Doubly Spin-Labeled RNA as an EPR Reporter for Studying Multicomponent Supramolecular Assemblies

mRNAs are involved in complicated supramolecular complexes with human 40S and 80S ribosomes responsible for the protein synthesis. In this work, a derivative of nonaribonucleotide pUUCGUAAAA with nitroxide spin labels attached to the 5′-phosphate and to the C8 atom of the adenosine in sixth position (mRNA analog) was used for studying such complexes using double electron-electron resonance/pulsed electron-electron double resonance spectroscopy. The complexes were assembled with participation of tRNA^Phe, which targeted triplet UUC of the derivative to the ribosomal peptidyl site and predetermined location of the adjacent GUA triplet coding for Val at the aminoacyl (A) site. The interspin distances were measured between the two labels of mRNA analog attached to the first nucleotide of the peptidyl site bound codon and to the third nucleotide of the A site bound codon, in the absence/presence of second tRNA bound at the A site. The values of the obtained interspin distances agree with those calculated for available near-atomic structures of similar complexes of 40S and 80S ribosomes, showing that neither 60S subunit nor tRNA at the A site have a noticeable effect on arrangement of mRNA at the codon-anticodon interaction area. In addition, the shapes of distance distributions in four studied ribosomal complexes allowed conclusions on conformational flexibility of mRNA in these complexes. Overall, the results of this study are the first, to our knowledge, demonstration of double electron-electron resonance/pulsed electron-electron double resonance application for measurements of intramolecular distances in multicomponent supramolecular complexes involving intricate cellular machineries and for evaluating dynamic properties of ligands bound to these machineries.     

EphA2 Mutation in Lung Squamous Cell Carcinoma Promotes Increased Cell Survival,Cell Invasion,Focal Adhesions,and Mammalian Target of Rapamycin Activation

Non-small cell lung cancer (NSCLC) has a poor prognosis and improved therapies are needed. Expression of EphA2 is increased in NSCLC metastases. In this study, we investigated EphA2 mutations in NSCLC and examined molecular pathways involved in NSCLC. Tumor and cell line DNA was sequenced. One EphA2 mutation was modeled by expression in BEAS2B cells, and functional and biochemical studies were conducted. A G391R mutation was detected in H2170 and 2/28 squamous cell carcinoma patient samples. EphA2 G391R caused constitutive activation of EphA2 with increased phosphorylation of Src, cortactin, and p130^Cas. Wild-type (WT) and G391R cells had 20 and 40% increased invasiveness; this was attenuated with knockdown of Src, cortactin, or p130^Cas. WT and G391R cells demonstrated a 70% increase in focal adhesion area. Mammalian target of rapamycin (mTOR) phosphorylation was increased in G391R cells with increased survival (55%) compared with WT (30%) and had increased sensitivity to rapamycin. A recurrent EphA2 mutation is present in lung squamous cell carcinoma and increases tumor invasion and survival through activation of focal adhesions and actin cytoskeletal regulatory proteins as well as mTOR. Further study of EphA2 as a therapeutic target is warranted.     

Recent methods of drought stress tolerance in plants

In the climate change scenario the drought has been diagnosed as major stress affecting crop productivity. This review demonstrates some recent findings on the amelioration of drought stress. Nanoparticles, synthetic growth regulators viz. Trinexapac-ethyl, and Biochar addition helps to economize the water budget of plants, enhances the bioavailability of water and nutrients as well as overcomes drought induced osmotic and oxidative stresses. Besides ABA, SA and JA are also involved in inducing tolerance to drought stress through modulation of physiological and biochemical processes in plants. Plant growth promoting rhizobacteria (PGPR) offer new opportunities in agricultural biotechnology. These beneficial microorganisms colonize the rhizosphere/endo-rhizosphere of plants and impart drought tolerance by improving root architechture, enhancing water use efficiency, producing exopolysaccharides, phytohormones viz, ABA, SA and IAA and volatile compounds. Further PGPR also play positive role in combating osmotic and oxidative stresses induced by drought stress through enhancing the accumulation of osmolytes, antioxidants and upregulation or down regulation of stress responsive genes. In transgenic plants stress inducible genes enhanced abiotic stress tolerance by encoding key enzymes regulating biosynthesis of compatible solutes. The role of genes/cDNAs encoding proteins involved in regulating other genes/proteins, signal transduction process and strategies to improve drought stress tolerance have also been discussed.     

