The role of presenilin cofactors in the gamma-secretase complex

Mutations in presenilin genes account for the majority of the cases of the familial form of Alzheimer's disease (FAD). Presenilin is essential for gamma-secretase activity, a proteolytic activity involved in intramembrane cleavage of Notch and beta-amyloid precursor protein (betaAPP). Cleavage of betaAPP by FAD mutant presenilin results in the overproduction of highly amyloidogenic amyloid beta42 peptides. gamma-Secretase activity requires the formation of a stable, high-molecular-mass protein complex that, in addition to the endoproteolysed fragmented form of presenilin, contains essential cofactors including nicastrin, APH-1 (refs 15-18) and PEN-2 (refs 16, 19). However, the role of each protein in complex formation and the generation of enzymatic activity is unclear. Here we show that Drosophila APH-1 (Aph-1) increases the stability of Drosophila presenilin (Psn) holoprotein in the complex. Depletion of PEN-2 by RNA interference prevents endoproteolysis of presenilin and promotes stabilization of the holoprotein in both Drosophila and mammalian cells, including primary neurons. Co-expression of Drosophila Pen-2 with Aph-1 and nicastrin increases the formation of Psn fragments as well as gamma-secretase activity. Thus, APH-1 stabilizes the presenilin holoprotein in the complex, whereas PEN-2 is required for endoproteolytic processing of presenilin and conferring gamma-secretase activity to the complex.     

不确定Sigma-Delta调制器的鲁棒滤波

研究级联Sigma-Delta调制器在非理想性电路中的鲁棒校正滤波器设计问题.基于MARKOV区间算法和线性区间系统的等价描述,将扰动抑制问题转化为增广系统的鲁棒稳定问题.给出以线性矩阵不等式描述的鲁棒滤波器存在的充分条件,该条件保证所设计的校正滤波器能有效抑制电路不确定性对调制器性能的影响.仿真结果表明,与传统校正滤波器相比,该调制器获得了更好的信号噪声比.方案降低了电路实现过程中器件精确度的要求,从而达到低应用成本的目的.     

Delta Sigma调制器非理想因素建模

提出并构建了开关电容积分器Delta Sigma调制器非理想因素行为级模型,该模型基于Matlab中的Simulink工具,包含开关非线性、时钟抖动、量化器非线件和积分器非线性等调制器非理想囚素,能为电路模块的设计提供精确的设计指标.重点研究并实现一种运放非线性直流增益模型,仿真结果表明它能更有效反映奇次谐波失真.同时综合考虑调制器其他非理想因素,例如采样噪声、开关非线性电阻以及运放参数(色化噪声.增益带宽,摆率,饱和电压),仿真得到其对调制器性能的影响.     

Photoactivated gamma-secretase inhibitors directed to the active site covalently label presenilin 1

Cleavage of amyloid precursor protein (APP) by the beta- and gamma-secretases generates the amino and carboxy termini, respectively, of the A beta amyloidogenic peptides A beta40 and A beta42--the major constituents of the amyloid plaques in the brain parenchyma of Alzheimer's disease patients. There is evidence that the polytopic membrane-spanning proteins, presenilin 1 and 2 (PS1 and PS2), are important determinants of gamma-secretase activity: mutations in PS1 and PS2 that are associated with early-onset familial Alzheimer's disease increase the production of A beta42 (refs 4-6), the more amyloidogenic peptide; gamma-secretase activity is reduced in neuronal cultures derived from PS1-deficient mouse embryos; and directed mutagenesis of two conserved aspartates in transmembrane segments of PS1 inactivates the ability of gamma-secretase to catalyse processing of APP within its transmembrane domain. It is unknown, however, whether PS1 (which has little or no homology to any known aspartyl protease) is itself a transmembrane aspartyl protease or a gamma-secretase cofactor, or helps to colocalize gamma-secretase and APP. Here we report photoaffinity labelling of PS1 (and PS2) by potent gamma-secretase inhibitors that were designed to function as transition state analogue inhibitors directed to the active site of an aspartyl protease. This observation indicates that PS1 (and PS2) may contain the active site of gamma-secretase. Interestingly, the intact, single-chain form of wild-type PS1 is not labelled by an active-site-directed photoaffinity probe, suggesting that intact wild-type PS1 may be an aspartyl protease zymogen.     

