Proliferating cells express mRNAs with shortened 3' untranslated regions and fewer microRNA target sites

Messenger RNA (mRNA) stability, localization, and translation are largely determined by sequences in the 3' untranslated region (3'UTR). We found a conserved increase in expression of mRNAs terminating at upstream polyadenylation sites after activation of primary murine CD4+ T lymphocytes. This program, resulting in shorter 3'UTRs, is a characteristic of gene expression during immune cell activation and correlates with proliferation across diverse cell types and tissues. Forced expression of full-length 3'UTRs conferred reduced protein expression. In some cases the reduction in protein expression could be reversed by deletion of predicted microRNA target sites in the variably included region. Our data indicate that gene expression is coordinately regulated, such that states of increased proliferation are associated with widespread reductions in the 3'UTR-based regulatory capacity of mRNAs.     

Modulation of hepatitis C virus RNA abundance by a liver-specific MicroRNA

MicroRNAs are small RNA molecules that regulate messenger RNA (mRNA) expression. MicroRNA 122 (miR-122) is specifically expressed and highly abundant in the human liver. We show that the sequestration of miR-122 in liver cells results in marked loss of autonomously replicating hepatitis C viral RNAs. A genetic interaction between miR-122 and the 5' noncoding region of the viral genome was revealed by mutational analyses of the predicted microRNA binding site and ectopic expression of miR-122 molecules containing compensatory mutations. Studies with replication-defective RNAs suggested that miR-122 did not detectably affect mRNA translation or RNA stability. Therefore, miR-122 is likely to facilitate replication of the viral RNA, suggesting that miR-122 may present a target for antiviral intervention.     

S6K1- and betaTRCP-mediated degradation of PDCD4 promotes protein translation and cell growth

The tumor suppressor programmed cell death protein 4 (PDCD4) inhibits the translation initiation factor eIF4A, an RNA helicase that catalyzes the unwinding of secondary structure at the 5' untranslated region (5'UTR) of messenger RNAs (mRNAs). In response to mitogens, PDCD4 was rapidly phosphorylated on Ser67 by the protein kinase S6K1 and subsequently degraded via the ubiquitin ligase SCF(betaTRCP). Expression in cultured cells of a stable PDCD4 mutant that is unable to bind betaTRCP inhibited translation of an mRNA with a structured 5'UTR, resulted in smaller cell size, and slowed down cell cycle progression. We propose that regulated degradation of PDCD4 in response to mitogens allows efficient protein synthesis and consequently cell growth.     

microRNA靶基因检测技术研究进展

microRNA是真核生物基因表达调控的重要因子,对其靶基因的验证是研究microRNA功能必不可少的环节。验证microRNA与其靶基因的相互作用,可以根据能否进行靶基因切割、能否造成目标靶基因RNA表达量或蛋白表达量降低以及能否形成RISC-mRNA复合体等特征进行检测。基于此,从以上三方面对验证microRNA靶基因的技术进行了综述。     

循环microRNA在冠心病及其中医证候诊断中的作用

microRNA是一类长度约为22个核苷酸的内源性非编码RNA,并能以特定的方式与靶基因结合,从而抑制靶基因的翻译或使其降解。microRNA只有在受到致病因素（如环境因素）的影响以及病理状态下,其表达量才会发生改变,因此microRNA具有成为各种疾病的诊断生物标志物的可能。目前有大量研究表明microRNA参与冠心病发生发展,但其作用机制尚未完全阐明,本文从microRNA的特性、microRNA与冠心病、microRNA与中医证候等方面对近年来国内外的研究做一综述。     

Dom34:Hbs1 promotes subunit dissociation and peptidyl-tRNA drop-off to initiate no-go decay

No-go decay (NGD) is one of several messenger RNA (mRNA) surveillance systems dedicated to the removal of defective mRNAs from the available pool. Two interacting factors, Dom34 and Hbs1, are genetically implicated in NGD in yeast. Using a reconstituted yeast translation system, we show that Dom34:Hbs1 interacts with the ribosome to promote subunit dissociation and peptidyl-tRNA drop-off. Our data further indicate that these recycling activities are shared by the homologous translation termination factor complex eRF1:eRF3, suggesting a common ancestral function. Because Dom34:Hbs1 activity exhibits no dependence on either peptide length or A-site codon identity, we propose that this quality-control system functions broadly to recycle ribosomes throughout the translation cycle whenever stalls occur.     

