Trans-Splicing in Drosophila

描述(由申请人提供)： 摘要：前信使RNA剪接是去除内含子将外显子连接在一起的过程。这一过程用于合成几乎所有的后生动物mRNAs(约占人类mRNAs的93%)，是调节基因表达的重要手段。除了少数例外，剪接发生在顺式结构中--连接在一起的外显子位于相同的前-mRNA上。在一些真核生物中，包括线虫、锥虫和平面动物，反式剪接可以发生。在这些情况下，一个专门的剪接前导RNA被剪接到蛋白质编码RNA的5‘端。有趣的是，在少数情况下，发现了一种不同类型的反式剪接，即蛋白质编码基因外显子的剪接。两个最典型的例子是果蝇的mod(MDG4)和loA基因。在这些基因中，一组共同的5‘外显子可以反式拼接到几个可变的3’外显子中的一个(mod(MDG4)为26个，LOLA为22个)。一个令人信服的反式剪接案例已经被证明发生在蚊子身上，最近在人类身上也出现了一个。在这些案例中，这些基因的反式剪接都是偶然发现的。因此，反式剪接的真实程度是未知的。此外，对反式剪接涉及的机制一无所知。我们最近使用深度测序技术鉴定了80个在果蝇体内反式剪接的新基因。该项目的长期目标是进一步探索果蝇的反式剪接情况，并确定这一过程中涉及的机制。我们将首先对不同果蝇物种发育过程中的反式剪接基因进行详尽的调查。其次，我们将进行实验，以测试反式剪接的mRNAs是否被翻译并具有功能。最后，这些实验将得到生化、遗传学、基因组、细胞生物学和生物信息学实验的补充，这些实验旨在确定反式剪接发生的机制。总之，这些实验将为反式剪接的机制提供巨大的洞察力，这是一个完全未被研究但却很重要的过程。考虑到反式剪接在治疗人类疾病中的潜在效用，以及最近在人类中反式剪接的mRNAs与癌症有关，我们的发现很可能与人类健康直接相关。 公共卫生相关性： 这些实验将为反式剪接的机制提供巨大的洞察力。考虑到反式剪接在治疗人类疾病中的潜在效用，以及最近在人类中反式剪接的mRNAs与癌症有关，我们的发现很可能与人类健康直接相关。

DESCRIPTION (provided by applicant): Abstract: Pre-messenger RNA (pre-mRNA) splicing is a process in which introns are removed to join exons together. This process is used during the synthesis of nearly all metazoan mRNAs (~93% of human mRNAs) and is an important means of regulating gene expression. With only a handful of exceptions, splicing occurs in cis - the exons that are joined together are located on the same pre-mRNA. In some eukaryotes, including nematodes, trypanosomes and planarians, splicing can occur in trans. In these cases, a specialized spliced leader RNA is spliced to the 5' end of protein coding RNAs. Interestingly, there are a few cases where a distinct type of trans-splicing has been shown to occur, namely, the splicing of exons from protein coding genes. The two best characterized examples are the mod(mdg4) and lola genes from Drosophila. In each of these genes, a group of common 5' exons can be spliced in trans to one of several (26 for mod(mdg4) or 22 for lola) variable 3' exons. One convincing case of trans-splicing has been shown to occur in mosquitos and one has been recently shown in humans. In each of these cases, trans-splicing of these genes was discovered fortuitously. As a result, the true extent of trans-splicing is unknown. Moreover, nothing is known about the mechanisms involved in trans-splicing. We have recently used deep sequencing to identify 80 new genes that are trans-spliced in Drosophila. The long- term goals of this project are to further explore the trans-splicing landscape in Drosophila and to determine the mechanisms involved in this process. We will first perform an exhaustive survey for trans-spliced genes throughout development in different Drosophila species. Second, we will perform experiments to test whether the trans-spliced mRNAs are translated and are functional. Finally, these experiments will be complemented by a combination of biochemical, genetic, genomic, cell biology, and bioinformatics experiments designed to determine the mechanisms by which trans- splicing occurs. Together these experiments will provide tremendous insight into the mechanisms of trans-splicing, a completely understudied yet important process. Given the potential utility of trans-splicing in treating human diseases and that trans-spliced mRNAs in humans have recently been linked to cancer, it is likely the discoveries we make will be of direct relevance to human health. PUBLIC HEALTH RELEVANCE: These experiments will provide tremendous insight into the mechanisms of trans- splicing. Given the potential utility of trans-splicing in treating human diseases and that trans-spliced mRNAs in humans have recently been linked to cancer, it is likely the discoveries we make will be of direct relevance to human health.