The PRiMA-linked Cholinesterase Tetramers Are Assembled from Homodimers: HYBRID MOLECULES COMPOSED OF ACETYLCHOLINESTERASE AND BUTYRYLCHOLINESTERASE DIMERS ARE UP-REGULATED DURING DEVELOPMENT OF CHICKEN BRAIN*

Acetylcholinesterase (AChE) is anchored onto cell membranes by the transmembrane protein PRiMA (proline-rich membrane anchor) as a tetrameric globular form that is prominently expressed in vertebrate brain. In parallel, the PRiMA-linked tetrameric butyrylcholinesterase (BChE) is also found in the brain. A single type of AChE-BChE hybrid tetramer was formed in cell cultures by co-transfection of cDNAs encoding AChE_T and BChE_T with proline-rich attachment domain-containing proteins, PRiMA I, PRiMA II, or a fragment of ColQ having a C-terminal GPI addition signal (Q_N-GPI). Using AChE and BChE mutants, we showed that AChE-BChE hybrids linked with PRiMA or Q_N-GPI always consist of AChE_T and BChE_T homodimers. The dimer formation of AChE_T and BChE_T depends on the catalytic domains, and the assembly of tetramers with a proline-rich attachment domain-containing protein requires the presence of C-terminal “t-peptides” in cholinesterase subunits. Our results indicate that PRiMA- or ColQ-linked cholinesterase tetramers are assembled from AChE_T or BChE_T homodimers. Moreover, the PRiMA-linked AChE-BChE hybrids occur naturally in chicken brain, and their expression increases during development, suggesting that they might play a role in cholinergic neurotransmission.     

副溶血性弧菌免疫逃逸及其可能机制的研究进展

副溶血性弧菌是全球范围内威胁人体健康和食品安全的食源性致病菌。在感染人体的过程中,副溶血性弧菌通过将其效应蛋白直接注射至宿主细胞中操纵宿主,介导毒力的发挥,并进化出了一套完美的免疫逃逸策略,成功躲避免疫系统的攻击,引起急性肠胃炎、败血症和坏死性筋膜炎等疾病。副溶血性弧菌入侵上皮细胞,使胞内囊泡酸化,在与溶酶体融合之前逃逸到细胞质中,并且限制活性氧的产生,促进其在胞内生存。副溶血性弧菌可以诱导自噬,抑制NLRC4炎症小体介导的caspase-1的激活,还可以通过抑制TAK1激酶,阻止MAPK和NF-κB信号通路的激活,干扰免疫系统激活,借助多种手段共同协作从而达到免疫逃逸。本文系统总结了副溶血性弧菌现已研究的免疫逃逸机制,并对其可能存在的免疫逃逸机制提供了新的见解和方向,对深入了解副溶血性弧菌的致病机理和防控药物靶向位点的选择及研发具有重要意义。     

Generation and Analysis of ESTs from the Grass Carp,Ctenopharyngodon idellus

Grass carp, Ctenopharyngodon idellus (Valenciennes, 1844), is an economically important species widely cultured in the world, but its genome research resources are largely lacking. The objectives of this study were to construct normalized cDNA libraries for efficient EST analysis, to generate ESTs from these libraries, and to identify EST-related molecular markers such as microsatellites and single nucleotide polymorphisms (SNPs) for genetic analysis of this species. A total of 6,269 ESTs were generated representing 4,815 unique sequences, from which 105 putative microsatellites and 5,228 SNPs were identified. These genome resources provide the material basis for future genetic and functional analyses in this species.     

Differential effect of three-repeat and four-repeat tau on mitochondrial axonal transport

Tau protein is present in six different splice forms in the human brain and interacts with microtubules via either 3 or 4 microtubule binding repeats. An increased ratio of 3 repeat to 4 repeat isoforms is associated with neurodegeneration in inherited forms of frontotemporal dementia. Tau over-expression diminishes axonal transport in several systems, but differential effects of 3 repeat and 4 repeat isoforms have not been studied. We examined the effects of tau on mitochondrial transport and found that both 3 repeat and 4 repeat tau change normal mitochondrial distribution within the cell body and reduce mitochondrial localization to axons; 4 repeat tau has a greater effect than 3 repeat tau. Further, we observed that the 3 repeat and 4 repeat tau cause different alterations in retrograde and anterograde transport dynamics with 3 repeat tau having a slightly stronger effect on axon transport dynamics. Our results indicate that tau-induced changes in axonal transport may be an underlying theme in neurodegenerative diseases associated with isoform specific changes in tau's interaction with microtubules.     

Optimization of recombinant aminolevulinate synthase production in<Emphasis Type="Italic"> Escherichia coli</Emphasis> using factorial design

The production of recombinant Rhodobacter sphaeroides aminolevulinate (ALA) synthase was optimized in two strains of Escherichia coli: the wild-type strain MG1655, and a ptsG mutant AFP111. The effects of initial succinate, glucose and isopropyl--d-thiogalactopyranoside (IPTG) concentrations and the time of induction on enzyme activity were studied. One-way analysis was used to approximate the optimal ranges for these factors, followed by a full factorial design to quantify the effects of each factor and the interactions between the factors. Initial succinate, glucose, and IPTG concentration were observed to be the key factors affecting ALA synthase activity with the optimal levels determined to be above 6 g/l succinate, 0 g/l glucose, and 0.10 mM IPTG. ALA synthase activity was generally lower with AFP111 than with MG1655, and the effect of these three key factors was also lower with AFP111 than with MG1655. Based on the full factorial design results, a fermentation was completed that yielded 296 mU/mg protein with a final ALA concentration of 5.2 g/l (39 mM).