Feeder Cells Regulate Telomere Homeostasis And Pluripotency Maintenance Of Mouse Embryonic Stem Cells

Feeder cells like mouse embryonic fibroblasts(MEFs)have been widely applied for culture of pluripotent stem cells(PSCs),but their roles remain elusive.Noticeably,mouse embryonic stem cells(ESCs)cultured on the feeders display transcriptional heterogeneity.Telomeres are regions of repetitive nucleotide sequences at the end of chromosomes,which maintain genomic stability with associated proteins and critical for naive state maintenance of mouse PSCs.We re-visited roles of feeder cells by examining the telomere homeostasis maintenance.Here we show that telomere is longer in mESCs cultured with than without the feeders.ESC cultures without MEF feeders exhibit telomere loss,chromosomal fusion and aneuploidy with increasing passages.Notably,feeders facilitate heterogeneous transcription of 2 cell-genes including Zscan4 and telomere elongation independent of telomerase.Consistently,frequency of telomere sister chromatin exchange(T-SCE)is increased in feeder-based ESCs.Moreover,feeders produce Fstl1 that together with BMP4 periodically activate Zscan4.Epigenetically,feeders repress the expression of Dnmt3 b.Feeder-free cultures also have been explored to sustain self-renewal of ESCs.Remarkably,N2B27 medium with 2i(inhibitors of Mek and Gsk3? signaling)in the absence of feeders was developed to achieve ground state of mouse ESCs.Interestingly,Zscan4 is significantly repressed in mESCs cultured in 2i media without feeders,associated with shorter telomeres and increased chromosome instability.These data suggest that MEF feeders can promote the appropriate expression of Zscan4 and other 2 cell-genes,maintain telomere lengths and genomic stability,and thus play an important role in maintaining long-term stable self-renewal of naive mESCs.