐较低的温度,虽非名贵鱼种,却具有一定的经济价值,在开展近海增殖时,不存在越冬和效益外流问题。因此,早在60年代,日本山口县和大分县的学者们,便开始着手该品种的人工孵化和增殖放流试验。80年代初已达年放流2—3cm的稚鱼苗7.0×10~6尾的水平。为了在渤海开展增殖工作,建立综     

Catalase, but not MnSOD, inhibits glucose deprivation-activated ASK1-MEK-MAPK signal transduction pathway and prevents relocalization of Daxx: hydrogen peroxide as a major second messenger of metabolic oxidative stress

Overexpression of catalase, but not manganese superoxide dismutase (MnSOD), inhibited glucose deprivation-induced cytotoxicity and c-Jun N-terminal kinase 1 (JNK1) activation in human prostate adenocarcinoma DU-145 cells. Suppression of JNK1 activation by catalase overexpression resulted from inhibition of apoptosis signal-regulating kinase 1 (ASK1) activation by preventing dissociation of thioredoxin (TRX) from ASK1. Overexpression of catalase also inhibited relocalization of Daxx from the nucleus to the cytoplasm as well as association of Daxx with ASK1 during glucose deprivation. Taken together, hydrogen peroxide (H(2)O(2)) rather than superoxide anion (O(2) (*-)) acts as a second messenger of metabolic oxidative stress to activate the ASK1-MAPK/extracellular signal-regulated kinase (ERK) kinase (MEK)-mitogen-activated protein kinase (MAPK) signal transduction pathway.     

Rht10基因对‘鲁麦15’农艺性状和光合生理特性的影响

利用鲁麦15、Rht10鲁麦15、Ms2鲁麦15及Ms2+Rht10鲁麦15近等基因系为试验材料,研究了Rht10基因对小麦农艺性状、净光合速率(Pn)、蒸腾速率(Tr)等光合生理特性的影响.结果表明,Rht10使小麦的挑旗、抽穗、开花期均推迟了4～5 d;Rht10对鲁麦15和Ms2鲁麦15的降秆强度分别为53.77%和53.00%,穗长显著缩短(P<0.05),千粒重显著减少,但对有效穂数无显著影响(P>0.05).含有Rht10基因的材料与不含此基因的材料之间的光合生理参数普遍存在显著差异,但在不同时期不同参数差异正负不同;在灌浆后期,Rht10对Pn有显著正效应(P<0.05);从开花期到灌浆后期,Pn与气孔导度(Gs)、胞间二氧化碳浓度(Ci)总体变化趋势相同,且相关性显著,表明Pn在一定程度上受气孔因素的限制;在灌浆中后期,Rht10对Gs有明显的正效应;在抽穗和开花期,Rht10对Tr有明显的正效应;Rht10对叶片水分利用效率(WUE)在4个生育期有明显的负效应.由此可见,Rht10基因对小麦的生育期、株高、穗长都有明显的不利影响,对各项光合生理参数也有普遍正向或负向的影响,在利用Rht10进行杂交育种时应充分考虑此基因带来的不利效应.