Phylogenetic Analysis of Zebrafish Basic Helix-Loop-Helix Transcription Factors

The basic helix-loop-helix (bHLH) proteins play important regulatory roles in eukaryotic developmental processes including neurogenesis, myogenesis, hematopoiesis, sex determination, and gut development. Zebrafish is a good model organism for developmental biology. In this study, we identified 139 bHLH genes encoded in the zebrafish genome. Phylogenetic analyses revealed that zebrafish has 58, 29, 21, 5, 19, and 5 bHLH members in groups A, B, C, D, E, and F, respectively, while 2 members were classified as “orphan.” A comparison between zebrafish and human bHLH repertoires suggested that both organisms have a certain number of specific bHLH members. Eight zebrafish bHLH genes were found to have multiple coding regions in the genome. Two of these, Bmal1 and MITF, are good anchor genes for identification of fish-specific whole-genome duplication events in comparison with mouse and chicken genomes. The present study provides useful information for future studies on gene family evolution and vertebrate development. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

重要转录因子FoxO3a在斑马鱼性腺中的表达分析

FoxO蛋白作为Forkhead Box家族的重要一员,在动物的生长发育、细胞分化、代谢、免疫和凋亡等方面起重要作用,研究发现FoxO蛋白是哺乳动物卵泡发育的重要调节因子.通过检测雌雄斑马鱼性腺中foxo3a基因的表达情况,初步研究FoxO在鱼类生殖发育中的作用.PCR和蛋白免疫杂交结果都显示foxo3a基因在斑马鱼雌性性腺中的表达量要明显高于雄性.这一结果提示,foxo3a基因在斑马鱼的卵巢发育中可能发挥重要作用.     

Induction of clusterin Expression by Neuronal Cell Death in Zebrafish

Clusterin, a protein associated with multiple functions, is expressed in a wide variety of mammalian tissues. Although clusterin is known to be involved in neurodegenerative diseases, ageing, and tumorigenesis, a detailed analysis of the consequences of gain- or loss-of-function approaches has yet to be performed to understand the underlying mechanisms of clusterin functions. Since clusterin levels change in neurological diseases, it is likely that clusterin contributes to cell death and degeneration in general. Zebrafish was investigated as a model system to study human diseases. During development, zebrafish clusterin was expressed in the notochord and nervous system. Embryonic overexpression of clusterin by mRNA microinjection did not affect axis formation, whereas its knock-down by anti-sense morpholino treatment resulted in neuronal cell death. To analyze the function of clusterin in neurodegeneration, a transgenic zebrafish was investigated, in which nitroreductase expression is regulated under the control of a neuron-specific huC promoter which is active between the stages of early neuronal precursors and mature neurons. Nitroreductase turns metronidazole into a cytotoxic agent that induces cell death within 12 h. After metronidazole treatment, transgenic zebrafish showed neuron-specific cell death. Interestingly, we also observed a dramatic induction of clusterin expression in the brain and spinal cord in these fish, suggesting a direct or indirect role of clusterin in neuronal cell death and thus, more generally, in neurodegeneration.     

植物非生物胁迫诱导启动子顺式元件及转录因子研究进展

顺式作用元件(cix-acting element)是与结构基因串联的特定DNA序列,是转录因子的结合位点,它们通过与转录因子结合调控基因转录的精确起始和转录效率,在植物基因表达调控过程中起着重要的作用.非生物胁迫诱导基因的表达受其上游启动子顺式作用元件及转录因子的调控,目前已发现了多种与非生物胁迫相关的顺势作用元件及转录因子,如DRE元件及DREB类转录因子、MYB元件及MYB类转录因子、GT-1元件及GT-1类转录因子等.顺式作用元件及转录因子的研究对研究植物非生物胁迫相关基因的表达调控具有重要意义,综述植物非生物胁迫诱导启动子功能元件及转录因子的研究进展.     

低氧诱导因子和角质细胞生长因子双基因重组腺病毒的构建及其在肺泡上皮细胞中的表达

目的：构建同时携带低氧诱导因子-1α（HIF-1α）和角质细胞生长因子（KGF）N腺病毒载体（pAdxsi-GFP-HIF-KGF）,观察其在防治肺损伤潜在的应用前景。方法：低氧处理A549细胞后提取总RNA并逆转录为eDNA作为模板,依据GeneBank公布的HIF-1α cDNA设计引物,并分别引入KpnI和BamHI酶切位点,PCR扩增后将目的基因HIF-1α连接到载体pShuttle-CMV-EGFP上,构建重组质粒pShuttle-GFP—HIF。然后以质粒plRES2-EGFP-KGF为模板,用引入NheI和PmeI酶切位点的引物PCR扩增KGF基因并克隆到重组质粒pShuttle-GFP-HIF上,获得穿梭质粒重组质粒pShuttle—GFP-HIF—KGF。采用细菌内重组方法将目的序列重组到pAdxsi病毒骨架栽体上构建携带HIF．10t和KGF双基因的重组腺病毒载体pAdxsi-GFP-HIF-KGF。检测重组腺病毒滴度后,转染人肺泡上皮细胞A549,检测目的基因的转染表达。结果：通过对构建质粒克隆进行测序及酶切,证实携带HIF—lot和KGF双基因的重组腺病毒载体pAdxsi-GFP-HIF-KGF构建成功,且构建的重组腺病毒纯度好、滴度高。用pAdxsi-GFP-HIF-KGF以100MOI转染A549细胞后24h后在荧光显微镜下可观察到细胞有较强的绿色荧光表达,48h时荧光更强;转染48hELISA法检测培养上清中HIF-1蛋白表达水平为（56．36±4．53）ng／mL,KGF蛋白表达水平为（60．20±2．92）ng／mL。结论：成功构建了腺病毒栽体pAdxsi-GFP-HIF-KGF,其转染效率及目的基因的蛋白表达水平较高,具有潜在的进一步在肺损伤局部应用的前景,为后期制备可以同时发挥KGF、HIF-1作用的基因治疗药物打下基础,同时为高海拔地区应激性急性肺损伤的有效防治提供实验基础。     

