Corces, R. M. | bioRxiv Science

bioRxiv2021

Clonal inactivation of telomerase promotes accelerated stem cell differentiation

Telomerase is intimately associated with stem cells and upregulated in cancer, where it serves essential roles through its catalytic action in elongating telomeres, nucleoprotein caps that protect chromosome ends1. Overexpression of the telomerase reverse transcriptase (TERT) enhances cell proliferation through telomere-independent means, yet definitive evidence for such a direct role in stem cell function has yet to be revealed through loss-of-function studies. Here, we show that conditional deletion of TERT in spermatogonial stem cells (SSCs) markedly impairs competitive clone formation. Using lineage-tracing from the Tert locus, we find that TERT-expressing SSCs yield long-lived clones, but that selective TERT-inactivation in SSCs causes accelerated stem cell differentiation thereby disrupting clone formation. This requirement for TERT in clone formation is bypassed by expression of a catalytically inactive TERT transgene and occurs independently of the canonical telomerase complex. TERT inactivation induces a genome-wide reduction in open chromatin evident in purified SSCs, but not in committed progenitor cells. Loss of TERT causes reduced activity of the MYC oncogene and transgenic expression of MYC in TERT-deleted SSCs efficiently rescues clone formation. These data reveal a required catalytic activity-independent role for TERT in preventing stem cell differentiation, forge a genetic link between TERT and MYC and suggest new means by which TERT may promote tumorigenesis.

developmental biology↗

bioRxiv2020

Polycomb-mediated Genome Architecture Enables Long-range Spreading of H3K27 methylation

Polycomb-group proteins play critical roles in gene silencing through the deposition of histone H3 lysine 27 trimethylation (H3K27me3) and chromatin compaction1-5. This process is essential for embryonic stem cell (ESCs) pluripotency, differentiation, and development. Polycomb repressive complex 2 (PRC2) can both read and write H3K27me3, enabling progressive spread of H3K27me3 on the linear genome6. Long-range Polycomb-associated DNA contacts have also been described, but their regulation and role in gene silencing remains unclear7-10. Here, we apply H3K27me3 HiChIP11-13, a protein-directed chromosome conformation method, and optical reconstruction of chromatin architecture14 to profile long-range Polycomb-associated DNA loops that span tens to hundreds of megabases across multiple topological associated domains in mouse ESCs and human induced pluripotent stem cells7-10. We find that H3K27me3 loop anchors are enriched for Polycomb nucleation points and coincide with key developmental genes, such as Hmx1, Wnt6 and Hoxa. Genetic deletion of H3K27me3 loop anchors revealed a coupling of Polycomb-associated genome architecture and H3K27me3 deposition evidenced by disruption of spatial contact between distant loci and altered H3K27me3 in cis, both locally and megabases away on the same chromosome. Further, we find that global alterations in PRC2 occupancy resulting from an EZH2 mutant15 selectively deficient in RNA binding is accompanied by loss of Polycomb-associated DNA looping. Together, these results suggest PRC2 acts as a "genomic wormhole", using RNA binding to enhance long range chromosome folding and H3K27me3 spreading. Additionally, developmental gene loci have novel roles in Polycomb spreading, emerging as important architectural elements of the epigenome.

genetics↗