RTEL1 knockout in human cells caused genome-wide DNA damage and compromised telomere elongation by telomerase

Eukaryotic chromosomes are capped by specialized structures composed of DNA repeats and associated proteins called telomeres. The telomeres distinguish the chromosome ends from broken ends. Telomeres in somatic cells gradually shorten with age and induce senescence and aging. In highly proliferative cells, telomeres are elongated by telomerase to extend their proliferation potential . Hoyeraal-Hreidarsson syndrome (HHS) is a fatal disease, characterized by severe telomere shortening, bone marrow failure and developmental defects. HHS is caused by germline mutations in telomerase subunits or accessory factors, and in the helicase RTEL1. RTEL1 was reported to have diverse genome-wide and telomeric functions. Human RTEL1 encodes for two main splice variants, RTEL1v1 RTEL1v2. To distinguish the different functions of RTEL1, we established RTEL1 knockout fibroblasts-like cells. These RTEL1^-/-, telomerase positive cells display severe telomere shortening and increased DNA damage. We introduced into these fibroblasts different variants and mutants of RTEL1 and examined the resulting phenotypes. Ectopic expression of WT RTELv2, but not RTEL1v1 or RTEL1v2 mutants, suppressed the genome-wide DNA damage. Furthermore, while the RTEL1^-/- cells did not display significant increase in DNA damage at telomeres, the expression of RTELv2 mutants or WT RTELv1 increased the telomeric damage, suggesting a dominant negative effect on telomeres. We did not observe telomere deletion in the RTEL1^-/- cells, but in vivo telomerase activity was dramatically reduced, while in vitro activity was normal. Altogether, these findings indicate that telomere shortening in RTEL1^-/- cells is mainly caused by inability of telomerase to elongate telomeres and not by telomere deletion.