Identification of long non-coding RNAs and small peptide-coding transcripts in soybean

Ph.D.Long non-coding RNAs (lncRNAs) are generally defined as non-protein-coding transcripts that are at least 200 nucleotides in lengths. They are known to play a pivotal role in regulation of gene expression at all levels. In plants, studies have also implicated them as having a possible role in stress responses. In this study, a large collection of in-house transcriptome data on different soybean tissues in various conditions to perform a comprehensive identification of soybean lncRNAs was taken as an advantage. Existing soybean transcriptome data from public sources that are of sufficient quality and sequencing depth were also retrieved to enrich our analysis. In total, 261 data samples have been used for this analysis. An integrated reference-based, de novo transcript assembly was developed, which identified ~90,000 lncRNA gene loci. It can be shown that lncRNA genes are distinct from both protein-coding genes and genomic background noises in terms of length, number of exons, transposable element composition and sequence conservation levels across legume species. They are possibly related to biological processes because they show tissue-specific and germplasm-specific transcriptional responses under various stress conditions. Transcription start sites of lncRNA gene loci tend to be close to their nearest protein-coding genes and they might be transcriptionally related to each other, particularly for antisense and intronic lncRNA genes. LncRNA gene loci also share distinct epigenetic marks, such as histone modification and DNA methylation, with protein-coding gene loci.The current definition of “non-protein-coding” in a transcript refers to the absence of open-reading frame (ORF) encoding at least 100 amino acids (aa) in length. However, small ORFs that encode between 10 and 100 aa were routinely predicted in the lncRNA sequences. Mass spectrometry was used to identify a novel subset of small peptide-coding transcripts in these novel lncRNA loci, which opened a new avenue for investigating their functional roles. The results also highlighted the current inadequacy of the bioinformatics definition of lncRNA, which does not consider the presence of small ORFs.In summary, results presented in this study have laid the groundwork for studying lncRNA in soybean and provided evidence for further functional study.長非編碼核糖核苷酸普遍被定義爲長度大於二百堿基、且不能被轉譯成蛋白序列的轉錄本。它們在不同層次的基因調控中的重要作用是衆所周知的。一些研究指出，它們可能在植物的壓力反應中會起一定的作用。本研究中利用了自家的龐大的轉錄組數據用以全面地預測大豆的長非編碼核糖核苷酸。這一龐大的數據涵蓋了豐富多樣的組織和環境因素。同時，大量高深度的優良的大豆轉錄組數據也被從公用數據庫提取，用以豐富本研究。總的來説，本研究一共使用了261個樣品的數據來進行預測分析。利用這些數據，一個自家的生物信息流程被開發并用以預測大豆的長非編碼基因。這個流程整合了基於參考基因組和全新轉錄組組裝的方法，成功預測了大約九萬個長非編碼基因位點。仔細分析發現，這些預測得到的長非編碼基因在轉錄本長度、外顯子數目、轉座子成分和在豆科植物中的序列保守程度都有別于蛋白編碼基因和基因組的背景。另外，這些長非編碼基因在不同環境中呈現出組織和品種特異的表達模式，進一步説明了它們可能與生物過程有關。長非編碼基因的轉錄起始點趨向靠近它們最近的蛋白編碼基因的轉錄起始點，反映著長非編碼基因和蛋白編碼基因在轉錄上的相關性。這一現象在反義和内含子長非編碼基因中尤其明顯。從表觀遺傳學的角度來看，長非編碼基因呈現出和蛋白編碼基因不一樣的模式，例如在組蛋白修飾和脫氧核糖核酸甲基化方面。目前來説，“非編碼”一般是指缺少大於一百氨基酸的開放閲讀框架。然而，在長非編碼核糖核苷酸中常會出現可能會被翻譯成長度介於十到一百氨基酸的小型開放閲讀框架。質譜實驗發現大豆長非編碼基因位點中的部分轉錄本具有編譯小型多肽的能力。這爲本研究長非編碼基因位點的功能開展了一個新的方向。同時，該研究結果强調了目前生物信息學對長非編碼核糖核苷酸定義上的不足。這一不足體現在它缺少考慮小型開放閲讀框架。總結來説，本研究結果爲研究大豆長非編碼核糖核苷酸打下了夯實的基礎，并爲之後將會開展的功能研究提供有力的證據。Lin, Xiao."December 2019."Thesis Ph.D. Chinese University of Hong Kong 2020.Includes bibliographical references (leaves 96-123).Abstracts also in Chinese.Title from PDF title page (viewed on 20 May, 2021)