低氧诱导因子在肾炎大鼠肾小管间质中的表达及氯沙坦的保护作用研究

目的：观察低氧诱导因子在肾炎大鼠肾小管间质中表达的变化以及氯沙坦对肾炎的保护作用。方法：雄性Wistar 大鼠32只,随机分成假手术组、肾炎模型组、氯沙坦小剂量组、氯沙坦大剂量组各8 只,假手术组不做肾脏切除,其他组在右肾切除后尾静脉注射标记抗体。给药组则按量分别灌胃给药,8 周时处死。结果：大剂量给药组的血肌酐（Scr）、收缩压和24 h尿蛋白量较模型组显著降低,且小剂量给药组的血肌酐（Scr）和24h 尿蛋白量较模型组也显著降低。小剂量组和大剂量组大鼠的肾间质面积较肾炎模型组均显著降低。HIF-1αmRNA 大量表达于肾小管上皮细胞胞质和间质细胞胞质,且与模型组比较,小剂量给药组的HIF-1αmRNA显著降低;大剂量给药组较肾炎模型组HIF-1琢mRNA 的含量也显著降低。结论：氯沙坦可能可以通过影响低氧诱导因子（HIF-1α）的表达发挥对肾脏的保护作用。     

A Novel Role for the Hypoxia Inducible Transcription Factor HIF-1alpha: Critical Regulation of Inflammatory Cell Function

No abstract available.     

斑马鱼NUP98基因的克隆及其时空表达图谱

目的：斑马鱼NUP98基因的克隆及其在个体早期发育过程中的表达情况研究。方法：提取斑马鱼胚胎的总RNA,制备地高辛标记的NUP98RNA反义探针,WISH（整体胚胎原位杂交）研究NUP98在斑马鱼早期发育过程中的表达;提取斑马鱼胚胎各时相和成鱼各组织的RNA,实时定量PCR检测斑马鱼胚胎各时相和成鱼各组织中的表达。结果：成功克隆斑马鱼NUP98基因,通过实时定量RT-PCR和原位杂交,获得NUP98基因在斑马鱼早期发育过程中的表达情况：NUP98在2-cell、32．cell、oblong、shield期、12h前普遍性表达（0．75h、1．7h、3．7h、6h、12h）;24h以后在眼部、头部表达较多,特别是在脊索表达较高;斑马鱼NUP98在0、0．5h、6h、12h、24h、48h表达逐渐降低,到72h和96h表达有所增加,但是仍低于24h其表达水平;NUP98在成鱼眼、脑、鳔、肾、肝、睾丸、胆囊、卵巢、鳍、心、肠、肌肉、腮、皮肤的表达中,眼的表达最高,明显高于其他组织,腮、卵巢、肠的表达次之,肌肉、鳔、胆囊、睾丸、皮肤、脑的表达紧随其后,鳍、肝、心、肾的表达最低。结论：NUP98基因可能在个体脑部、脊索及眼部的早期发育过程中起到了重要作用;NUP98基因可能具有抑制肿瘤发生的作用,该基因的调节异常对白血病的发生发展可能有重要影响。这些研究结果为进一步研究NUP98基因在造血系统中的作用,评估其是否适合作为血液系统恶性肿瘤的新的治疗靶点等奠定了理论基础。     

缺氧诱导因子-1的调控及其转录后修饰(英文)

缺氧诱导因子-1(hypoxia inducible factor-1,HIF-1)是一种异源二聚体转录因子,由结构表达型β亚基和氧调节型α亚基组成。在低氧环境下,HIF-1调控一系列促进细胞成活的基因,这些基因涉及血管生成、铁代谢、葡萄糖代谢和细胞增殖与存活。α亚基主要受到诸如乙酰化、羟基化、磷酸化和相扑化等转录后修饰,这些修饰可以稳定或激活HIF-1的活性。除氧环境外,胞内氧化还原稳态、铁代谢、线粒体代谢物和生长因子还可通过影响转录后修饰进而调节HIF-1的活性。此外,近来的研究表明HIF-1在病原学方面也发挥重要作用,在中风和神经退行性疾病这样的脑紊乱疾病中提供潜在神经保护作用。本文总结了HIF-1研究的最新进展,谨以此文献给忻文娟教授80周年诞辰。