Two transmembrane aspartates in presenilin-1 required for presenilin endoproteolysis and gamma-secretase activity

Accumulation of the amyloid-beta protein (Abeta) in the cerebral cortex is an early and invariant event in the pathogenesis of Alzheimer's disease. The final step in the generation of Abeta from the beta-amyloid precursor protein is an apparently intramembranous proteolysis by the elusive gamma-secretase(s). The most common cause of familial Alzheimer's disease is mutation of the genes encoding presenilins 1 and 2, which alters gamma-secretase activity to increase the production of the highly amyloidogenic Abeta42 isoform. Moreover, deletion of presenilin-1 in mice greatly reduces gamma-secretase activity, indicating that presenilin-1 mediates most of this proteolytic event. Here we report that mutation of either of two conserved transmembrane (TM) aspartate residues in presenilin-1, Asp 257 (in TM6) and Asp 385 (in TM7), substantially reduces Abeta production and increases the amounts of the carboxy-terminal fragments of beta-amyloid precursor protein that are the substrates of gamma-secretase. We observed these effects in three different cell lines as well as in cell-free microsomes. Either of the Asp --> Ala mutations also prevented the normal endoproteolysis of presenilin-1 in the TM6 --> TM7 cytoplasmic loop. In a functional presenilin-1 variant (carrying a deletion in exon 9) that is associated with familial Alzheimer's disease and which does not require this cleavage, the Asp 385 --> Ala mutation still inhibited gamma-secretase activity. Our results indicate that the two transmembrane aspartate residues are critical for both presenilin-1 endoproteolysis and gamma-secretase activity, and suggest that presenilin 1 is either a unique diaspartyl cofactor for gamma-secretase or is itself gamma-secretase, an autoactivated intramembranous aspartyl protease.     

应用于连续时间∑Δ调制器的时钟抖动建模方法

提出了一种能够快速而精确地模拟时钟抖动的建模方法,可应用于连续时间Sigma-Delta调制器(continuous-time sigma-delta-modulator,CT-SDM)等系统的仿真与验证。相较于传统的基于离散时间的建模方法,所提出的一种基于连续时间的模型,可以灵活地应用于各种连续时间电路中,且可在保证精度的情况下,快速完成仿真。给出了关于时钟抖动的理论分析和该模型的数学理论推导,并通过搭建一个完整的连续时间Sigma-Delta调制器,验证了所提时钟抖动方法的正确性与可行性,仿真时间在数十秒内。     

近红外波段硅基石墨烯电光调制器研究进展

 近红外波段的电光调制器是未来光信号处理和计算系统中的关键功能元器件,硅基石墨烯电光调制器在结构尺寸、调制速率、调制带宽及大规模片上集成等方面具有诸多潜在优点而引起人们的广泛关注和重视。本文介绍了石墨烯的光电特性及光调制机理,结合石墨烯在近红外波段电光调制器中的研究及应用,综述了国内外近红外波段硅基石墨烯电光调制器的研究进展,重点叙述了条形波导结构、谐振结构、纳米梁结构的电光调制器的工作原理及各器件的特性,展望了硅基石墨烯电光调制器的研究方向。     

TMP21 is a presenilin complex component that modulates gamma-secretase but not epsilon-secretase activity

The presenilin proteins (PS1 and PS2) and their interacting partners nicastrin, aph-1 (refs 4, 5) and pen-2 (ref. 5) form a series of high-molecular-mass, membrane-bound protein complexes that are necessary for gamma-secretase and epsilon-secretase cleavage of selected type 1 transmembrane proteins, including the amyloid precursor protein, Notch and cadherins. Modest cleavage activity can be generated by reconstituting these four proteins in yeast and Spodoptera frugiperda (sf9) cells. However, a critical but unanswered question about the biology of the presenilin complexes is how their activity is modulated in terms of substrate specificity and/or relative activities at the gamma and epsilon sites. A corollary to this question is whether additional proteins in the presenilin complexes might subsume these putative regulatory functions. The hypothesis that additional proteins might exist in the presenilin complexes is supported by the fact that enzymatically active complexes have a mass that is much greater than predicted for a 1:1:1:1 stoichiometric complex (at least 650 kDa observed, compared with about 220 kDa predicted). To address these questions we undertook a search for presenilin-interacting proteins that differentially affected gamma- and epsilon-site cleavage events. Here we report that TMP21, a member of the p24 cargo protein family, is a component of presenilin complexes and differentially regulates gamma-secretase cleavage without affecting epsilon-secretase activity.     

基于电吸收调制器的新型40 Gbit/s光学3R重生器

全光3R重生技术（重整形、重定时、重放大）是将来高速大容量全光网中的关键技术.提出了一种结构简单而且性能稳定的40 Gbit/s光学3R重生器,它是基于电吸收调制器的2个波长转换器与基于行波电吸收调制器的光钟恢复器构成的,具有造价低、性能稳定等优点,在未来大容量超高速全光网中具有很大的应用潜力.     

