Telomere heterogeneity linked to metabolism and pluripotency state revealed by simultaneous analysis of telomere length and RNA-seq in the same human embryonic stem cell

Abstract

BackgroundTelomere length heterogeneity has been detected in various cell types, including stem cells and cancer cells. Cell heterogeneity in pluripotent stem cells, such as embryonic stem cells (ESCs), is of particular interest; however, the implication and mechanisms underlying the heterogeneity remain to be understood. Single-cell analysis technology has recently been developed and effectively employed to investigate cell heterogeneity. Yet, methods that can simultaneously measure telomere length and analyze the global transcriptome in the same cell have not been available until now.ResultsWe have established a robust method that can simultaneously measure telomere length coupled with RNA-sequencing analysis (scT&R-seq) in the same human ESC (hESC). Using this method, we show that telomere length varies with pluripotency state. Compared to those with long telomere, hESCs with short telomeres exhibit the lowest expressions of TERF1/TRF1, and ZFP42/REX1, PRDM14 and NANOG markers for pluripotency, suggesting that these hESCs are prone to exit from the pluripotent state. Interestingly, hESCs ubiquitously express NOP10 and DKC1, stabilizing components of telomerase complexes. Moreover, new candidate genes, such as MELK, MSH6, and UBQLN1, are highly expressed in the cluster of cells with long telomeres and higher expression of known pluripotency markers. Notably, short telomere hESCs exhibit higher oxidative phosphorylation primed for lineage differentiation, whereas long telomere hESCs show elevated glycolysis, another key feature for pluripotency.ConclusionsTelomere length is a marker of the metabolic activity and pluripotency state of individual hESCs. Single cell analysis of telomeres and RNA-sequencing can be exploited to further understand the molecular mechanisms of telomere heterogeneity.Electronic supplementary materialThe online version of this article (doi:10.1186/s12915-017-0453-8) contains supplementary material, which is available to authorized users.

Teratogenic Rubella Virus Alters the Endodermal Differentiation Capacity of Human Induced Pluripotent Stem Cells

“…Additionally, RV infection did not alter stemness-related expression signatures as indicated by the transcriptomic activity of the GO gene set telomere maintenance (Figure 1F). Telomere maintenance is active in stem cells, but gets deactivated in differentiated somatic cells [24].…”

Role of CD133 in human embryonic stem cell proliferation and teratoma formation

“…WA26 and RuES2 human embryonic cells were routinely maintained in undifferentiated state in E8 medium (A1517001, Life technologies) on Matrigel (356230, BD Bioscience)-coated tissue culture plates with daily medium feeding and passaged every 3-4 days with 0.5 mM EDTA in phosphate buffered saline (PBS) with 10 μM Rocki (sc-281642A, Santa Cruz) for maintenance. Osteosarcoma cells (U2OS) (HTB96, ATCC), cervix adenocarcinoma cells (HeLa) (CCL-2, ATCC), and human embryonic fibroblast (HEF) cells (derived from an aborted fetus) [18,19] were cultured in high glucose Dulbecco's modified Eagle's media (DMEM) plus 10% fetal bovine serum (FBS) (SH30070.03, Hyclone) with 1% penicillin and streptomycin. Colon carcinoma cell line (HCT116) (CCL-247, ATCC) was cultured in RPMI1640 (11875085, Life technologies) plus 10% FBS and 1% penicillin and streptomycin.…”

“…Based on our single cell analysis, hESCs express CD133 at high levels [18]. To validate the high expression of CD133 in hESCs, we assessed CD133 expression by qPCR using two paired primers (different loci at the cDNA of PROM1) in six human cell lines, including two hESC lines (WA26 and RuES2), a colon adenocarcinoma cell line (HCT116), an osteosarcoma cell line (U2OS), a cervix adenocarcinoma cell line (HeLa), and a human embryonic fibroblast (HEF) cell line (Fig.…”

The role of telomere-binding modulators in pluripotent stem cells

“…Cells with longer telomeres may have higher pluripotency potential and be positively selected from a heterogeneous population. Singlecell telomere length analysis coupled with RNA sequencing (scT&R-seq) revealed that hESCs with short telomeres exhibited more characteristics associated with differentiated lineages, whereas those with longer telomeres better maintained pluripotency (Wang et al, 2017). Telomere length also plays an important role in stem cell differentiation.…”