高等植物中的磷酸烯醇式丙酮酸羧激酶

简要介绍了近年来有关高等植物中磷酸烯醇式丙酮酸羧激酶（ＰＥＰＣＫ）的研究进展,并讨论了此酶的结构、功能和调节等方面的问题。     

中国人扩张型心肌病与受磷蛋白基因关系的研究

受磷蛋白（phospholamban）是心肌收缩的一个重要调节因子,可抑制心肌肌浆网钙ATPase的活性、降低其对钙的亲和力。正常情况下,受磷蛋白可被不同的蛋白激酶磷酸化从而解除其肌浆网钙ATPase的抑制作用。国外两个DCM家系研究发现受磷蛋白基因突变与DCM的发生有关,研究目的旨在探讨中国人群心肌特异性受磷蛋白基因突变与特发性扩张型心肌病发病的关系。收集60例确诊的特发性扩张型心肌病患者临床资料,采集血样本并分离白细胞和提取基因组DNA。应用PCR扩增肌特异性受磷蛋白基因片段,经测序检测基因突变的存在。结果显示60例扩张型心肌病人受磷蛋白基因开放阅读框未发现突变,仅有两例患者出现阅读框3′部分非翻译区分别离终止密码153和173位单个碱基T的缺失。两例患者终止密码后非翻译区单个碱基T的缺失据分析并无实际意义。因此我们认为绝大部分中国人扩张型心肌病的发病可能与受磷蛋白基因突变无关。Abstract: Phospholamban (PLB) is a prominent regulator of myocardial contractility and a reversible inhibitor of the cardiac sarcoplasmic reticulum Ca2+ ATPase activity. In normal cardiac muscles, phospholamban can be phosphorylated at distinct sites by various protein kinases and release its inhibition to sarcoplasmic reticulum Ca2+ ATPase. The studies of pedigrees have shown dilated cardiomyopathy (DCM) is related with mutation of PLB. The aim of present study is to investigate the relationship between mutation of PLB gene and DCM. Sixty patients with idiotic DCM were enrolled in present study. The clinical data were collected, including clinical symptoms, ECG and echocardiography. Peripheral blood samples of all these subjects were collected to extract genome DNA. The fragments of PLB gene were amplified by PCR and PCR fragment sequencing was performed to study weather mutation of phospholamban gene exists. phospholamban gene did not show any mutation in these patients. Most Chinese DCM patients may not be related with mutation of PLB gene.     

饲料铜水平对斑节对虾血细胞中酚氧化酶原系统相关基因表达的影响

以不同铜离子添加水平(0 mg·kg-1、10 mg·kg-1、25 mg·kg-1、40 mg·kg-1、55 mg·kg-1和110 mg·kg-1)的饲料喂养斑节对虾Penaeus monodon 8周,测定对虾血细胞中酚氧化酶原系统相关基因(pro PO1、pro PO2、PE、PPAF和BGBP)表达水平的变化。结果显示,与对照组相比,当铜添加水平为40~55 mg·kg-1时,proPO1的表达量显著上升;当铜添加水平为25~55 mg·kg-1时,pro PO2、PE和PPAF的表达量显著上升;铜添加水平对BGBP的表达量没有显著影响。血清PO活力也在铜添加水平为25~55 mg·kg-1时显著提高。这些结果表明,饲料中添加一定量的铜离子可诱导pro PO1、pro PO2、PE和PPAF表达量的上调,从而提高血清PO活力。根据上述结果,针对提高酚氧化酶原系统免疫功能,在本基础饲料配方中适宜的铜添加水平为25~55 mg·kg-1。     

兰州市主要绿化植物气孔性状特征

气孔是植物叶片与外界环境进行水汽交换的门户, 利用气孔特征反映植物对环境变化的响应与适应有助于了解干旱区绿化植物的适应策略。于2019年7月通过气孔印迹法对兰州市40种主要绿化植物气孔性状进行观察与测定。采用标准化主轴估计和系统独立比较分析建立气孔性状间的相关关系; 通过计算Blomgerg’sK值以检验系统发育信号; 利用聚类分析和主成分分析对气孔性状特征进行功能群划分。结果表明: (1)在所研究的植物中, 气孔性状特征在个体间差异显著, 植物生长型(乔木、小乔木、灌木和草本)会显著影响气孔长度、宽度、开度和密度, 叶习性(落叶和常绿)仅对气孔开度有显著影响; (2)气孔长度与宽度、气孔开度与面积间均为显著的异速生长关系, 气孔密度与面积和开度间为负异速生长关系; (3)系统发育会对气孔性状的相关关系产生显著影响, 虽然气孔性状的系统发育保守性不强(K < 1), 但气孔开度和气孔开张比具有显著的系统发育信号; 4)依据气孔性状特征可以将绿化植物划分为: 低密度-大面积、高密度-小面积和中等密度-适中面积共3种功能群。结合系统发育和异速生长理论能更好地解释气孔性状变异及适应策略。     