基于电吸收调制器的新型40 Gbit/s光学3R重生器

全光3R重生技术（重整形、重定时、重放大）是将来高速大容量全光网中的关键技术。提出了一种结构简单而且性能稳定的40 Gbit/s光学3R重生器，它是基于电吸收调制器的2个波长转换器与基于行波电吸收调制器的光钟恢复器构成的，具有造价低、性能稳定等优点，在未来大容量超高速全光网中具有很大的应用潜力。     

基于Systemview的新型数字带通调制系统的设计及仿真

为了提高数字带通传输系统的性能,人们对数字调制系统不断加以改进,提出了多种新的调制体系。这些新型调制解调系统的某些硬件电路可以用软件实现,从而节省设计时间,缩短设计周期,提高设计效率。利用SystemView软件对通信系统中的正交振幅键控和最小频移键控进行了设计和仿真,仿真结果正确。     

Effects of modulators of myosin light-chain kinase activity in single smooth muscle cells

Phosphorylation of myosin light chains by a calmodulin-myosin light-chain kinase (MLCK) pathway is considered to be responsible for coupling increased calcium concentration with contraction in smooth muscle. This simple view has, however, recently been questioned. To test this hypothesis directly, we microinjected individual smooth muscle cells with modulators of the MLCK pathway while measuring contraction and calcium-ion concentration. Injection of a constitutively active proteolyzed form of MLCK causes contraction but no change in calcium concentration. By contrast, injection of peptide inhibitors of MLCK blocks contraction in response to K+ depolarization, despite the fact that the change in calcium concentration in response to stimulation was enhanced over controls. These results provide a direct demonstration at the level of a single cell that activation of the calmodulin-MLCK pathway is both necessary and sufficient to trigger contraction of smooth muscle.     

Viral immune modulators perturb the human molecular network by common and unique strategies

Viruses must enter host cells to replicate, assemble and propagate. Because of the restricted size of their genomes, viruses have had to evolve efficient ways of exploiting host cell processes to promote their own life cycles and also to escape host immune defence mechanisms. Many viral open reading frames (viORFs) with immune-modulating functions essential for productive viral growth have been identified across a range of viral classes. However, there has been no comprehensive study to identify the host factors with which these viORFs interact for a global perspective of viral perturbation strategies. Here we show that different viral perturbation patterns of the host molecular defence network can be deduced from a mass-spectrometry-based host-factor survey in a defined human cellular system by using 70 innate immune-modulating viORFs from 30 viral species. The 579 host proteins targeted by the viORFs mapped to an unexpectedly large number of signalling pathways and cellular processes, suggesting yet unknown mechanisms of antiviral immunity. We further experimentally verified the targets heterogeneous nuclear ribonucleoprotein?U, phosphatidylinositol-3-OH kinase, the WNK (with-no-lysine) kinase family and USP19 (ubiquitin-specific peptidase 19) as vulnerable nodes in the host cellular defence system. Evaluation of the impact of viral immune modulators on the host molecular network revealed perturbation strategies used by individual viruses and by viral classes. Our data are also valuable for the design of broad and specific antiviral therapies.     

Arachidonic acid metabolites as intracellular modulators of the G protein-gated cardiac K+ channel

Arachidonic acid is released from cell membranes in response to receptor-dependent as well as receptor-independent stimulation in various cells, including cardiac myocytes. Arachidonic acid is converted to prostaglandins by cyclooxygenase and to leukotrienes by 5-lipoxygenase, metabolites which are very biologically active and modulate cellular functions such as platelet aggregation, smooth muscle contraction and neural excitation. The molecular mechanisms underlying their modulations are, however, still badly understood. Here, we report that the 5-lipoxygenase metabolites of arachidonic acid activate the pertussis toxin-sensitive G protein-gated muscarinic K+ channel (IK.ACh): arachidonic acid activation of IK.ACh was prevented by the lipoxygenase inhibitors, nordihydroguaiaretic acid and AA-861; leukotriene A4 and C4 activated IK.ACh. The activation occurred in pertussis toxin-treated atrial cells and ceased when inside-out patches were formed but the patches were still susceptible to stimulation by GTP and to inhibition by GDP-beta-S. These results indicate that arachidonic acid metabolites may stimulate the G-protein in a receptor-independent way.     

Development of a functional cell-based HTS assay for identification of NKCC1-negative modulators

Na–K–Cl cotransporter 1(NKCC1) cotransports Na+, K+, and Cl-ions across the plasma membrane into cells. Accumulation of Cl-ions in dorsal root ganglion neurons induces depolarizing GABAA receptors, which mediate presynaptic inhibition and filtration of sensory noise. The activity of the Na–K–Cl cotransporter is modulated by high-dose loop diuretics, such as furosemide and bumetanide. To identify NKCC1 modulators, we developed a functional cell-based assay feasible for highthroughput screening(HTS), in which the activity of NKCC1 was detected by a BTC-AM dye-based thallium transportation assay. We demonstrated that the influx of Tl?was mediated by NKCC1, which required the existence of Cl-ions and could be inhibited by bumetanide and furosemide. Our results demonstrated that the assay was stable, reproducible, and suitable for HTS of negative modulators for NKCC1.