APP substitutions V715F and L720P alter PS1 conformation and differentially affect Abeta and AICD generation

The 37-43 amino acid Abeta peptide is the principal component of beta-amyloid deposits in Alzheimer's disease (AD) brain, and is derived by serial proteolysis of the amyloid precursor protein (APP) by beta- and gamma-secretase. gamma-Secretase also cleaves APP at Val50 in the Abeta numbering (epsilon cleavage), resulting in the release of a fragment called APP intracellular domain (AICD). The aim of this study was to determine whether amino acid substitutions in the APP transmembrane domain differentially affect Abeta and AICD generation. We found that the APPV715F substitution, which has been previously shown to dramatically decrease Abeta40 and Abeta42 while increasing Abeta38 levels, does not affect in vitro generation of AICD. Furthermore, we found that the APPL720P substitution, which has been previously shown to prevent in vitro generation of AICD, completely prevents Abeta generation. Using a fluorescence resonance energy transfer (FRET) method, we next found that both the APPV715F and APPL720P substitutions significantly increase the distance between the N- and C-terminus of presenilin 1 (PS1), which has been proposed to contain the catalytic site of gamma-secretase. In conclusion, both APPV715F and APPL720P change PS1 conformation with differential effects on Abeta and AICD production.     

A triple mutation in the second transmembrane domain of mouse dopamine transporter markedly decreases sensitivity to cocaine and methylphenidate

Previously, we reported that Phe105 in transmembrane domain 2 of the mouse dopamine transporter (DAT) is crucial for high-affinity cocaine binding. In the current study, we investigated whether other residues surrounding Phe105 also affect the potency of cocaine inhibition. After three rounds of sequential random mutagenesis at these residues, we found a triple mutant (L104V, F105C and A109V) of mouse DAT that retained over 50% uptake activity and was 69-fold less sensitive to cocaine inhibition when compared with the wild-type mouse DAT. The triple mutation also resulted in a 47-fold decrease in sensitivity to methylphenidate inhibition, suggesting that the binding sites for cocaine and methylphenidate may overlap. In contrast, the inhibition of dopamine uptake by amphetamine or methamphetamine was not significantly changed by the mutations, suggesting that the binding sites for the amphetamines differ from those for cocaine and methylphenidate. Such functional but cocaine-insensitive DAT mutants can be used to generate a knock-in mouse line to study the role of DAT in cocaine addiction.     

胸腔闭式引流联合尿激酶注入对包裹性胸腔积液的临床研究

目的:探讨胸腔闭式引流联合尿激酶注入对包裹性胸腔积液的临床疗效。方法:对我院2007年2月-2011年4月收治的包裹性胸腔积液患者87例,随机分为实验组以及对照组,实验组采用胸腔闭式引流联合尿激酶注入进行治疗,对照组采用常规治疗。结果:实验组患者临床疗效明显优于对照组(P<0.05);实验组患者治疗后其积液中蛋白量以及白细胞含量明显低于对照组(P<0.05);实验组患者治疗时间、胸膜壁厚度等比较明显优于对照组(P<0.05)。结论:对包裹性胸腔积液患者采用胸腔闭式引流联合尿激酶注入进行治疗,可有效改善患者预后,提高患者临床治疗效果。     

Design and synthesis of diarylamines and diarylethers as cytotoxic antitumor agents

Based on a shared structural core of diarylamine in several known anticancer drugs as well as a new cytotoxic hit 6-chloro-2-(4-cyanophenyl)amino-3-nitropyridine (7), 30 diarylamines and diarylethers were designed, synthesized, and evaluated for cytotoxic activity against A549, KB, KB-vin, and DU145 human tumor cell lines (HTCL). Four new leads 11e, 12, 13a, and 13b were discovered with GI(50) values ranging from 0.33 to 3.45μM. Preliminary SAR results revealed that a diarylamine or diarylether could serve as an active structural core, meta-chloro and ortho-nitro groups on the A-ring (either pyridine or phenyl ring) were necessary and crucial for cytotoxic activity, and the para-substituents on the other phenyl ring (B-ring) were related to inhibitory selectivity for different tumor cells. In an investigation of potential biological targets of the new leads, high thoughput kinase screening discovered that new leads 11e, 12 and 13b especially inhibit Mer tyrosine kinase, a proto-oncogene associated with munerous tumor types, with IC(50) values of 2.2-3.0μM. Therefore, these findings provide a good starting point to optimize a new class of compounds as potential anticancer agents, particularly targeting Mer tyrosine kinase.     

