Rothmund-Thomson Syndrome-like RECQL4 truncating mutations cause a haploinsufficient low bone mass phenotype in mice.
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ABSTRACT:
SUBMITTER: Castillo-Tandazo W
PROVIDER: S-EPMC8088275 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:Mutations within the gene encoding the DNA helicase RECQL4 underlie the autosomal recessive cancer-predisposition disorder Rothmund-Thomson syndrome, though it is unclear how these mutations lead to disease. Here, we demonstrated that somatic deletion of Recql4 causes a rapid bone marrow failure in mice that involves cells from across the myeloid, lymphoid, and, most profoundly, erythroid lineages. Apoptosis was markedly elevated in multipotent progenitors lacking RECQL4 compared with WT cells. While the stem cell compartment was relatively spared in RECQL4-deficent mice, HSCs from these animals were not transplantable and even selected against. The requirement for RECQL4 was intrinsic in hematopoietic cells, and loss of RECQL4 in these cells was associated with increased replicative DNA damage and failed cell-cycle progression. Concurrent deletion of p53, which rescues loss of function in animals lacking the related helicase BLM, did not rescue BM phenotypes in RECQL4-deficient animals. In contrast, hematopoietic defects in cells from Recql4Δ/Δ mice were fully rescued by a RECQL4 variant without RecQ helicase activity, demonstrating that RECQL4 maintains hematopoiesis independently of helicase activity. Together, our data indicate that RECQL4 participates in DNA replication rather than genome stability and identify RECQL4 as a regulator of hematopoiesis with a nonredundant role compared with other RecQ helicases.
| S-EPMC4109528 | biostudies-literature
Project description:Rothmund-Thomson syndrome is a rare genodermatosis caused by biallelic mutations of the RECQL4 gene and is characterised by poikiloderma, sparse hair, eyelashes and/or eyebrows, small stature, skeletal and dental abnormalities and cancer predisposition. Mutations predicted to result in the loss of RECQL4 protein have been associated with osteosarcoma risk, but mutation(s)-phenotype correlations are better addressed by combined DNA and RNA analyses. We describe two siblings with a mild phenotype, mainly restricted to the skin, who carry the unreported paternal c.2272C>T alteration in exon 14 and the previously reported maternal exon 15 c.2492_2493delAT, both predicted to result in premature termination codons (p.(Arg758*), p.(His831Argfs*52)). However real-time and transcript analysis showed, in the carrier father and affected daughter, increased levels of a novel RECQL4 physiological alternative transcript with partial in-frame skipping of exon 14, generated by increased usage of a weak cryptic splice site. This alternative transcript is expressed in all controls and tested tissues, its upregulation is specific to the paternal c.2272C>T mutation and depends on the abrogation of the binding motifs for SF2 and SRp55 serine/arginine-rich proteins with bypass of the mutation site located in the skipped exon 14 portion. Moreover, in the proband the increased levels of the alternative transcript, likely encoding a protein isoform with residual activity, may compensate for the dearth of the canonical transcript with the c.2492_2493delAT, accounting for the mild clinical phenotype of the siblings. Our results emphasise the value of RNA analysis to better predict the effects of RECQL4 mutations on the clinical phenotype.
| S-EPMC4200430 | biostudies-literature
Project description:BackgroundPoikiloderma is defined as a chronic skin condition presenting with a combination of punctate atrophy, areas of depigmentation, hyperpigmentation and telangiectasia. In a variety of hereditary syndromes such as Rothmund-Thomson syndrome (RTS), Clericuzio-type poikiloderma with neutropenia (PN) and Dyskeratosis Congenita (DC), poikiloderma occurs as one of the main symptoms. Here, we report on genotype and phenotype data of a cohort of 44 index patients with RTS or related genodermatoses.MethodsDNA samples from 43 patients were screened for variants in the 21 exons of the RECQL4 gene using PCR, SSCP-PAGE analysis and/or Sanger sequencing. Patients with only one or no detectable mutation in the RECQL4 gene were additionally tested for variants in the 8 exons of the USB1 (C16orf57) gene by Sanger sequencing. The effect of novel variants was evaluated by phylogenic studies, single-nucleotide polymorphism (SNP) databases and in silico analyses.ResultsWe identified 23 different RECQL4 mutations including 10 novel and one homozygous novel USB1 (C16orf57) mutation in a patient with PN. Moreover, we describe 31 RECQL4 and 8 USB1 sequence variants, four of them being novel intronic RECQL4 sequence changes that may have some deleterious effects on splicing mechanisms and need further evaluation by transcript analyses.ConclusionThe current study contributes to the improvement of genetic diagnostic strategies and interpretation in RTS and PN that is relevant in order to assess the patients' cancer risk, to avoid continuous and inconclusive clinical evaluations and to clarify the recurrence risk in the families. Additionally, it shows that the phenotype of more than 50% of the patients with suspected Rothmund-Thomson disease may be due to mutations in other genes raising the need for further extended genetic analyses.
