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Long Non-Coding RNAs Control Hematopoietic Stem Cell Function

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摘要 : Hematopoietic stem cells (HSCs) possess unique gene expression programs that enforce their identity and regulate lineage commitment. Long non-coding RNAs (lncRNAs) have emerged as important regulators of gene expression and cell fate decisions, although their functions in HSCs are unclear. Here we profiled the transcriptome of purified HSCs by deep sequencing and identified 323 unannotated lncRNAs. Comparing their expression in differentiated lineages revealed 159 lncRNAs enriched in HSCs, some
Hematopoietic stem cells (HSCs) possess unique gene expression programs that enforce their identity and regulate lineage commitment. Long non-coding RNAs (lncRNAs) have emerged as important regulators of gene expression and cell fate decisions, although their functions in HSCs are unclear. Here we profiled the transcriptome of purified HSCs by deep sequencing and identified 323 unannotated lncRNAs. Comparing their expression in differentiated lineages revealed 159 lncRNAs enriched in HSCs, some of which are likely HSC specific (LncHSCs). These lncRNA genes share epigenetic features with protein-coding genes, including regulated expression via DNA methylation, and knocking down two LncHSCs revealed distinct effects on HSC self-renewal and lineage commitment.We mapped the genomic binding sites of one of these candidates and found enrichment for key hematopoietic transcription factor binding sites, especially E2A. Together, these results demonstrate that lncRNAs play important roles in regulating HSCs, providing an additional layer to the genetic circuitry controlling HSC function.
Long Non-Coding RNAs Control Hematopoietic Stem Cell Function
Figure 5. ChIRP-Seq Reveals LncHSC-2 Binding Sites in the genome (A) LncHSC-2 binding sites are enriched in promoter-proximal regions. Left: the distribution of LncHSC-2 binding sites across the indicated intergenic or intragenic regions. Right: enrichment of LncHSC-2 sites (versus the genomic background) among transcript features. CDS, coding sequence. (B) Enriched sequence motif associated with lncHSC-2 binding sites (bottom) strongly reseMBLing the mouse Tcfe2a secondary motif (top). (C) Co-enrichment analysis of lncHSC-2 binding sites with sequence features, ChIP-seq profiles of hematopoietic transcriptional regulators and epigenetic marks in HSCs, multipotent progenitors (HPC-7 cells), bone marrow, thymus, and spleen. The enrichment for LncHSC-2 binding compared with the LacZ negative control was assessed by Fisher’s exact test with multiple testing correction. Colors subdivide the results into three classes: epigenetic mark (red), sequence feature (green), and TF binding site (blue). Dot sizes are proportionate to the odds ratio. The x axis values represent the –log Benjamini-Hochberg-corrected p value. (D) Hierarchical clustering of genomic regions bound by lncHSC-2 and published hematopoietic lineage TFs or epiGENEtic marks. The major partition of columns separates LncHSC-2 occupancy into two main branches, with unmethylated promoter-proximal regions associated with transcriptional activation marks (H3K4me3/H3K27ac/PolII) and Erg/Fli/Meis1/Pu.1 TFs to the left and promoter distal intronic or intergenic regions associated with bone marrow tissue enhancer or insulator elements (CTCF) E2A sequence motifs to the right. Each line corresponds to a LncHSC-2 peak, where blue/white coloring indicates the presence/absence of the additional given factors. (E) LncHSC-2 occupancy at the genes Pml (top) and Itpkb (bottom). Shown are LncHSC-2 and LacZ control ChIRP-seq signal density tracks generated by MACS2 representing the fragment pileup signal per million reads. Additional overlaid tracks are HSC H3K4me3, rna-seq, undermethylated regions (Jeong et al., 2014), and hematopoietic lineage TF binding sites (Wilson et al., 2010). (F) ChIP QPCR to show E2A binding to three LncHSC-2 binding peaks after LncHSC-2 KD in primary Sca-1+ cells. The y axis represents the percentage of immunoprecipitated DNA compared with the input. Mean ± SEM values are shown (n = 4). 原文链接:http://www.cell.com/cell-stem-cell/abstract/S1934-5909(15)00058-2 作者:广州赛诚生物 点击:
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