Functional genomics of evolutionary conserved long noncoding RNAs in Drosophila

果蝇进化保守长非编码RNA的功能基因组学

• 批准号: 1716532
• 负责人: Carlos Machado
• 金额: $ 69万
• 依托单位: University of Maryland, College Park
• 依托单位国家: 美国
• 项目类别: Standard Grant
• 财政年份: 2017
• 资助国家: 美国
• 起止时间: 2017-08-01 至 2022-07-31
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=1716532&HistoricalAwards=false
• 关键词: Functional; genomics; evolutionary; conserved; long

Biologists have recently discovered that a large fraction of the genomes of most organisms, previously thought to be "junk DNA", has biochemical activity and may be functional. Some of the genetic elements found in those regions of the genome do not code for proteins (the building blocks of cells) and have thus been named "non-coding RNAs". A few of these non-coding RNAs are known to play important roles in the biology of organisms, mainly by their capacity to turn other genes on or off. However, it is not clear if most of the non-coding RNAs that have been catalogued are even functional. This project seeks to address that gap in understanding by using patterns of evolutionary conservation in two fruit fly species to identify candidate non-coding RNAs for functional characterization. Because non-coding RNAs are found in many organisms, from flies to plants to humans, the results could have far-reaching impact. The work will be carried out by college students at graduate and undergraduate levels, thus providing them with valuable training experiences. In addition, the project will provide research experience and support with curriculum development for K-12 science teachers. One of the most exciting recent advances in molecular biology has been the discovery of a large number of long non-coding RNAs (lncRNAs) in the transcriptomes of eukaryotic organisms. Although about a dozen lncRNAs are known to play important roles in a broad spectrum of developmental and physiological phenomena, our understanding of lncRNA function lags significantly behind our ability to quickly catalog them using RNA sequencing and computational approaches. Given that there are thousands of lncRNAs in any given genome, one essential approach to select relevant lncRNAs for functional characterization is to focus on lncRNAs with strong patterns of evolutionary conservation. This project will focus on a group of lncRNAs that are conserved between Drosophila melanogaster and D. pseudoobscura, species that diverged from their common ancestor 25-40 million years ago. These lncRNAs show small but detectable levels of DNA sequence conservation, conserved synteny, and conserved developmental expression patterns that suggest shared function in the two species. This project will characterize the phenotypic effects of that group of evolutionary conserved lncRNAs in Drosophila. Aims will include first confirming that the conserved putative lncRNAs are independent untranslated transcripts using a combination of IsoSeq and Riboseq data, then identifying their patterns of tissue-specific expression in both species using RNAseq, and finally targeting a group of the lncRNAs for functional analysis using using transcript disruption enabled by CRISPR/Cas9 genome editing in D. melanogaster. The methods developed in this study should have broad applicability across other organisms, and the functional studies should add substantially to our understanding of what lncRNAs do.

生物学家最近发现，大多数生物体的大部分基因组，以前被认为是“垃圾DNA”，具有生化活性，可能是功能性的。在基因组的这些区域中发现的一些遗传元件不编码蛋白质（细胞的构建块），因此被命名为“非编码RNA”。已知这些非编码RNA中的一些在生物体生物学中发挥重要作用，主要是通过它们打开或关闭其他基因的能力。然而，目前尚不清楚大多数已被编目的非编码RNA是否具有功能。该项目旨在通过使用两种果蝇物种的进化保守模式来识别候选非编码RNA以进行功能表征来解决理解上的差距。由于非编码RNA存在于许多生物体中，从苍蝇到植物再到人类，这一结果可能会产生深远的影响。这项工作将由研究生和本科生进行，从而为他们提供宝贵的培训经验。此外，该项目还将为K-12科学教师的课程开发提供研究经验和支持。分子生物学领域最令人兴奋的最新进展之一是在真核生物的转录组中发现了大量长链非编码RNA（lncRNA）。虽然已知大约有十几种lncRNA在广泛的发育和生理现象中发挥重要作用，但我们对lncRNA功能的理解明显落后于我们使用RNA测序和计算方法快速对其进行分类的能力。考虑到在任何给定的基因组中存在数千种lncRNA，选择相关lncRNA用于功能表征的一种基本方法是关注具有强进化保守模式的lncRNA。本项目将重点研究一组在果蝇和果蝇之间保守的lncRNA。pseudoobscura，2500 - 4000万年前从共同祖先分化出来的物种。这些lncRNA显示出小但可检测水平的DNA序列保守性、保守的同线性和保守的发育表达模式，表明这两个物种具有共同的功能。这个项目将描述果蝇中进化保守lncRNA组的表型效应。目的将包括首先使用IsoSeq和Riboseq数据的组合确认保守的推定lncRNA是独立的非翻译转录物，然后使用RNAseq鉴定它们在两个物种中的组织特异性表达模式，最后使用由D中的CRISPR/Cas9基因组编辑实现的转录物破坏来靶向一组lncRNA进行功能分析。黑腹菌这项研究中开发的方法应该在其他生物体中具有广泛的适用性，并且功能研究应该大大增加我们对lncRNA作用的理解。

项目成果

Inversions shape the divergence of Drosophila pseudoobscura and D. persimilis on multiple timescales

反演塑造了伪暗果蝇和 D. persimilis 在多个时间尺度上的分歧

• DOI: 10.1101/842047
• 发表时间: 2021
• 期刊: Evolution
• 影响因子: 3.3
• 作者: Korunes, Katharine L.;Machado, Carlos A.;Noor, Mohamed A.
• 通讯作者: Noor, Mohamed A.

Phylogenetic diversity of two common Trypanosoma cruzi lineages in the Southwestern United States

美国西南部两个常见克氏锥虫谱系的系统发育多样性

• DOI: 10.1016/j.meegid.2022.105251
• 发表时间: 2022
• 期刊: Genetics and Evolution
• 作者: Flores-López, Carlos A.;Mitchell, Elizabeth A.;Reisenman, Carolina E.;Sarkar, Sahotra;Williamson, Philip C.;Machado, Carlos A.
• 通讯作者: Machado, Carlos A.