不同光强对吴茱萸生长动态和光合日变化的影响

报道了不同生长光强(透光率分别为15%、30%、50%、100%)对吴茱萸的生长和光合日变化的影响。结果表明:生长环境光强对吴茱萸苗期株高、地径和冠面积的影响较大,100%RI下的株高、地径和冠面积明显小于各遮荫处理,其中30%RI下吴茱萸的株高、地径和冠面积最大,这表明苗期强光不利于吴茱萸的生长。随着光强的减弱,吴茱萸增大冠面积与株高有利于截获更多光能。10月下旬,100%RI下吴茱萸净光合速率(Pn)日变化曲线呈"双峰型",11:30～14:30,吴茱萸的Pn、胞间二氧化碳浓度与气孔导度均下降,表明此时的光合午休现象是由气孔部分关闭造成的。各遮荫处理下,吴茱萸的Pn日变化均呈"单峰型",并无第二峰的出现,可能与遮荫条件下,下午光强较弱,环境温度较高,空气相对湿度较低有关。     

Self-renewal and Differentiation of Muscle Satellite Cells Are Regulated by the Fas-associated Death Domain

Making the decision between self-renewal and differentiation of adult stem cells is critical for tissue repair and homeostasis. Here we show that the apoptotic adaptor Fas-associated death domain (FADD) regulates the fate decisions of muscle satellite cells (SCs). FADD phosphorylation was specifically induced in cycling SCs, which was high in metaphase and declined in later anaphase. Furthermore, phosphorylated FADD at Ser-191 accumulated in the uncommitted cycling SCs and was asymmetrically localized in the self-renewing daughter SCs. SCs containing a phosphoryl-mimicking mutation at Ser-191 of FADD (FADD-D) expressed higher levels of stem-like markers and reduced commitment-associated markers. Moreover, a phosphoryl-mimicking mutation at Ser-191 of FADD suppressed SC activation and differentiation, which promoted the cycling SCs into a reversible quiescent state. Therefore, these data indicate that FADD regulates the fate determination of cycling SCs.     

Molecular mechanism of ERK dephosphorylation by striatal‐enriched protein tyrosine phosphatase

Striatal‐enriched tyrosine phosphatase (STEP) is an important regulator of neuronal synaptic plasticity, and its abnormal level or activity contributes to cognitive disorders. One crucial downstream effector and direct substrate of STEP is extracellular signal‐regulated protein kinase (ERK), which has important functions in spine stabilisation and action potential transmission. The inhibition of STEP activity toward phospho‐ERK has the potential to treat neuronal diseases, but the detailed mechanism underlying the dephosphorylation of phospho‐ERK by STEP is not known. Therefore, we examined STEP activity toward para‐nitrophenyl phosphate, phospho‐tyrosine‐containing peptides, and the full‐length phospho‐ERK protein using STEP mutants with different structural features. STEP was found to be a highly efficient ERK tyrosine phosphatase that required both its N‐terminal regulatory region and key residues in its active site. Specifically, both kinase interaction motif (KIM) and kinase‐specific sequence of STEP were required for ERK interaction. In addition to the N‐terminal kinase‐specific sequence region, S245, hydrophobic residues L249/L251, and basic residues R242/R243 located in the KIM region were important in controlling STEP activity toward phospho‐ERK. Further kinetic experiments revealed subtle structural differences between STEP and HePTP that affected the interactions of their KIMs with ERK. Moreover, STEP recognised specific positions of a phospho‐ERK peptide sequence through its active site, and the contact of STEP F311 with phospho‐ERK V205 and T207 were crucial interactions. Taken together, our results not only provide the information for interactions between ERK and STEP, but will also help in the development of specific strategies to target STEP‐ERK recognition, which could serve as a potential therapy for neurological disorders.