Single cell transcriptome analysis of MCF-7 reveals consistently and inconsistently expressed gene groups each associated with distinct cellular localization and functions

PLOS ONE, Nov 2019

Single cell transcriptome (SCT) analysis provides superior resolution to illustrate tumor cell heterogeneity for clinical implications. We characterized four SCTs of MCF-7 using 143 housekeeping genes (HKGs) as control, of which lactate dehydrogenase B (LDHB) expression is silenced. These SCT libraries mapped to 11,423, 11,486, 10,380, and 11,306 RefSeq genes (UCSC), respectively. High consistency in HKG expression levels across all four SCTs, along with transcriptional silencing of LDHB, was observed, suggesting a high sensitivity and reproducibility of the SCT analysis. Cross-library comparison on expression levels by scatter plotting revealed a linear correlation and an 83–94% overlap in transcript isoforms and expressed genes were also observed. To gain insight of transcriptional diversity among the SCTs, expressed genes were split into consistently expressed (CE) (expressed in all SCTs) and inconsistently expressed (IE) (expressed in some but not all SCTs) genes for further characterization, along with the 142 expressed HKGs as a reference. Distinct transcriptional strengths were found among these groups, with averages of 1,612.0, 88.0 and 1.2 FPKM for HKGs, CE and IE, respectively. Comparison between CE and IE groups further indicated that expressions of CE genes vary more significantly than that of IE genes. Gene Ontology analysis indicated that proteins encoded by CE genes are mainly involved in fundamental intracellular activities, while proteins encoded by IE genes are mainly for extracellular activities, especially acting as receptors or ion channels. The diversified gene expressions, especially for those encoded by IE genes, may contribute to cancer drug resistance.

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Single cell transcriptome analysis of MCF-7 reveals consistently and inconsistently expressed gene groups each associated with distinct cellular localization and functions

June Single cell transcriptome analysis of MCF-7 reveals consistently and inconsistently expressed gene groups each associated with distinct cellular localization and functions Yih-Shien Chiang 0 1 Yu-Feng Huang 0 1 Mohit K. Midha 0 1 Tzu-Han Chen 0 1 Hsin- Chieh Shiau 0 1 Kuo-Ping Chiu 0 1 0 Genomics Research Center , Academia Sinica, Taipei, Taiwan , 2 Institute of Biochemistry and Molecular Biology, National Yang-Ming University , Taipei, Taiwan , 3 College of Life Science, National Taiwan University , Taipei , Taiwan 1 Editor: Aamir Ahmad, University of South Alabama Mitchell Cancer Institute , UNITED STATES Single cell transcriptome (SCT) analysis provides superior resolution to illustrate tumor cell heterogeneity for clinical implications. We characterized four SCTs of MCF-7 using 143 housekeeping genes (HKGs) as control, of which lactate dehydrogenase B (LDHB) expression is silenced. These SCT libraries mapped to 11,423, 11,486, 10,380, and 11,306 RefSeq genes (UCSC), respectively. High consistency in HKG expression levels across all four SCTs, along with transcriptional silencing of LDHB, was observed, suggesting a high sensitivity and reproducibility of the SCT analysis. Cross-library comparison on expression levels by scatter plotting revealed a linear correlation and an 83±94% overlap in transcript isoforms and expressed genes were also observed. To gain insight of transcriptional diversity among the SCTs, expressed genes were split into consistently expressed (CE) (expressed in all SCTs) and inconsistently expressed (IE) (expressed in some but not all SCTs) genes for further characterization, along with the 142 expressed HKGs as a reference. Distinct transcriptional strengths were found among these groups, with averages of 1,612.0, 88.0 and 1.2 FPKM for HKGs, CE and IE, respectively. Comparison between CE and IE groups further indicated that expressions of CE genes vary more significantly than that of IE genes. Gene Ontology analysis indicated that proteins encoded by CE genes are mainly involved in fundamental intracellular activities, while proteins encoded by IE genes are mainly for extracellular activities, especially acting as receptors or ion channels. The diversified gene expressions, especially for those encoded by IE genes, may contribute to cancer drug resistance. - Funding: This work was supported by Summit Project provided by Academia Sinica and PI funding provided by Genomics Research Center, Academia Sinica. The funder had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. Introduction Cancer is known to result from progressive accumulation of genomic and epigenomic alterations, leading to dysregulated cell growth [ 1, 2 ]. The process of carcinogenesis and subsequent cancer development is strongly enhanced by chromosome instability, a hallmark of cancer, causing the formation of diversified cancer genomes in the cancer mass, each of which drives a cell-specific transcriptome. Through an evolutionary process, cancer cells carrying lethal mutations undergo apoptosis or necrosis, leaving the rest to continue the journey of cancer development in an uncontrolled manner [ 3, 4 ]. Due to the advances in Next-Generation Sequencing (NGS) technologies, cancer genomes and associated transcriptomes can be analyzed at single cell level [ 5, 6 ]. Due to the fact that it is able to provide a superior resolution as enhanced by NGS, single cell transcriptome (SCT) analysis has a strong potential to facilitate our understanding of cancer evolution and its transcriptional heterogeneity [7, 8], which is believed to be associated with acquired drug resistance [ 9 ]. However, although with a great potential [ 10 ], SCT data from cancerous cells remain very limited. Here, we conducted SCT sequencing on MCF-7 breast cancer cell line to reveal their transcriptional diversity and potential clinical implications. MCF-7 is one of the most widely used metastatic breast cancer cell line. It retains several characteristics of differentiated mammary epithelium (https://www.atcc.org/Products/All/HTB-22.aspx#characteristics). On the other hand, breast cancer cells such as those characterized as HER2-positive and TNBC (Triple Negative Breast Cancer) have already experienced intensive selection process to abolish the expressions of some key genes, which may further affect the expressions of many other genes to make the transcriptome more subtype-specific. We prefer to use MCF-7 due to the fact that it represents a more general form of breast cancer and has a better potential to express both consistently expressed and inconsistently expressed genes. The analysis employs RNA-Seq approach [ 11 ], in conjunction with Tophat and Cufflinks software encompassed in UCSC Galaxy pipeline. To evaluate the reliability of our approach, we used 143 housekeeping genes (HKGs) agreed by two independent groups [ 12, 13 ], as control. HKGs, originally def (...truncated)


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Yih-Shien Chiang, Yu-Feng Huang, Mohit K. Midha, Tzu-Han Chen, Hsin-Chieh Shiau, Kuo-Ping Chiu. Single cell transcriptome analysis of MCF-7 reveals consistently and inconsistently expressed gene groups each associated with distinct cellular localization and functions, PLOS ONE, 2018, Volume 13, Issue 6, DOI: 10.1371/journal.pone.0199471