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.
Highlights
Telomere length heterogeneity has been detected in various cell types, including stem cells and cancer cells
Simultaneous measurement of telomere length and gene expression in the same cell To acquire the total mRNA and genomic DNA from the same cell, we used the biotinylated oligo-dT primer [35], targeting to mRNA containing a polyadenylated tail (poly (A)+) and applied a magnet to separate the gDNA from the same cell (Fig. 1a)
High expression levels of NOP10 and DKC1 may suggest their potential role in telomere maintenance of human ESC (hESC), presumably through the activities of the telomerase enzyme complex with the shelterin complex (TERF1, TRF2, TINF2, POT1, TPP1, and RAP1) [51]
Summary
Telomere length heterogeneity has been detected in various cell types, including stem cells and cancer cells. Functional telomeres are required for embryo development and for the pluripotency and differentiation capacity of embryonic stem cells (ESCs) [7, 8]. Pluripotent stem cells, including ESCs, induced pluripotent stem cells (iPSCs), and nuclear transfer ESCs with sufficient telomere lengths, are able to give rise to offspring shown by germline chimera production or tetraploid embryo complementation tests, the most stringent and functional test of naïve pluripotency [7, 9,10,11]. While short telomeres impair stem cell differentiation [12], ESCs with hyper-long telomeres may delay aging as evidenced by the generation of healthier chimera mice that exhibit reduced cell senescence and DNA damage with age and better skin wound healing [13]
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
