Dyskeratosis congenita (DC) is a multisystem bone marrow failure syndrome characterized by a triad of mucocutaneous abnormalities and a predisposition to cancer. The genetic basis of DC remains unknown in more than 60% of patients. Mutations have been identified in components of the telomerase complex (dyskerin, TERC, TERT, NOP10, and NHP2), and recently in one component of the shelterin complex TIN2 (gene TINF2). To establish the role of TINF2 mutations, we screened DNA from 175 uncharacterised patients with DC as well as 244 patients with other bone marrow failure disorders. Heterozygous coding mutations were found in 33 of 175 previously uncharacterized DC index patients and 3 of 244 other patients. A total of 21 of the mutations affected amino acid 282, changing arginine to histidine (n = 14) or cysteine (n = 7). A total of 32 of 33 patients with DC with TINF2 mutations have severe disease, with most developing aplastic anaemia by the age of 10 years. Telomere lengths in patients with TINF2 mutations were the shortest compared with other DC subtypes, but TERC levels were normal. In this large series, TINF2 mutations account for approximately 11% of all DC, but they do not play a significant role in patients with related disorders. This study emphasises the role of defective telomere maintenance on human disease.

Dyskeratosis congenita (DC) is characterized by multiple features including mucocutaneous abnormalities, bone marrow failure and an increased predisposition to cancer. It exhibits marked clinical and genetic heterogeneity. DKC1 encoding dyskerin, a component of H/ACA small nucleolar ribonucleoprotein (snoRNP) particles is mutated in X-linked recessive DC. Telomerase RNA component (TERC), the RNA component and TERT the enzymatic component of telomerase, are mutated in autosomal dominant DC, suggesting that DC is primarily a disease of defective telomere maintenance. The gene(s) involved in autosomal recessive DC remains elusive. This paper describes studies aimed at defining the genetic basis of AR-DC. Homozygosity mapping in 16 consanguineous families with 25 affected individuals demonstrates that there is no single genetic locus for AR-DC. However, we show that NOP10, a component of H/ACA snoRNP complexes including telomerase is mutated in a large consanguineous family with classical DC. Affected homozygous individuals have significant telomere shortening and reduced TERC levels. While a reduction of TERC levels is not a universal feature of DC, it can be brought about through a reduction of NOP10 transcripts, as demonstrated by siRNA interference studies. A similar reduction in TERC levels is also seen when the mutant NOP10 is expressed in HeLa cells. These findings identify the genetic basis of one subtype of AR-DC being due to the first documented mutations in NOP10. This further strengthens the model that defective telomere maintenance is the primary pathology in DC and substantiates the evidence in humans for the involvement of NOP10 in the telomerase complex and telomere maintenance.

Dyskeratosis congenita is a premature aging syndrome characterized by muco-cutaneous features and a range of other abnormalities, including early greying, dental loss, osteoporosis, and malignancy. Dyskeratosis congenita cells age prematurely and have very short telomeres. Patients have mutations in genes that encode components of the telomerase complex (dyskerin, TERC, TERT, and NOP10), important in the maintenance of telomeres. Many dyskeratosis congenita patients remain uncharacterized. Here, we describe the analysis of two other proteins, NHP2 and GAR1, that together with dyskerin and NOP10 are key components of telomerase and small nucleolar ribonucleoprotein (snoRNP) complexes. We have identified previously uncharacterized NHP2 mutations that can cause autosomal recessive dyskeratosis congenita but have not found any GAR1 mutations. Patients with NHP2 mutations, in common with patients bearing dyskerin and NOP10 mutations had short telomeres and low TERC levels. SiRNAmediated knockdown of NHP2 in human cells led to low TERC levels, but this reduction was not observed after GAR1 knockdown. These findings suggest that, in human cells, GAR1 has a different impact on the accumulation of TERC compared with dyskerin, NOP10, and NHP2. Most of the mutations so far identified in patients with classical dyskeratosis congenita impact either directly or indirectly on the stability of RNAs. In keeping with this effect, patients with dyskerin, NOP10, and now NHP2 mutations have all been shown to have low levels of telomerase RNA in their peripheral blood, providing direct evidence of their role in telomere maintenance in humans.GAR1 ͉ bone marrow failure ͉ telomeres

