1H, 13C, and 15N resonance assignments of FK506-binding domain of Plasmodium falciparum FKBP35

The immunosuppressant FK506 binds Plasmodium falciparum FK-506 binding protein 35 (PfFKBP35) and shows anti-malarial activity. To understand molecular mechanism of the drug on the parasite, we have done NMR studies. Here, we report the assignment of FK506-binding domain of PfFKBP35.     

Somaclonal variation at the nucleotide sequence level in rice (<Emphasis Type="Italic">Oryza sativa</Emphasis> L.) as revealed by RAPD and ISSR markers,and by pairwise sequence analysis

The nature of somaclonal variation at the nucleotide sequence level was studied in rice cv. Nipponbare. First, we investigated genomic variations by using 2 molecular marker systems: RAPD (random amplified polymorphic DNA) and ISSR (inter-simple sequence repeat). This was followed by sequencing of selected bands that represented genomic variations, and pairwise sequence analysis taking advantage of the whole genome sequence of rice. In addition, transpositional activity of the active MITE, mPing, was analysed by locus-specific PCR amplifications. The 2-year-old calli and their regenerated plants, analysed with 24 RAPD and 20 ISSR primers, showed moderate levels of genomic variation (20.83% and 17.04%, respectively). To test whether DNA methylation plays a role in somaclonal variation, the calli were treated with 5-azacytidine, a chemical agent that reduces cytosine methylation by blocking the activity of DNA methyltransferase. Though dwarfism occurred in regenerants from treated calli (a hallmark of the drug treatment), there was only a slight increase in the frequency of somaclonal variation detected in the treated calli and their regenerated plants relative to untreated controls. The transposon mPing also remained immobile in both treated and untreated calli. Nevertheless, dendrograms constructed according to the Jaccard coefficient calculated by UPGMA of the ISSR bands revealed that the 5-azacytidine-treated and untreated somaclones were grouped into 2 distinct clusters, suggesting a possible indirect effect of the treatment on the genomic changes, depending on the marker used. Sequence analysis indicated a low level of variation (0.31%), with single-base-pair substitutions predominating.     

Fosmid Library Construction and Initial Analysis of End Sequences in Zhikong Scallop (<Emphasis Type="Italic">Chlamys farreri</Emphasis>)

Zhikong scallop (Chlamys farreri Jones et Preston, 1904) is one of the most commercially important bivalves in China, but research on its genome is underdeveloped. In this study, we constructed the first Zhikong scallop fosmid library, and analyzed the fosmid end sequences to provide a preliminary assessment of the genome. The library consists of 133,851 clones with an average insert size of about 40 kb, amounting to 4.3 genome equivalents. Fosmid stability assays indicate that Zhikong scallop DNA was stable during propagation in the fosmid system. Library screening with two genes and seven microsatellite markers yielded between two and eight positive clones, and none of those tested was absent from the library. End-sequencing of 480 individual clones generated 828 sequences after trimming, with an average sequence length of 624 bp. BLASTN searches of the nr and EST databases of GenBank and BLASTX searches of the nr database resulted in 213 (25.72%) and 44 (5.31%) significant hits (E < e⁻⁵), respectively. Repetitive sequences analysis resulted in 375 repeats, accounting for 15.84% of total length, which were composed of interspersed repetitive sequences, tandem repeats, and low-complexity sequences. The fosmid library, in conjunction with the fosmid end sequences, will serve as a useful resource for physical mapping and positional cloning, and provide a better understanding of the Zhikong scallop genome.     

Binding properties of palmatine to DNA: spectroscopic and molecular modeling investigations

Palmatine, an isoquinoline alkaloid, is an important medicinal herbal extract with diverse pharmacological and biological properties. In this work, spectroscopic and molecular modeling approaches were employed to reveal the interaction between palmatine and DNA isolated from herring sperm. The absorption spectra and iodide quenching results indicated that groove binding was the main binding mode of palmatine to DNA. Fluorescence studies indicated that the binding constant (K) of palmatine and DNA was ~ 10⁴ L·mol^?1. The associated thermodynamic parameters, ΔG, ΔH, and ΔS, indicated that hydrogen bonds and van der Waals forces played major roles in the interaction. The effects of chemical denaturant, thermal denaturation and pH on the interaction were investigated and provided further support for the groove binding mode. In addition to experimental approaches, molecular modeling was conducted to verify binding pattern of palmatine–DNA. Copyright © 2015 John Wiley & Sons, Ltd.     

