On October17th, 2018, Junling Jia’ s team of the Life Sciences Institute of Zhejiang University and the MinZheng’ s team of the First Affiliated Hospital of Zhejiang University jointly publisheda paper entitled"Holo-Seq: single-cell sequencing ofholo-transcriptome" in Genome Biology.
Single-cell RNA-seq technology has become a very powerful tool for studying cellular heterogeneity and has driven many important advances in developmental biology and cancer biology.Current single-cell RNA-seq approaches are hindered by preamplification bias, loss of strand of origin information and the inability to observe small-RNA and mRNA dual transcriptomes. Here, we introduce a single-cell holo-transcriptome sequencing (Holo-Seq)adapting the conventional bulk RNA-Seq approach that overcomes all three hurdles. Holo-Seq has the same quantitative accuracy and uniform coverage with a complete strand of origin information as bulk RNA-seq. Most importantly, Holo-Seq can simultaneously observe small-RNAs and mRNAs in a single cell.
a. An RPKM scatterplot of expressed genes between Smart-Seq2 and bulk mRNA-Seq. b. An RPKM scatterplot of expressed genes between Holo-Seq (mRNA) and bulk mRNA-Seq.
Furthermore, we applied Holo-Seq to probe small RNA-mRNAdual transcriptomes from 32 single cells isolated from a human hepatocellular carcinoma. We found three expression-based-subpopulations (Exp-subpopulations) with six featured transcript groupsand three super-enhancer-based-subpopulations (SE-subpopulations). We also inferred a potential hepatic neoplasm kinetics model showing that:1.HCC malignant transition couples with genome-wide super-enhancer remodeling; 2. the downregulation of mitochondrial activity and the upregulation of both tumor suppressor miRNA and oncomiRs happen at the early stage of malignant transition before activating tumorigenesis signaling pathways.These findings have important implications and reference for the detection and diagnosis of hepatocellular carcinoma.
a. Unsupervised hierarchical clustering analysis of 32 HCC single cells based on the variant mRNAs and miRNAs
b. Correlation heat map of the genome-wide super-enhancer activity of32 HCC single cells (Left panel).The cells are rankedin the same order as figure a.Right panels is the relative expression heat map of the gene sets related to mitochondrial activity, oncogenic signals pathways, tumor suppressor miRNAs and oncomiRs of32 HCC single cells.
Holo-Seq technology has high precision and low cost, and it has high feasibility and practicability. It also provides a safe and effective technical foundation for the establishment of quality control system for stem cell clinical research.
Ph.D. students Zhengyun Xiao and Guo Cheng in Prof.Jia’s group are the co-first authors. Dr.Chao Wu is the co-corresponding author of this article. This work was supported bythe Natural Science Foundation of Zhejiang Province, the special program for stem cells of the Ministry of Science and Technology, and the key specialty of precision medicine in the Ministry of Science and Technology, China.
Links:https://genomebiology.biomedcentral.com/articles/10.1186/s13059-018-1553-7