Dyskeratosis congenita (DC) is a multisystem bone marrow failure syndrome characterized by a triad of mucocutaneous abnormalities and an increased predisposition to malignancy. X-linked DC is due to mutations in DKC1, while heterozygous mutations in TERC (telomerase RNA component) and TERT (telomerase reverse transcriptase) have been found in autosomal dominant DC. Many patients with DC remain uncharacterized, particularly families displaying autosomal recessive (AR) inheritance. We have now identified novel homozygous TERT mutations in 2 unrelated consanguineous families, where the index cases presented with classical DC or the more severe variant, Hoyeraal-Hreidarsson (HH) syndrome. These TERT mutations resulted in reduced telomerase activity and extremely short telomeres. As these mutations are homozygous, these patients are predicted to have significantly reduced telomerase activity in vivo. Interestingly, in contrast to patients with heterozygous TERT mutations or hemizygous DKC1 mutations, these 2 homozygous TERT patients were observed to have higherthan-expected TERC levels compared with controls. Collectively, the findings from this study demonstrate that homozygous TERT mutations, resulting in a pure but severe telomerase deficiency, produce a phenotype of classical AR-DC and its severe variant, the HH syndrome. IntroductionDyskeratosis congenita (DC) is a genetically and clinically heterogeneous inherited bone marrow (BM) failure syndrome that is classically characterized by a mucocutaneous triad of abnormal skin pigmentation, nail dystrophy, and leukoplakia, as well as a wide range of other cutaneous abnormalities. 1 The X-linked recessive form of DC was linked to mutations in DKC1, which encodes the highly conserved multifunctional protein dyskerin. [2][3][4][5][6][7] Dyskerin associates with a subclass of small nucleolar RNAs (snoRNAs) through the RNAs H/ACA domain. [8][9][10] These 2 molecules ultimately form a ribonucleoprotein (RNP) complex with the additional proteins GAR1, NHP2, and NOP10. [11][12][13] This complex has at least 2 significant biologic functions. First, it is responsible for the pseudouridylation of ribosomal RNA at residues specified by the snoRNA contained within the RNP. 14 Second, in addition to these pseudouridylation functions, the vertebrate version of this RNP complex associates with a reverse transcriptase (RT) to form the telomerase complex. [15][16][17] Telomeres are unique tandem repeat structures located at the ends of linear eukaryotic chromosomes. 18 Their presence serves many functions including prevention of chromosome end-to-end fusion, maintenance of chromosomal stability, and prevention of chromosomal degradation. 19 Telomerase is a specialized RNP complex that elongates the G-rich telomeric repeats. 20,21 It is proposed that telomerase acts after DNA replication where this specialized enzyme extends the leading DNA strand, reestablishing the 3Ј overhang that is lost during normal cell division due to end processing and the "end replication problem." 22,23 Th...

Dyskeratosis congenita (DC) is an inherited poikiloderma which in addition to the skin abnormalities is typically associated with nail dystrophy, leucoplakia, bone marrow failure, cancer predisposition and other features. Approximately 50% of DC patients remain genetically uncharacterized. All the DC genes identified to date are important in telomere maintenance. To determine the genetic basis of the remaining cases of DC, we undertook linkage analysis in 20 families and identified a common candidate gene region on chromosome 16 in a subset of these. This region included the C16orf57 gene recently identified to be mutated in poikiloderma with neutropenia (PN), an inherited poikiloderma displaying significant clinical overlap with DC. Analysis of the C16orf57 gene in our uncharacterized DC patients revealed homozygous mutations in 6 of 132 families. In addition, three of six families previously classified as Rothmund–Thomson syndrome (RTS—a poikiloderma that is sometimes confused with PN) were also found to have homozygous C16orf57 mutations. Given the role of the previous DC genes in telomere maintenance, telomere length was analysed in these patients and found to be comparable to age-matched controls. These findings suggest that mutations in C16orf57 unify a distinct set of families which clinically can be categorized as DC, PN or RTS. This study also highlights the multi-system nature (wider than just poikiloderma and neutropenia) of the clinical features of affected individuals (and therefore house-keeping function of C16orf57), a possible role for C16orf57 in apoptosis, as well as a distinct difference from previously characterized DC patients because telomere length was normal.