Enhanced Gene Transduction into Skeletal Muscle of Mice In Vivo with Pluronic Block Copolymers and Ultrasound Exposure

The pluronic block copolymers are able to enhance the ultrasound-induced gene delivery in vitro. In the present study, the effects of pluronics on the efficiency of gene transfer into skeletal muscle in vivo under sonoporation were investigated. Plasmid DNA encoding green fluorescent protein (GFP) in combination with three different pluronics, F127, L61, and P85, was injected into the tibialis anterior (TA) muscle of mice with and without adjunct ultrasound (1 MHz, 3 W/cm² 1 min, 20% duty cycle). Mice were killed 1 week after injection. The TA muscles were removed and snap frozen immediately in isopentane cooled by liquid nitrogen and sections of 7 μm thick were cut. Transfection efficiency was assessed by counting the number of GFP-positive fibers under fluorescence microscopy, and tissue damage by hematoxylin and eosin staining. The results suggested that all three pluronics significantly enhanced transgene expression in skeletal muscle (P < 0.01), especially the P85 showed significantly higher efficiency than the other two pluronics (P < 0.05). Ultrasound synergistically enhanced the gene delivery efficiency with P85 (P < 0.01), but was unable to do so with F127 and L61 groups. In short, P85 displays significantly synergistic effect with ultrasound for enhancing plasmid DNA transduction in skeletal muscle of mice in vivo.     

A transferable heterogeneous two-hybrid system in <Emphasis Type="Italic">Escherichia coli</Emphasis> based on polyhydroxyalkanoates synthesis regulatory protein PhaR

Background  

Polyhydroxyalkanoate (PHA) synthesis regulatory protein PhaR contains a DNA binding domain (DBD) and a PHA granule binding domain (GBD), it anchors to the promoter region of PHA granule-associated protein (PhaP) to repress phaP expression. However, PhaR will bind to PHB granules and be released from phaP promoter region when PHA granules are formed in vivo, initiating expression of phaP gene. Based on this regulatory mechanism, a bacterial two-hybrid system was developed: PhaR was separated into two parts: DBD was used to fuse with the bait, GBD with the prey, and phaP was replaced by a reporter gene lacZ. However, GBD protein expressed in vivo formed inclusion bodies. Thus, PhaP with strong binding ability to PHB granules was employed to replace GBD.     

Metschnikowia henanensis sp. nov., a new anamorphic yeast species isolated from rotten wood in China

Four yeast strains were isolated from rotting wood samples collected from two sites in the Baotianman Nature Reserve and the Laojieling Nature Reserve in China. DNA sequence comparison and other taxonomic characteristics identified the strains as a single novel species of the genus Metschnikowia. The name Metschnikowia henanensis sp. nov. is proposed to accommodate these highly divergent organisms with the type strain BY-97^T (= CICC 1982^T = CBS 12677^T). The novel species produced chlamydospores, but it did not exhibit ascospore formation in sporulation media for 4 weeks. Molecular phylogeny from the D1/D2 domains of the large subunit (LSU) rRNA gene sequences placed this new species in a basal position to the Metschnikowia viticola/Candida kofuensis/Metschnikowia noctiluminum subclade, and an undescribed Candida species namely strains IMB-EMP4 and IMB-EMP5 was a close sister to M. henanensis.     

