ACTIVITY-DEPENDENT POLYPEPTIDE ELONGATION IN MUSCLE

这一项目的近期目标是确定新生人口的减少 多肽伸长率导致蛋白质合成迅速减少 在非负重的骨骼肌中观察到。对…的明确识别 肌肉萎缩最初事件的作用部位将有助于 定义维持正常基因表达的细胞内信号。 这些信号可能影响骨骼肌对慢性疲劳的适应。 卧床休息、截瘫、四肢瘫痪和失重。 如果阻止大鼠比目鱼肌失去80%的肌原纤维蛋白 举重。尽管所有水平的基因表达都会受到影响，但 大多数快速变化发生在翻译层面；在五个小时内 起病的非负重肌原纤维蛋白合成率有 在朝着最终下降50%的方向发展的过程中，至少减少了25%。 初步数据表明，最初的缺陷可能是降低的比率 指新生的多肽合成，表现为多聚体增多 尺码。这一提议的工作假设是，放慢的速度 伸长导致80S核糖体在mRNA上积累。这个 实验还将确定这种形式的监管是否专门针对 收缩蛋白。两个实验将测试更多的80s核糖体 MRNA.第一个将测量放射性氨基酸的掺入量。 进入新生的多肽池。修正了氨基酰基-tRNA特异性 活性，此值与新生多肽的数量成正比 (因此，80S核糖体的数量)与该mRNA相关。这个 第二个实验将直接测量核糖体亚基的数量 与α-肌动蛋白和细胞色素c mRNAs相关。以测试是否 调节是针对收缩蛋白、非肌肉基因(β- 半乳糖苷酶和酒精脱氢酶)将被结构性表达 通过逆转录病毒介导的基因转移在萎缩的肌肉中。信使核糖核酸 因为这些基因不会反映多聚体相对于阿尔法的大小移动- 肌动蛋白，如果调节是收缩蛋白所特有的。

The immediate goal of this project is to determine if a decrease in nascent polypeptide elongation rate causes the rapid decrease in protein synthesis observed in non-weightbearing skeletal muscle. A clear identification of the site-of-action of the initial events in muscle atrophy will help to define the intracellular signals that maintain normal gene expression. These signals may influence the adaptation of skeletal muscle to chronic bedrest, paraplegia, limb immobilization, and weightlessness. Rat soleus muscle loses 80% of its myofibril protein if prevented from weightbearing. Although all levels of gene expression are affected, the most rapid changes occur at the level of translation; within five hours of the onset of non-weightbearing myofibril protein synthesis rate has diminished by at least 25% as it evolves toward an eventual 50% decline. Preliminary data indicate that the initial defect may be a decreased rate of nascent polypeptide synthesis that is manifest as in increased polysome size. The working hypothesis of this proposal is that a slowed rate of elongation causes an accumulation of 80S ribosomes on the mRNA. The experiments will also determine if this form of regulation is specific for contractile proteins. Two experiments will test for more 80S ribosomes per mRNA. The first will measure amount of radioactive amino acid incorporated into the nascent polypeptide pool. Corrected for aminoacyl-tRNA specific activity, this value is proportional to the number of nascent polypeptides (and hence the number of 80S ribosomes) associated with the mRNA. The second experiment will directly measure the number of ribosomal subunits associated with both alpha-actin and cytochrome c mRNAs. To test if the regulation is specific for contractile proteins, non-muscle genes (beta- galactosidase and alcohol dehydrogenase) will be constitutively expressed in the atrophying muscle by retroviral-mediated gene transfer. The mRNAs for these genes will not reflect the polysome size shift relative to alpha- actin if regulation is specific for contractile proteins.