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Single cell lncRNAs in hematopoiesis
tein-coding gene pairs (see Online Supplementary Results “Characterization of lncRNAs defined in human CD34+ hematopoietic cells”; Online Supplementary Figure S3)].
Detection of long noncoding RNAs with single cell RNA-sequencing
Expression of lncRNAs showed more variation among single cells than did the expression of coding transcripts (Figure 2A). Across all percentiles of gene expression lev- els, lncRNAs were expressed in smaller proportions of cells than were mRNAs (Figure 2B). Low overall expres-
sion of lncRNAs in bulk samples was likely partly attrib- utable to limited but high expression of lncRNAs in a minority of cells or in small cell populations. Seven bulk samples of the CD34+ population from the nine individu- als studied were sequenced in parallel with single cells. We sought to compare the maximum abundance of mRNAs or lncRNAs versus housekeeping genes in bulk samples and individual cells,28 to quantify the power of gene expression detection by these different technical approaches. mRNAs were detected at a similar ratio to housekeeping genes in both bulk samples and single cells,
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D
Figure 2. Detection of long noncoding RNAs by single cell RNA sequencing. (A) Variance of long noncoding RNA (lncRNA) and messenger RNA (mRNA) expression among single cells. x axis, Log (TPM+1); y axis, variance. (B) Proportion of CD34+ cells (individual dots) that express individual lncRNAs (blue) and mRNAs (red), sep- arated by expression quantile of the set of all transcripts (lncRNAs and mRNAs combined). x axis, average expression level quantiles; y axis, proportion of cells. (C) Comparison of single cell and bulk tissue maximum expression levels of mRNAs and lncRNAs. Gray, housekeeping genes; green, mRNAs; red, lncRNAs. Projected density plots summarized expression levels of scatter plots along the single-cell (horizontal) and bulk tissue (vertical) axes. Short lines noted alongside the histogram plots represent the difference of the median expression of lncRNAs or mRNAs to the median expression of housekeeping genes in single cell or bulk tissue RNA-seq. (D) Gene-ontology semantic similarity matrix of protein-coding genes defined by a guilt-by-association approach of lncRNAs in human CD34+ cells. Gene ontology terms involved in a similar functional matrix were adjacent and formed a block with Pearson R values ranging from -1 to 1. Terms noted on the right side depict com- mon biological processes of the block of gene-ontology terms.
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