TRANSLATIONAL CONTROL OF GENE EXPRESSION BY MRNA

mRNA 对基因表达的翻译控制

• 批准号: 3283543
• 负责人: ROSEMARY JAGUS
• 金额: $ 11.74万
• 依托单位: UNIVERSITY OF PITTSBURGH AT PITTSBURGH
• 依托单位国家: 美国
• 财政年份: 1983
• 资助国家: 美国
• 起止时间: 1983-12-01 至 1992-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3283543
• 关键词: Nematoda; adenosine triphosphate; antibody; binding proteins; calcium; cell free system; chemical binding; chemical structure function; crosslink; density gradient ultracentrifugation; developmental genetics; early embryonic stage; fertilization; gel electrophoresis; gene expression; genetic manipulation; genetic transcription; genetic translation; germ cells; histones; hydrolysis; immunochemistry; messenger RNA; nonmammalian vertebrate embryology; nuclease; nucleic acid probes; protein biosynthesis; protein kinase; protein structure; radiotracer; ribonucleoproteins; ribosomes; saltwater environment; sea urchins; synthetic nucleic acid

During oogenesis and maturation of the sea urchin egg, large stores of mRNA are accumulated for subsequent use during early embryogenesis. Utilization of stored maternal mRNA occurs at a very low rate prior to fertilization. After fertilization, dramatic changes in mRNA availability and recruitment occur, allowing rapid increases in the synthetic rate of many proteins during early cleavage. Changes have been documented in both overall rates of maternal mRNA utilization and in the selection of particular mRNA species at different developmental stages. Our previous studies have allowed the development of cell-free translation systems from eggs and embryos that reproduce the overall differences observed in the intact system. This investigation proposes to utilize these cell-free systems, in conjunction with other approaches, to identify the cellular components involved in the utilization of maternal mRNA. Using the cell-free systems an inhibitor of eIF-4F activity has been discovered in the unfertilized egg, that decreases gradually in activity over the first 2-3 hr after fertilization. The identity and nature of the eIF-4F inhibitory activity will be characterized. The mechanism by which this activity inhibits the initiation sequence will also be investigated. The manner in which the inhibitor is inactivated after fertilization will be explored. To complement these studies, a systematic search for developmentally regulated mRNA-interactive proteins will be undertaken, using a protein blotting and in vitro binding technique, with capped 32p-labelled mRNA's synthesized in vitro.

在海胆卵的卵子发生和成熟过程中，大 mRNA 的储存被积累以供早期使用 胚胎发生。 储存的母体 mRNA 的利用发生在 受精前的比率非常低。 受精后，戏剧性的 mRNA 可用性和募集发生变化，从而允许 早期许多蛋白质的合成率迅速增加 乳沟。 总体比率的变化已记录在案 母体 mRNA 的利用和特定的选择 处于不同发育阶段的mRNA种类。 我们之前的 研究已经允许无细胞翻译的发展 由卵子和胚胎组成的系统，可再现整体 在完整系统中观察到的差异。 这项研究建议利用这些无细胞系统 与其他方法结合，识别细胞 参与母体 mRNA 利用的成分。 使用 在无细胞系统中，eIF-4F 活性的抑制剂已被 在未受精卵中发现，逐渐减少 受精后2-3小时内的活动。 身份和 eIF-4F 抑制活性的性质将被表征。 该活性抑制启动的机制 序列也将被调查。 方式 将探讨受精后抑制剂失活的情况。 到 补充这些研究，系统地寻找 发育调节的 mRNA 相互作用蛋白将 使用蛋白质印迹和体外结合进行 技术，在体外合成加帽 32p 标记的 mRNA。