The Oxidative Damage and Disbalance of Calcium Homeostasis in Brain of Chicken Induced by Selenium Deficiency

Dietary selenium (Se) deficiency can influence the function of the brain. Our objective was to investigate the effects of Se deficiency on oxidative damage and calcium (Ca) homeostasis in brain of chicken. In the present study, 1-day-old chickens were fed either a commercial diet (as control group) with 0.15 mg/kg Se or a Se-deficient diet (as L group) with 0.033 mg/kg Se for 75 days. Then, brain injury biomarkers were examined, including histological analysis, ultrastructure assay, and apoptosis assay. We also examined the effect of Se deficiency on the Se-containing antioxidative enzyme glutathione peroxidase (GSH-Px), the level of glutathione (GSH), and the Ca homeostasis in brain of chicken. The results showed that the levels of Se and GSH and activity of GSH-Px are seriously reduced by 33.8–96 % (P?<?0.001), 24.51–27.84 % (P?<?0.001), and 20.70–64.24 % (P?<?0.01), respectively. In the present study, we also perform histological analysis and ultrastructure assay and find that Se deficiency caused disorganized histological structure, damage to the mitochondria, fusion of nuclear membrane and nucleus shrinkage, higher apoptosis rate (P?<?0.001), and increase of Ca homeostasis (P?<?0.05 or P?<?0.01 or P?<?0.001) in the brain of chicken. In conclusion, the results demonstrated that Se deficiency induced oxidative damage and disbalance of Ca homeostasis in the brain of chicken. Similar to mammals, chickens brain is also extremely susceptible to oxidative damage and selenium deficiency.     

Mitochondrial DNA polymorphisms in Phytophthora infestans: New haplotypes are identified and re-defined by PCR

Polymorphisms of mitochondrial DNA (mt-DNA) are particularly useful for monitoring specific pathogen populations like Phytophthora infestans. Basically type I and II of P. infestans mt-DNA were categorized by means of polymorphism lengths caused by an ~ 2 kb insertion, which can be detected via restriction enzyme digestion. In addition genome sequencing of haplotype Ib has been used as a simple Polymerase Chain Reaction–Restriction Fragment Length Polymorphism (PCR–RFLP) method to indirectly identify type I and II alterations through EcoR I restriction enzyme DNA fragment patterns of the genomic P4 area. However, with the common method, wrong mt-DNA typing occurs due to an EcoR I recognition site mutation in the P4 genomic area. Genome sequencing of the four haplotypes (Ia, Ib, IIa, and IIb) allowed us to thoroughly examine mt-DNA polymorphisms and we indentified two hypervariable regions (HVRs) named HVRi and HVRii. The HVRi length polymorphism caused by a 2 kb insertion/deletion was utilized to identify mt-DNA types I and II, while another length polymorphism in the HVRii region is caused by a variable number of tandem repeats (n = 1, 2, or 3) of a 36 bp sized DNA stretch and was further used to determine mt-DNA sub-types, which were described as R_{n=1, 2, or 3}. Finally, the P. infestans mt-DNA haplotypes were re-defined as IR₁ or IIR₂ according to PCR derived HVRi and HVRii length polymorphisms. Twenty-three isolates were chosen to verify the feasibility of our new approach for identifying mt-DNA haplotypes and a total of five haplotypes (IR₁, IR₂, IR₃, IIR₂ and IIR₃) were identified. Additionally, we found that six isolates determined as type I by our method were mistakenly identified as type II by the PCR–RFLP technique. In conclusion, we propose a simple and rapid PCR method for identification of mt-DNA haplotypes based on sequence analyses of the mitochondrial P. infestans genome.     

Enhanced remediation of black liquor by activated sludge bioaugmented with a novel exogenous microorganism culture

Black liquor (BL) is a notoriously difficult wastewater to treat due to the economic and efficiency limitations of physiochemical methods and intrinsic difficulties with bioremediation strategies caused by the high pH (10–13) and lignin content. This study investigated the feasibility of a novel bioaugmentation strategy for BL treatment, which uses a mixed microorganism culture of lignocellulose-degrading microorganisms isolated from degraded bamboo slips. Black liquor treatment was assessed in terms of chemical oxygen demand (COD) and color removal with a sequencing batch reactor organic loading rate of 9 kg COD/L·day under highly alkaline conditions (pH?10). Results revealed that bioaugmented activated sludge treatment of BL with special mixed microorganisms significantly enhanced the removal efficiency of COD, color, and lignin from the wastewater up to 64.8, 50.5, and 53.2 %, respectively. Gel permeation chromatography profiles showed that the bioaugmentation system could successfully degrade high molecular lignin fragments in black liquor. This work confirms bioaugmentation as a feasible alternative strategy for enhanced biological treatment of wastewater with high lignin content and high organic load rate under strongly alkaline conditions.