TRANSLATIONAL REGULATION DURING RED CELL DIFFERENTIATION

本实验室的研究旨在了解细胞 Friend小鼠诱导分化的机制 红白血病（MEL）细胞。 最近的研究结果表明：1） 这些成红细胞的转化直接影响翻译 启动;和2）暴露于诱导分化迅速 降低了翻译速率，这种降低先于分化。 这一点和其他大量证据表明，翻译是一个重要的 转化细胞中基因表达的调节因子。 实验 本提案中概述的内容将决定翻译的作用 MEL转化和分化过程中的基因表达调控 细胞 诱导剂被证明影响一个以前未描述的网站， 翻译调节，其中mRNA以单体形式积累， 慢慢地转化为多聚核糖体。 单体的积累 似乎是由于在mRNA的起始AUG密码子处“停滞”。 的 本提案中概述的实验将确定 翻译受诱导物的影响，受“足迹”的影响， 核糖体和mRNA，并分析存在于核糖体中的tRNA和肽。 停滞的单体 调节失速的因素将通过 开发用于该事件的体外测定，并通过纯化和进一步 两种核糖体蛋白的特征， 从这些停滞的单体中。 翻译启动率将为 选择性增加或减少MEL细胞利用最近的 描述了遗传学方法。 这将决定翻译的作用 在转化这些细胞及其诱导的 分化 这些实验的结果将提供洞察力 转化为真核生物生长调节和翻译）调节机制。

Research in this laboratory is directed at understanding the cellular mechanisms responsible for induced differentiation of Friend murine erythroleukemia (MEL) cells. Recent results demonstrate that: 1) transformation of these erythroblasts directly affects translation initiation; and 2) exposure to inducers of differentiation rapidly decreases translation rate and this decrease precedes differentiation. This and other abundant evidence indicate that translation is a significant regulator of gene expression in transformed cells. The experiments outlined in this proposal will determine the role of translational regulation of gene expression in transformation and differentiation of MEL cells. Inducers are shown to affect a previously undescribed site for translational regulation where MRNA accumulates as monosomes and only slowly progresses to translating polysomes. The accumulation of monosomes appears to be due to "stalling" at the initiating AUG codon of MRNA. The experiments outlined in this proposal will determine the precise step in translation affected by inducers by "footprinting" the interaction of ribosomes and MRNA and analyzing the tRNAs and peptides present in the stalled monosome. Factors which regulate stalling will be analyzed by developing an in vitro assay for this event and by purification and further characterization of two ribosomal proteins which are specifically absent from these stalled monosomes. Translation initiation rate will be selectively increased or decreased in MEL cells utilizing a recently described genetic approach. This will determine the role of translation initiation in transforming these cells and in their induced differentiation. The results of these experiments will provide insight into eukaryotic growth regulation and translations) regulatory mechanisms.