Wong JMY, Collins K.. Telomerase RNA level limits telomere maintenance in X-linked dyskeratosis congenita. Genes Dev 20: 2848-2858

Department of Molecular and Cell Biology, University of California at Berkeley, Berkeley, California 94720, USA.
Genes & Development (Impact Factor: 10.8). 11/2006; 20(20):2848-58. DOI: 10.1101/gad.1476206
Source: PubMed


Dyskeratosis congenita (DC) patients suffer a progressive and ultimately fatal loss of hematopoietic renewal correlating with critically short telomeres. The predominant X-linked form of DC results from substitutions in dyskerin, a protein required both for ribosomal RNA (rRNA) pseudouridine modification and for cellular accumulation of telomerase RNA (TER). Accordingly, alternative models have posited that the exhaustion of cellular renewal in X-linked DC arises as a primary consequence of ribosome deficiency or telomerase deficiency. Here we test, for the first time, whether X-linked DC patient cells are compromised for telomerase function at telomeres. We show that telomerase activation in family-matched control cells allows telomere elongation and telomere length maintenance, while telomerase activation in X-linked DC patient cells fails to prevent telomere erosion with proliferation. Furthermore, we demonstrate by phenotypic rescue that telomere defects in X-linked DC patient cells arise solely from reduced accumulation of TER. We also show that X-linked DC patient cells averted from premature senescence support normal levels of rRNA pseudouridine modification and normal kinetics of rRNA precursor processing, in contrast with phenotypes reported for a proposed mouse model of the human disease. These findings support the significance of telomerase deficiency in the pathology of X-linked DC.

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    • "Labeling of a tagged fusion protein by SFP synthase in vitro occurred with >80% efficiency (Yin et al., 2005). Human TERT tagged at the N-terminus supports telomere elongation, whereas telomerase assembled with C-terminally tagged TERT does not (Counter et al., 1998; Wong and Collins, 2006). "
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    ABSTRACT: Telomerase synthesizes chromosome-capping telomeric repeats using an active site in telomerase reverse transcriptase (TERT) and an integral RNA subunit template. The fundamental question of whether human telomerase catalytic activity requires cooperation across two TERT subunits remains under debate. In this study, we describe new approaches of subunit labeling for single-molecule imaging, applied to determine the TERT content of complexes assembled in cells or cell extract. Surprisingly, telomerase reconstitutions yielded heterogeneous DNA-bound TERT monomer and dimer complexes in relative amounts that varied with assembly and purification method. Among the complexes, cellular holoenzyme and minimal recombinant enzyme monomeric for TERT had catalytic activity. Dimerization was suppressed by removing a TERT domain linker with atypical sequence bias, which did not inhibit cellular or minimal enzyme assembly or activity. Overall, this work defines human telomerase DNA binding and synthesis properties at single-molecule level and establishes conserved telomerase subunit architecture from single-celled organisms to humans.
    Full-text · Article · Oct 2015 · eLife Sciences
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    • "Dyskerin and the other H/ACA snoRNP proteins are also required, in vertebrates, for the assembly and function of telomerase. Although no pseudouridylation defects or alterations in ribosome biogenesis or function have been shown in patients [6], mouse models with mutations in Dkc1 have decreased levels of pseudouridine in rRNA [7] [8] and a variety of other defects including changes in translational fidelity [2] [9] and perturbed ribosome biogenesis [7] [8]. In one case it was shown that in Dkc1 mutant mouse cells, pseudouridylation at a specific site was decreased and that rRNA in the cells, and in cells from a DC patient, had altered electrophoretic mobility under conditions of incomplete denaturation, suggesting changes in the stability or the secondary structure of rRNA [10]. "
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    ABSTRACT: Pseudouridine is the most abundant modified nucleotide in ribosomal RNA throughout eukaryotes and archaea but its role is not known. Here we produced mouse embryonic fibroblast cells expressing only catalytically inactive dyskerin, the pseudouridine synthase that converts uridine to pseudouridine in ribosomal RNA. The mutant dyskerin protein, D125A, was extremely unstable but cells were able to divide and grow very slowly. Abnormalities in ribosome RNA synthesis were apparent but mature cytoplasmic RNAs lacking pseudouridine were produced and were very unstable. We conclude that pseudouridine is required to stabilize the secondary structure of ribosomal RNA that is essential for its function.
    Preview · Article · May 2013 · FEBS letters
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    • "Despite telomerase is highly associated with cancer, inherited mutations in telomerase genes are known to be causative (as already described) of aging-related impairments, including DC. Mutations in DKC1 gene, for example, lead to lower levels of TERC in both humans and mice, resulting in premature telomere dysfunction in highly renewable tissues, as the bone marrow (Wong et al., 2004; Mochizuki et al., 2004; Wong and Collins, 2006). Telomerase activity is also important for telomere reprogramming during the early embryonic development (Wright et al., 1996, 2001; Xu and Yang, 2000), which is a fundamental event to generate young offspring and, consequently, for species maintenance along generations. "
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    ABSTRACT: Dyskeratosis congenita (DC) encompasses a large spectrum of diseases and clinical manifestations generally related to premature aging, including bone marrow failure and cancer predisposition. The major risk factor for DC is to carry germline telomere-related mutations-in telomerase or telomere shelterin genes-which results in premature telomere dysfunction, thus increasing the risk of premature aging impairments. Despite the advances that have been accomplished in DC research, the molecular aspects underlying the phenotypic variability of the disease remain poorly understood. Here different aspects of telomere biology, concerning adult stem cells senescence, tumor suppression and cancer are considered in the context of DC, resulting in two translational models: late onset of DC symptoms in telomere-related mutations carriers is a potential indicator of increased cancer risk and differences in tumor suppression capacities among the genetic subgroups are (at least partial) causes of different clinical manifestations of the disease. The limitations of both models are presented, and further experiments for their validation, as well as clinical implications, are discussed.
    Full-text · Article · Mar 2013 · Ageing research reviews
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