MicroRNA-21与平滑肌蛋白22α在哈萨克族食管癌组织中的表达及其临床病理学意义

目的探讨microRNA21与SM22a基因在哈萨克族食管癌发生发展中的作用及临床意义。方法免疫组织化学法检测162例石蜡包埋食管鳞状细胞癌组织及RT-PCR方法检测47例哈萨克族食管癌标本中microRNA21、SM22a表达水平,分析这些基因与临床病理特征的关系。结果SM22a在162例食管鳞癌组织中的阳性表达率(87.0%)显著高于食管正常黏膜组织(36.0%);在47例哈萨克族食管癌组织中,SM22a表达水平较远端无癌组织增高。与远端无癌组织相比,microRNA21在哈萨克族食管癌组织中表达水平增高。MicroRNA21高表达与分化程度、淋巴结转移、临床分期相关,SM22a高表达与临床分期相关、淋巴结转移相关;microRNA21与SM22a的表达呈正相关。结论MicroRNA21、SM22a协同高表达共同参与哈萨克族食管癌的侵袭发展过程。     

2′-OH of mRNA are critical for the binding of its codons at the 40S ribosomal P site but not at the mRNA entry site

The roles of 2′-OH groups in the binding of mRNA to human ribosomes were studied using site-directed cross-linking. We found that both mRNA and mDNA analogues bearing a cross-linker can modify ribosomal proteins (rps) S3e and S2e at the mRNA entry site independently on tRNA presence, but only mRNA analogues were capable of a tRNA^Phe-dependent binding to human ribosomes and cross-linking to rpS26e in the mRNA binding centre. Thus, 2′-OH groups of mRNA are unimportant for binding at the entry site but they are crucial for codon-anticodon interactions at the P site, implying the existence of mRNA-ribosome contacts that do not occur in bacteria.     

Does glutathione S-transferase Pi (GST-Pi) a marker protein for cancer?

Glutathione S-transferases (GSTs, EC 2.5.1.18) are multifunctional and multigene products. They are versatile enzymes and participate in the nucleophilic attack of the sulphur atom of glutathione on the electrophilic centers of various endogenous and xenobiotic compounds. Out of the five, , and , major classes of GSTs, GST- has significance in the diagnosis of cancers as it is expressed abundantly in tumor cells. This protein is a single gene product, coded by seven exons, that is having 24 kDa mass and pI value of 7.0. Four upstream elements such as two enhancers, and one of each of AP-1 site and GC box regulate gene. During chemical carcinogenesis because of jun/fos oncogenes (AP-1) regulatory elements, specifically GST- is expressed in liver. Therefore this gene product could be used as marker protein for the detection of chemical toxicity and carcinogenesis.     

甘肃、新疆、内蒙苹果蠹蛾成虫消长规律

苹果蠹蛾Cydia pomonella(L.)是我国重要的果树害虫和检疫对象。2005年至2010年,本研究在甘肃、新疆及内蒙古的不同区县选取了16个果园,使用性信息素诱捕器对其中的苹果蠹蛾成虫发生规律进行长期监测。结果表明,苹果蠹蛾在西北地区每年发生2.5个世代;在正常气候条件下,3个成虫发生高峰分别出现在5月上旬、7月中下旬和8月中下旬,但不同地区及同一地区不同果园之间存在较大差异;化学防治、迷向防治等防治措施对苹果蠹蛾成虫捕获量的影响较大,因此生活史研究为主的监测并不适合在上述果园中开展。综合上述研究结果,未对苹果蠹蛾的季节动态进行准确的预测,需要对苹果蠹蛾除成虫外的其他虫态的季节性变化进行详细研究,并建议选择3个以上的果园进行监测,综合各个果园的监测结果并得出结论。     

Lymphatic Endothelial Differentiation in Pulmonary Lymphangioleiomyomatosis Cells

Pulmonary lymphangioleiomyomatosis (LAM) is a rare, low-grade neoplasm affecting almost exclusively women of childbearing age. LAM belongs to the family of perivascular epithelioid cell tumors, characterized by spindle and epithelioid cells with smooth muscle and melanocytic differentiation. LAM cells infiltrate the lungs, producing multiple, bilateral lesions rich in lymphatic channels and forming cysts, leading to respiratory insufficiency. Here we used antibodies against four lymphatic endothelial markers—podoplanin (detected by D2-40), prospero homeobox 1 (PROX1), vascular endothelial growth factor receptor 3 (VEGFR-3), and lymphatic vessel endothelial hyaluronan receptor 1 (LYVE1)—to determine whether LAM cells show lymphatic differentiation. Twelve of 12 diagnostic biopsy specimens (early-stage LAM) and 19 of 19 explants (late-stage LAM) showed immunopositivity for D2-40 in most neoplastic cells. PROX1, VEGFR-3, and LYVE1 immunoreactivity varied from scarce in the early stage to abundant in the late stage. Lymphatic endothelial, smooth muscle, and melanocytic markers were partially co-localized. These findings indicate that lymphatic endothelial differentiation is a feature of LAM and provide evidence of a previously unidentified third lineage of differentiation in this neoplasm. This study has implications for the histological diagnosis of LAM, the origin of the neoplastic cells, and potential future treatment with drugs targeting lymphangiogenesis.