| S-EPMC4867568 | biostudies-literature
Project description:Telomeres are structures at the ends of chromosomes and are composed of long tracks of short tandem repeat DNA sequences bound by a unique set of proteins (shelterin). Telomeric DNA is believed to form G-quadruplex and D-loop structures, which presents a challenge to the DNA replication and repair machinery. Although the RecQ helicases WRN and BLM are implicated in the resolution of telomeric secondary structures, very little is known about RECQL4, the RecQ helicase mutated in Rothmund-Thomson syndrome (RTS). Here, we report that RTS patient cells have elevated levels of fragile telomeric ends and that RECQL4-depleted human cells accumulate fragile sites, sister chromosome exchanges, and double strand breaks at telomeric sites. Further, RECQL4 localizes to telomeres and associates with shelterin proteins TRF1 and TRF2. Using recombinant proteins we showed that RECQL4 resolves telomeric D-loop structures with the help of shelterin proteins TRF1, TRF2, and POT1. We also found a novel functional synergistic interaction of this protein with WRN during D-loop unwinding. These data implicate RECQL4 in telomere maintenance.
| S-EPMC3249070 | biostudies-literature
Project description:RecQ-like helicase 4 (RECQL4) is mutated in patients suffering from the Rothmund-Thomson syndrome, a genetic disease characterized by premature aging, skeletal malformations, and high cancer susceptibility. Known roles of RECQL4 in DNA replication and repair provide a possible explanation of chromosome instability observed in patient cells. Here, we demonstrate that RECQL4 is a microtubule-associated protein (MAP) localizing to the mitotic spindle. RECQL4 depletion in M-phase-arrested frog egg extracts does not affect spindle assembly per se, but interferes with maintaining chromosome alignment at the metaphase plate. Low doses of nocodazole depolymerize RECQL4-depleted spindles more easily, suggesting abnormal microtubule-kinetochore interaction. Surprisingly, inter-kinetochore distance of sister chromatids is larger in depleted extracts and patient fibroblasts. Consistent with a role to maintain stable chromosome alignment, RECQL4 down-regulation in HeLa cells causes chromosome misalignment and delays mitotic progression. Importantly, these chromosome alignment defects are independent from RECQL4's reported roles in DNA replication and damage repair. Our data elucidate a novel function of RECQL4 in mitosis, and defects in mitotic chromosome alignment might be a contributing factor for the Rothmund-Thomson syndrome.
| S-EPMC6362308 | biostudies-literature
Project description:Cellular senescence refers to irreversible growth arrest of primary eukaryotic cells, a process thought to contribute to aging-related degeneration and disease. Deficiency of RecQ helicase RECQL4 leads to Rothmund-Thomson syndrome (RTS), and we have investigated whether senescence is involved using cellular approaches and a mouse model. We first systematically investigated whether depletion of RECQL4 and the other four human RecQ helicases, BLM, WRN, RECQL1 and RECQL5, impacts the proliferative potential of human primary fibroblasts. BLM-, WRN- and RECQL4-depleted cells display increased staining of senescence-associated β-galactosidase (SA-β-gal), higher expression of p16(INK4a) or/and p21(WAF1) and accumulated persistent DNA damage foci. These features were less frequent in RECQL1- and RECQL5-depleted cells. We have mapped the region in RECQL4 that prevents cellular senescence to its N-terminal region and helicase domain. We further investigated senescence features in an RTS mouse model, Recql4-deficient mice (Recql4(HD)). Tail fibroblasts from Recql4(HD) showed increased SA-β-gal staining and increased DNA damage foci. We also identified sparser tail hair and fewer blood cells in Recql4(HD) mice accompanied with increased senescence in tail hair follicles and in bone marrow cells. In conclusion, dysfunction of RECQL4 increases DNA damage and triggers premature senescence in both human and mouse cells, which may contribute to symptoms in RTS patients.
| S-EPMC4047874 | biostudies-literature
Project description:RECQ helicase protein-like 4 (RECQL4) is a member of the human RECQ family of DNA helicases. Two-thirds of patients with Rothmund-Thomson syndrome (RTS) carry biallelic inactivating mutations in the RECQL4 gene. RTS is an autosomal recessive disorder characterized by poikiloderma, sparse hair, small stature, skeletal abnormalities, cataracts, and an increased risk of cancer. