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.
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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)