Control of the reversibility of cellular quiescence by the transcriptional repressor HES1

The mechanisms by which quiescent cells, including adult stem cells, preserve their ability to resume proliferation after weeks or even years of cell cycle arrest are not known. We report that reversibility is not a passive property of nondividing cells, because enforced cell cycle arrest for a period as brief as 4 days initiates spontaneous, premature, and irreversible senescence. Increased expression of the gene encoding the basic helix-loop-helix protein HES1 was required for quiescence to be reversible, because HES1 prevented both premature senescence and inappropriate differentiation in quiescent fibroblasts. In some human tumors, the HES1 pathway was activated, which allowed these cells to evade differentiation and irreversible cell cycle arrest. We conclude that HES1 safeguards against irreversible cell cycle exit both during normal cellular quiescence and pathologically in the setting of tumorigenesis.     

miRNA-mediated gene silencing by translational repression followed by mRNA deadenylation and decay

microRNAs (miRNAs) regulate gene expression through translational repression and/or messenger RNA (mRNA) deadenylation and decay. Because translation, deadenylation, and decay are closely linked processes, it is important to establish their ordering and thus to define the molecular mechanism of silencing. We have investigated the kinetics of these events in miRNA-mediated gene silencing by using a Drosophila S2 cell-based controllable expression system and show that mRNAs with both natural and engineered 3' untranslated regions with miRNA target sites are first subject to translational inhibition, followed by effects on deadenylation and decay. We next used a natural translational elongation stall to show that miRNA-mediated silencing inhibits translation at an early step, potentially translation initiation.     

拟南芥miRNA及其靶序列配对特征分析

microRNAs(miRNAs)是真核生物中一类长度约为22 nt的非编码小分子RNA,miRNA与AGO等蛋白形成RISC沉默复合体,通过剪切或翻译抑制对靶基因起负调控作用。对拟南芥miRNA序列及其配对的靶序列间的特征进行了统计分析,结果表明miRNA序列5′端富含A、U,第1、第19碱基位对U、C具有较强的倾向性;miRNA与靶序列间常有1~4个碱基错配,错配碱基常出现在第1,第2和第21位,而第3~第6,第9~第10,第16~第17碱基配对较为保守,为人工合成miRNA的设计及miRNA靶基因的预测以提供了依据。     

血清microRNA在癌症诊断中的进展

microRNA是长约19到25单位的内生性的微小RNA,它通过与靶信使RNA的3’UTR的特异性结合从而调节靶基因的表达。大量的试验证明了其在肿瘤的发生发展中microRNA发挥了重大的作用。近年来,在血清中发现了稳定存在的microRNA,而对microRNA的研究主要聚焦在血清microRNA与疾病的关系上。通过对患者和正常血清microRNA的检测和对比后,发现了一批可以用于肿瘤临床检测的microRNA。作为新的检测生物标志物,血清microRNA具有方便、快捷以及较高的准确性等优点,甚至可以区分在临床很难诊断的早期肿瘤,诸如非小细胞肺癌、胃癌以及胰腺癌。     

Pokemon敲减对结肠癌Lovo细胞周期、迁移和侵袭能力的影响

目的观察Pokemon敲减对结肠癌Lovo细胞周期、迁移和侵袭能力的影响。方法转染shRNA质粒构建Pokemon敲减的稳定细胞株Lovo-KD和阴性对照细胞株Lovo-NC,用荧光定量PCR检测Pokemon基因敲减效率,流式细胞术检测细胞周期变化,Transwell实验观察细胞迁移和侵袭能力。结果与Lovo-NC细胞比较,Lovo-KD细胞Pokemon mRNA水平下降65.0%（P〈0.01）,G1期细胞比例升高[（56.7±0.8）%vs（26.7±1.6）%,P〈0.01],迁移细胞数[（8±4）vs（189±19）个/视野,P〈0.01]和侵袭细胞数[（6±3）vs（183±13）个/视野,P〈0.01]减少。结论 Pokemon基因敲减可将Lovo细胞阻滞于G1期,使细胞迁移和侵袭能力显著下降。     

