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Telomere position effect in embryonic stem cells : heterogeneity, imprinting, and modifier screen Szeto, Rochelle Justina

Abstract

Telomeres are repetitive sequences found at the end of linear chromosomes. Their main function is to protect chromosome ends from degradation and to ensure proper DNA replication. Telomere position effect (TPE) refers to the epigenetic phenomenon of a gene being stochastically active or silenced as a consequence of its proximity to telomeric heterochromatin. TPE is a subset of position effect variegation (PEV), which also involves variable transcriptional silencing due to the proximity to centromeric heterochromatin or transposable elements. We have observed a TPE-like effect on GFP expression from the DelTel7 allele in embryonic stem cell (ESC) lines. The DelTel7 allele is an engineered chromosome truncation carrying a GFP reporter next to an array of telomere repeats. The truncation breakpoint is in the middle of a large cluster of imprinted genes found on distal mouse chromosome 7. Our results suggest that the GFP reporter is regulated by TPE in undifferentiated DelTel7/+ ESCs. The studies described in this thesis first addressed the relationship between GFP heterogeneity seen in DelTel7/+ ESCs and the previously described phenotypic heterogeneity existing in undifferentiated ESCs grown in serum. TPE was found to be active in all ESCs grown in serum, irrespective of their state. Growth in KSR+2i serum-free medium, which forces ESCs into a naїve state, up regulates the promoter driving GFP expression, at both the DelTel7 allele and at an interstitial control transgene. My results show that, as previously described in a yeast model of TPE, the silencing imposed by proximity to the telomeres can be at least partially overcome by increased transcription. A parent-of-origin effect at the GFP reporter of the DelTel7 allele was revealed in vivo, but TPE does not spread to nearby imprinted genes in ESCs. Finally, while working on the development of an episomal system to screen for modifiers of TPE, I noted that prolonged zeocin exposure has drastic effects on TPE. My results suggest a previously unappreciated relationship between DSB repair and TPE.

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