Nuclear translocation of PKM2 modulates astrocyte proliferation via p27 and β-catenin pathway after spinal cord injury

Aberrant functionality of the cell cycle has been implicated in the pathology of traumatic SCI. Although it has been reported that the expressions of various cell cycle related proteins were altered significantly following SCI, detailed information on the subject remains largely unclear. The embryonic pyruvate kinase M2 (PKM2) is an important metabolic kinase in aerobic glycolysis or the warburg effect, however, its functions in central nervous system (CNS) injury remains elusive. Here we demonstrate that PKM2 was not only significantly upregulated by western blot and immunohistochemistry but certain traumatic stimuli also induced translocation of PKM2 into the nucleus in astrocytes following spinal cord injury (SCI). Furthermore, the expression levels and localization of p-β-catenin, p27, cyclin D1 and PCNA were correlated with PKM2 after SCI. In vitro, we also found that PKM2 co-immunoprecipitation with p-β-catenin and p27 respectively. Knockdown of PKM2 apparently decreased the level of PCNA, cyclinD1, p27 in primary astrocyte cells. Taken together, our findings indicate that nuclear translocation of PKM2 promotes astrocytes proliferation after SCI through modulating cell cycle signaling. These discoveries firstly uncovered the role of PKM2 in spinal cord injury and provided a potential therapeutic target for CNS injury and repair.     

Curcumin inhibits hypoxia-induced migration in K1 papillary thyroid cancer cells

Curcumin, traditionally used as food and medicinal purposes, has recently been reported to have protective efficacy against hypoxia. Hypoxia is one of the important reactive factors in tumor metastasis, which is a key problem in clinical thyroid cancer therapy. In present study, we investigate the anti-metastatic effect of curcumin on the K1 papillary thyroid cancer cells as well as its potential mechanisms. The results show that curcumin effectively inhibits hypoxia-induced reactive oxygen species (ROS) upregulation and significantly decreases the mRNA and protein expression levels of hypoxia-inducible factor-1α (HIF-1α) in K1 cells. Curcumin also decreases the DNA binding ability of HIF-1α to hypoxia response element (HRE). Furthermore, curcumin enhances E-cadherin expression, inhibits metalloproteinase-9 (MMP-9) enzyme activity, and weakens K1 cells migration under hypoxic conditions. In summary, these results indicate that curcumin possesses a potent anti-metastatic effect and might be an effective tumoristatic agent for the treatment of aggressive papillary thyroid cancers.     

药用野生稻叶中淀粉合成酶基因家族的序列分化和特异表达

淀粉不仅是植物自身和后代生长繁殖的重要营养与能量储备,而且是人类膳食中碳水化合物的主要来源。植物中淀粉合成主要发生在两个阶段,一是在形成临时淀粉的光合作用阶段,另一个则是在成为贮藏淀粉的营养积累阶段。相对于最后的淀粉贮藏阶段,临时淀粉的形成阶段在植物整个碳水化合物代谢过程中扮演着更为重要的角色,然而却一直少有关注。为深入研究初始淀粉合成过程中相关酶在植物中的进化模式,选取了药用野生稻(Oryza officinalis)为研究对象,通过对其全叶转录组的重测序,定性、定量地调查了淀粉合成酶基因家族在稻属野生物种光合器官中的基因类型和表达变化。共有8个淀粉合成酶基因的完整编码序列在药用野生稻的叶中首次被识别。系统发育分析表明,这8个基因分别隶属SSI、SSII、SSIII、SSIV、SSV和GBSSII基因家族。序列比较和相对表达定量分析显示,药用野生稻与栽培稻的淀粉合成酶基因家族的进化模式具有高度的一致性,两个物种的同源基因在m RNA水平的序列相似度达到95%–98%。基于非同义置换和同义置换比率的统计检验表明,8个基因在两个物种间均经历了严格的纯化选择。另外,3个在栽培稻胚乳中特异表达的基因在药用野生稻的叶转录组中未筛查出来,而4个在栽培稻叶中优势表达的基因在药用野生稻叶中同样呈现相对较高水平的表达。     

SINC,a type III secreted protein of Chlamydia psittaci,targets the inner nuclear membrane of infected cells and uninfected neighbors

SINC, a new type III secreted protein of the avian and human pathogen Chlamydia psittaci, uniquely targets the nuclear envelope of C. psittaci–infected cells and uninfected neighboring cells. Digitonin-permeabilization studies of SINC-GFP–transfected HeLa cells indicate that SINC targets the inner nuclear membrane. SINC localization at the nuclear envelope was blocked by importazole, confirming SINC import into the nucleus. Candidate partners were identified by proximity to biotin ligase-fused SINC in HEK293 cells and mass spectrometry (BioID). This strategy identified 22 candidates with high confidence, including the nucleoporin ELYS, lamin B1, and four proteins (emerin, MAN1, LAP1, and LBR) of the inner nuclear membrane, suggesting that SINC interacts with host proteins that control nuclear structure, signaling, chromatin organization, and gene silencing. GFP-SINC association with the native LEM-domain protein emerin, a conserved component of nuclear “lamina” structure, or with a complex containing emerin was confirmed by GFP pull down. Our findings identify SINC as a novel bacterial protein that targets the nuclear envelope with the capability of globally altering nuclear envelope functions in the infected host cell and neighboring uninfected cells. These properties may contribute to the aggressive virulence of C. psittaci.