番茄microRNA调控生长发育及逆境响应的研究进展

[目的]回顾与总结番茄MicroRNA(miRNA)调控其生长发育及逆境响应的研究现状及进展,为番茄育种的应用提供理论和科学依据.[方法]查阅国内外相关文献,汇总并对比分析文献数据.[结果]miRNA是一类广泛存在于植物体内,位于基因组非编码区长约21～25个核苷酸的内源性非编码小分子RNA.其通过定向降解靶基因mRN...     

奶牛OLR1 3′UTR A223C多态的生物学信息分析

[目的]探索氧化型低密度脂蛋白受体OLR1 3＇UTR A223C多态与奶牛乳脂率高低紧密关联的分子机制。[方法]用miRanda（1.0）软件装载牛microRNA数据库,预测OLR1 3＇UTR潜在的microRNA。[结果]共预测16个microRNA靶标,OLR1 3＇UTR A223C突变定位于bta-miR-370靶标种子序列的互补区。[结论]由于OLR1 3＇UTR A→C突变导致bta-miR-370靶标的消失,为OLR1 3＇UTR A223C多态与奶牛乳脂率高低紧密关联的分子机制深入研究提供依据。     

Protein tyrosine kinase Wee1B is essential for metaphase II exit in mouse oocytes

Waves of cyclin synthesis and degradation regulate the activity of Cdc2 protein kinase during the cell cycle. Cdc2 inactivation by Wee1B-mediated phosphorylation is necessary for arrest of the oocyte at G2-prophase, but it is unclear whether this regulation functions later during the metaphase-to-anaphase transition. We show that reactivation of a Wee1B pathway triggers the decrease in Cdc2 activity during egg activation. When Wee1B is down-regulated, oocytes fail to form a pronucleus in response to Ca(2+) signals. Calcium-calmodulin-dependent kinase II (CaMKII) activates Wee1B, and CaMKII-driven exit from metaphase II is inhibited by Wee1B down-regulation, demonstrating that exit from metaphase requires not only a proteolytic degradation of cyclin B but also the inhibitory phosphorylation of Cdc2 by Wee1B.     

干扰KISS1基因对猪卵巢颗粒细胞功能的影响

【背景】卵泡是卵巢的基本结构和功能单位,其主要功能是排卵和分泌激素。颗粒细胞能促进卵泡发育,其过度凋亡能抑制卵泡发育,诱导卵泡闭锁,进而降低雌性动物发情频率,影响雌性动物繁殖力。现已有研究发现,KISS1在卵巢组织中发挥着重要作用。【目的】研究通过干扰KISS1,以阐释KISS1对猪卵巢颗粒细胞凋亡、周期及分泌雌激素能力的影响,为完善KISS1在猪颗粒细胞中的分子调控机制提供一定的依据。【方法】设计KISS1的干扰片段KISS1-siRNA,转染体外培养的母猪卵巢颗粒细胞,通过实时定量PCR(quantitative real time PCR, qRT-PCR)检测干扰KISS1对母猪卵巢颗粒细胞中磷脂酰肌醇3-激酶(phosphoinositide3-kinase, PI3K)信号通路部分基因转录水平的影响;采用流式检测法、Annexin V-FITC及ELISA技术,分别探究干扰KISS1对颗粒细胞周期、凋亡及雌二醇(estradiol, E2)分泌量的影响,最后使用qRT-PCR技术检测KISS1对雌激素受体及雌激素信号通路关键基因转录水平的影响。【结果】在猪颗粒细胞内,干扰K...