STRUCTURE AND FUNCTION OF NUCLEAR RNP PARTICLES

在真核细胞中，信使RNA（mRNA），功能性可翻译的 基因表达的中间产物，是通过大量的 初级基因转录物（hnRNA）。 我们和其他人最近 证明hnRNA存在于细胞核中，与一种 hnRNP颗粒是由一组特定的蛋白质形成的独特结构，称为hnRNP颗粒。 hnRNP颗粒是细胞核中最丰富的结构之一， 是RNA加工的场所。 RNP蛋白是包装、加工和功能所必需的 这些多核苷酸。 RNA相关蛋白可以是 光化学交联到它在完整的细胞，和交联的 可以容易地分离复合物。 我们用交联的 hnRNP复合物，并产生了针对几种 不同真核生物中的主要hnRNP蛋白。 有了这些抗体， 已经开始表征hnRNP蛋白，并分离出hnRNP 复杂. 其中一种针对主要hnRNP C蛋白（41 K和 43 K）在体外特异性抑制mRNA前体的剪接。 我们将研究hnRNP蛋白的结构和功能， hnRNP颗粒，以了解它们在mRNA形成中的作用 以及hnRNA在细胞核中的包装和组织。 的 将使用抗体分离hnRNP蛋白的cDNA克隆 通过表达载体文库的免疫学筛选。 的氨基酸 蛋白质的序列将由cDNA核苷酸确定 序列和它们的mRNA及其基因将被表征。 额外 将获得并表征针对hnRNP蛋白的抗体。 的 hnRNP蛋白与体外合成的特异性RNA的相互作用将 为了理解hnRNP的组装和拆卸， 复杂的-知识，这是必要的了解结构和 hnRNP颗粒的形成和核质融合过程 mRNA的运输。 蛋白质和hnRNP复合物的功能， 转录和RNA剪接将在体外系统中进行研究。 的 分离纯的和完整的hnRNP颗粒的能力以及 针对单个hnRNP蛋白的抗体将使进一步的 它们的结构和它们在mRNA生物发生中的作用的表征。

In eukaryotic cells, messenger RNAs (mRNAs), the functional translatable intermediates of gene expression, are formed by extensive processing of primary gene transcripts (hnRNAs). We and others have recently demonstrated that hnRNAs exist in the nucleus in association with a specific set of proteins to form unique structures termed hnRNP particles. The hnRNP particles, one of the most abundant structures in the nucleus, are the sites of RNA processing. The RNP proteins are essential for the packaging, processing and function of these polynucleotides. The RNA-associated proteins can be photochemically crosslinked to it in intact cells, and the crosslinked complexes can be readily isolated. We have immunized mice with crosslinked hnRNP complexes and have generated monoclonal antibodies against several of the major hnRNP proteins in divergent eukaryotes. With these antibodies we have begun to characterize the hnRNP proteins and have isolated the hnRNP complex. One of the antibodies to the prominent hnRNP C proteins (41K and 43K) specifically inhibits the splicing of mRNA precursors in vitro. We shall investigate the structure and function of the hnRNP proteins and the hnRNP particles in order to understand their role in mRNA formation and in the packaging and organization of hnRNA in the nucleus. The antibodies will be used to isolate the cDNA clones for the hnRNP proteins by immunological screening of expression vector libraries. The amino acid sequence of the proteins will be determined form the cDNA nucleotide sequence and their mRNAs and their genes will be characterized. Additional antibodies to hnRNP proteins will be obtained and characterized. The interaction of hnRNP proteins with specific RNAs synthesized in vitro will be studied in order to understand the assembly and disassembly of the hnRNP complex -- knowledge which is necessary for understanding the structure and formation of hnRNP particles and the process of nucleo-cytoplasmic transport of mRNA. The function of the proteins and the hnRNP complexes in transcription and in RNA splicing will be studied in in vitro systems. The ability to isolate pure and intact hnRNP particles and the availability of antibodies to individual hnRNP proteins will enable the further characterization of their structure and their role in mRNA biogenesis.