INVADEseq to identify cell-adherent or invasive bacteria and the associated host transcriptome at single-cell-level resolution.
INVADEseq to identify cell-adherent or invasive bacteria and the associated host transcriptome at single-cell-level resolution.
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DOI：
10.1038/s41596-023-00888-7
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发表时间：
2023-11
期刊：
影响因子：
14.8
通讯作者：
Bullman, Susan
中科院分区：
文献类型：
--
作者：
Nino, Jorge Luis Galeano;Wu, Hanrui;Lacourse, Kaitlyn D.;Srinivasan, Harini;Fitzgibbon, Matthew;Minot, Samuel S.;Sather, Cassie;Johnston, Christopher D.;Bullman, Susan
单细胞RNA测序（scRNAseq）技术在揭示和描述哺乳动物组织（包括实体瘤）内的细胞异质性方面是有益的。然而，这些技术中的许多应用RNA的多聚（A）选择，并且因此主要集中于确定肿瘤微环境的真核细胞组分的基因特征。微生物组分析揭示了主要癌症类型的人类肿瘤组织中存在微生物生态系统，包括细菌和真菌。影像资料显示，肿瘤内的细菌可能位于上皮细胞和免疫细胞类型。然而，由于细菌RNA通常缺乏poly（A）尾，标准scRNAseq方法捕获肿瘤微环境的这种微生物组分的能力有限。为了克服这一点，我们描述了侵入粘附定向表达测序（INVADEseq）方法，通过引入除了真核多聚（A）RNA选择的标准引物之外的靶向细菌16S核糖体RNA基因保守区域的引物，我们调整了10x Genomics 5′ scRNAseq方案。这种“附加”方法能够以真核单细胞水平分辨率生成真核和细菌DNA文库，利用10 x条形码来鉴定具有细胞内细菌的单细胞。INVADEseq方法需要30小时才能完成，包括组织处理，测序和计算分析。作为输出，INVADEseq已被证明是人类癌细胞系和患者肿瘤标本中的可靠工具，其通过检测携带细菌的人类细胞的比例以及人类细胞和细胞内细菌的身份，沿着鉴定基于相关细菌调节的宿主转录程序。
Single-cell RNA sequencing (scRNAseq) technologies have been beneficial in revealing and describing cellular heterogeneity within mammalian tissues, including solid tumors. However, many of these techniques apply poly(A) selection of RNA, and thus have primarily focused on determining the gene signatures of eukaryotic cellular components of the tumor microenvironment. Microbiome analysis has revealed the presence of microbial ecosystems, including bacteria and fungi, within human tumor tissues from major cancer types. Imaging data have revealed that intratumoral bacteria may be located within epithelial and immune cell types. However, as bacterial RNA typically lacks a poly(A) tail, standard scRNAseq approaches have limited ability to capture this microbial component of the tumor microenvironment. To overcome this, we describe the invasion–adhesion-directed expression sequencing (INVADEseq) approach, whereby we adapt 10x Genomics 5′ scRNAseq protocol by introducing a primer that targets a conserved region of the bacterial 16S ribosomal RNA gene in addition to the standard primer for eukaryotic poly(A) RNA selection. This ‘add-on’ approach enables the generation of eukaryotic and bacterial DNA libraries at eukaryotic single-cell level resolution, utilizing the 10x barcode to identify single cells with intracellular bacteria. The INVADEseq method takes 30 h to complete, including tissue processing, sequencing and computational analysis. As an output, INVADEseq has shown to be a reliable tool in human cancer cell lines and patient tumor specimens by detecting the proportion of human cells that harbor bacteria and the identities of human cells and intracellular bacteria, along with identifying host transcriptional programs that are modulated on the basis of associated bacteria.
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影响因子：
30.8
作者：
Joanito, Ignasius;Wirapati, Pratyaksha;Zhao, Nancy;Nawaz, Zahid;Yeo, Grace;Lee, Fiona;Eng, Christine L. P.;Macalinao, Dominique Camat;Kahraman, Merve;Srinivasan, Harini;Lakshmanan, Vairavan;Verbandt, Sara;Tsantoulis, Petros;Gunn, Nicole;Venkatesh, Prasanna Nori;Poh, Zhong Wee;Nahar, Rahul;Oh, Hsueh Ling Janice;Loo, Jia Min;Chia, Shumei;Cheow, Lih Feng;Cheruba, Elsie;Wong, Michael Thomas;Kua, Lindsay;Chua, Clarinda;Nguyen, Andy;Golovan, Justin;Gan, Anna;Lim, Wan-Jun;Guo, Yu Amanda;Yap, Choon Kong;Tay, Brenda;Hong, Yourae;Chong, Dawn Qingqing;Chok, Aik-Yong;Park, Woong-Yang;Han, Shuting;Chang, Mei Huan;Seow-En, Isaac;Fu, Cherylin;Mathew, Ronnie;Toh, Ee-Lin;Hong, Lewis Z.;Skanderup, Anders Jacobsen;DasGupta, Ramanuj;Ong, Chin-Ann Johnny;Lim, Kiat Hon;Tan, Emile K. W.;Koo, Si-Lin;Leow, Wei Qiang;Tejpar, Sabine;Prabhakar, Shyam;Tan, Iain Beehuat
通讯作者：
Tan, Iain Beehuat
单细胞 RNA 测序强调了炎性癌症相关成纤维细胞在膀胱尿路上皮癌中的作用。
影响因子：
16.6
作者：
Chen Z;Zhou L;Liu L;Hou Y;Xiong M;Yang Y;Hu J;Chen K
通讯作者：
Chen K
影响因子：
64.5
作者：
Dohlman, Anders B.;Klug, Jared;Mesko, Marissa;Gao, Iris H.;Lipkin, Steven M.;Shen, Xiling;Iliev, Iliyan D.
通讯作者：
Iliev, Iliyan D.
影响因子：
64.8
作者：
通讯作者：
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影响因子：
64.8
作者：
Kennedy, Katherine M.;de Goffau, Marcus C.;Walter, Jens
通讯作者：
Walter, Jens