探究日本鰻成長與成熟階段編碼與長非編碼RNA轉錄體的表現

Abstract

這項研究使用了NCBI資料庫中的日本鰻（Anguilla japonica）RNA-seq原始定序資料，涵蓋卵、前期柳葉鰻、柳葉鰻、玻璃鰻和銀鰻等五個不同發育階段的28筆轉錄體定序數據。利用Trinity轉錄體組裝程式，參考實驗室先前組裝的日本鰻基因體，建立了從卵階段到銀鰻階段的完整轉錄體序列，共獲得約97萬條轉錄序列。通過一系列篩選機制，成功鑑定了31,189條長非編碼RNA序列，並將這些資料與原有的58,329條蛋白質編碼序列結合，構建了一個更全面的轉錄體註解資料庫，用於探討不同狀態下的轉錄體變化及差異基因群的生理調控機制。 基於親緣關係相近的8個物種蛋白質體分析結果顯示，GO富集分析結果顯示，日本鰻和歐洲鰻特有的基因群組在細胞過程和代謝過程方面表現顯著。這些基因群組在多種重要的分子功能和細胞結構中發揮關鍵作用，顯示出這兩種鰻魚在生物學功能上的差異與相似之處。 根據不同環境下大腦、肝臟和卵巢的基因分析結果，野生雌鰻在嗅覺感知、基因動態變化、信號傳導、代謝調節和免疫反應等多方面的基因表現顯著高於養殖雌鰻，反映出野生環境對其生理功能的更高需求。隨著鰻魚從玻璃鰻階段逐步成長至銀鰻階段，鈣離子動態調節、GTPase調節因子活性及酶調控等基因表現顯著提升，以適應生長需求和環境變化。 關於長非編碼RNA（LncRNA）的表達，結果顯示其在日本鰻不同組織中的表達在野生和養殖環境下存在顯著差異。在大腦組織中，野生環境下的LncRNA（如TRINITY_GG_176304_c0_g1_i5）表現量顯著高於養殖環境，與免疫調節和蛋白質運輸相關。相反，養殖環境下的LncRNA（如TRINITY_GG_215607_c0_g3_i10）顯著高於野生環境，與免疫相容性抗原相關。在肝臟和卵巢組織中，野生環境下的LncRNA表現量顯著高於養殖環境，分別與醣類代謝和生殖過程相關。這些結果反映了鰻魚在不同環境壓力下的基因調控機制和生理功能變化。 本研究分析了日本鰻在不同環境下的基因表達差異，發現養殖與野生環境中特定基因表達顯著不同，這可能與免疫和生理功能變化相關。特別是一些與免疫反應增強相關的基因在養殖環境中高表達，這為未來基因功能研究和對鰻魚生物學的理解提供了重要資訊。此外，研究還深入分析了影響生殖調控的轉錄序列，探討這些基因在不同發育階段如何影響鰻魚的性成熟和繁殖能力。通過分析與性腺發育相關的轉錄因子和激素信號，揭示了鰻魚生殖調控機制的重要方面，這有助於理解野生和養殖環境下日本鰻的生殖行為差異，並為促進養殖業的可持續發展提供理論支持。

This study utilized raw RNA-seq data from the NCBI database for Japanese eel (Anguilla japonica), covering 28 transcriptome sequencing datasets across five different developmental stages: egg, early leptocephalus, leptocephalus, glass eel, and silver eel. Using the Trinity transcriptome assembly program and referencing the previously assembled genome of the Japanese eel from the laboratory, a complete transcriptome sequence was established from the egg stage to the silver eel stage, obtaining approximately 970,000 transcript. Through a series of filtering mechanisms, 31,189 long non-coding RNA (lncRNA) sequences were successfully identified. These data were combined with the existing 58,329 protein-coding sequences to construct a more comprehensive transcriptome annotation database, which was used to explore transcriptome changes under different environments and the physiological regulatory mechanisms of differential gene groups. Based on proteomic analysis results of eight phylogenetically related species, GO enrichment analysis showed that gene groups specific to the Japanese eel and European eel exhibited significant expression in cellular and metabolic processes. These gene groups play key roles in various important molecular functions and cellular structures, indicating both differences and similarities in biological functions between these two eel species. According to gene analysis results from three tissues under different environments, wild female eels showed significantly higher gene expression than farmed female eels in olfactory perception, gene dynamic changes, signal transduction, metabolic regulation, and immune response, reflecting a higher demand for physiological functions in the wild environment. As the eels grew from the glass eel stage to the silver eel stage, gene expressions related to calcium ion dynamic regulation, GTPase regulatory factor activity, and enzyme regulation were significantly elevated to adapt to growth needs and environmental changes. Regarding the expression of long non-coding RNAs (lncRNAs), the results showed significant differences in their expression in different tissues of Japanese eels under wild and farmed conditions. In brain tissue, lncRNAs (such as TRINITY_GG_176304_c0_g1_i5) were significantly more expressed in the wild environment than in the farmed environment, related to immune regulation and protein transport. Conversely, in the farmed environment, lncRNAs (such as TRINITY_GG_215607_c0_g3_i10) were significantly more expressed than in the wild environment, related to immune compatibility antigens. In liver and ovary tissues, lncRNA expressions were significantly higher in the wild environment than in the farmed environment, associated with carbohydrate metabolism and reproductive processes, respectively. These results reflect the gene regulatory mechanisms and physiological function changes of eels under different environmental stresses. This study analyzed the gene expression differences of Japanese eels under different environments and found significant differences in the expression of specific genes in farmed and wild environments, which may be related to immune and physiological function changes. Notably, some genes associated with enhanced immune response were highly expressed in the farmed environment, providing important information for future gene function research and understanding eel biology. Additionally, the study deeply analyzed the transcript sequences affecting reproductive regulation, exploring how these genes influence the sexual maturity and reproductive capacity of eels at different developmental stages. By analyzing transcription factors and hormone signals related to gonadal development, important aspects of eel reproductive regulation mechanisms were revealed, aiding in understanding the reproductive behavior differences of Japanese eels in wild and farmed environments, and providing theoretical support for the sustainable development of the aquaculture industry.