利用核酸定序技術研究麩醯胺酸匱乏對大腸癌細胞外泌體RNA組成的影響;Exploring the Impact of Glutamine Deprivation on RNA Cargo Profiles of Colorectal Cancer Cell Exosomes by Nucleotide Sequencing

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Title:	利用核酸定序技術研究麩醯胺酸匱乏對大腸癌細胞外泌體RNA組成的影響;Exploring the Impact of Glutamine Deprivation on RNA Cargo Profiles of Colorectal Cancer Cell Exosomes by Nucleotide Sequencing
Authors:	劉柏渝;Liu, Bo-Yu
Contributors:	生命科學系
Keywords:	外泌體;大腸癌細胞;核酸定序;Exosomes;Colorectal Cancer Cell;Nucleotide Sequencing
Date:	2024-09-20
Issue Date:	2024-10-09 15:29:05 (UTC+8)
Publisher:	國立中央大學
Abstract:	麩醯胺酸（Glutamine）在癌細胞中扮演著重要的角色。作為一種非必需胺基酸，其在正常細胞中的需求遠低於腫瘤細胞。在癌症細胞中，麩醯胺酸主要通過麩醯胺酸分解代謝（glutaminolysis）途徑被代謝，轉化為麩胺酸（glutamate）和氨（ammonia），進一步生成α-酮戊二酸（α-Ketoglutarate）並進入三羧酸循環（TCA cycle），從而為腫瘤細胞的快速生長和增殖提供能量和代謝中間體。在腫瘤微環境中，由於癌細胞的大量消耗，麩醯胺酸往往呈現匱乏狀態。這種匱乏狀態對外泌體的產生和功能有顯著影響。有研究表明，麩醯胺酸匱乏會增加生成於Rab11陽性回收內涵體(recycling endosomes)的癌細胞外泌體數量，這些外泌體中攜帶的分子能夠影響腫瘤微環境中的其他細胞，幫助腫瘤適應代謝壓力。外泌體是由細胞釋放的微小細胞外囊泡，其直徑在30~150 nm，其中富含蛋白質、核酸和脂質。其中，RNA分子在外泌體中的功能極為關鍵，它們能夠被轉移到腫瘤微環境中的其他細胞，執行功能，從而促進腫瘤的增長和擴散。近年來，有研究利用定序技術，如奈米孔定序和NGS Illumina平台，進一步探討外泌體中RNA的組成以及推測其生物機制功能，這些先進的定序技術不僅提供了對外泌體RNA種類和豐富度的全面觀察，還能使研究者能夠識別特定RNA分子在癌症進程中的角色。由於麩醯胺酸匱乏與癌症惡化相關的研究仍然不足，因此，本研究利用奈米孔定序和NGS定序來研究麩醯胺酸匱乏下的外泌體的RNA組成。本研究發現，大腸癌HCT116細胞在正常麩醯胺酸與麩醯胺酸匱乏下分泌的外泌體RNA組成並不相同。重要的是，藉由KEGG訊息路徑富集分析，我們發現這些表現量不同的RNA的蛋白產物集中參與於RAS、PI3k-AKT與Rap1訊息傳遞，而這些訊息傳遞路徑多與癌症惡化有關。因此，本研究增進了我們對於麩醯胺酸匱乏所誘發之外泌體的功能了解。我們期待這些研究發現不僅能夠增進對腫瘤生物學的理解，也為發展新的癌症治療策略提供了可能。;Glutamine plays a crucial role in cancer cells. As a nonessential amino acid, its demand in tumor cells far exceeds that in normal cells. In cancer cells, glutamine is primarily metabolized through the glutaminolysis pathway, converting into glutamate and ammonia, further synthesizing α-ketoglutarate and entering the tricarboxylic acid (TCA) cycle, thereby providing energy and metabolic intermediates necessary for the rapid growth and proliferation of tumor cells. In the tumor microenvironment, due to the extensive consumption by cancer cells, glutamine often becomes depleted. This state of depletion significantly affects the production and function of exosomes. Studies have shown that glutamine depletion can increase the number of cancer exosomes released by Rab11-positive recycling endosomes, which carry molecules that can affect other cells in the tumor microenvironment and help the tumor adapt to metabolic stress. Exosomes are tiny extracellular vesicles released by cells, ranging in size from 30 to 150 nm and rich in proteins, nucleic acids, and lipids. Particularly, the role of RNA molecules within exosomes is crucial; they can be transferred to other cells in the tumor microenvironment, thus promoting tumor growth and disease spread. Recent studies using sequencing technologies, such as the nanopore sequencing and the NGS Illumina platforms, have further explored the RNA composition, leading to a better understanding of biological mechanisms of exosomes. These advanced sequencing techniques not only provide a comprehensive observation of the types and abundance of RNA within exosomes but also enable researchers to identify specific RNA molecules′ roles in cancer progression. Due to the insufficient research on the relationship between glutamine deprivation and cancer progression, this study utilizes nanopore sequencing and NGS sequencing to investigate the RNA composition of exosomes under glutamine deprivation. This study found that the RNA composition of exosomes secreted by colorectal cancer HCT116 cells differs between normal glutamine conditions and glutamine deprivation. Importantly, using the KEGG pathway enrichment analysis we found that the protein products of these differentially expressed RNAs are primarily involved in the RAS, PI3K-AKT, and Rap1 signaling pathways, which are often associated with cancer progression. Therefore, this study enhances our understanding of the functions of exosomes induced by glutamine deprivation. We hope that these findings will not only advance the understanding of tumor biology but also provide potential avenues for developing new cancer treatment strategies.
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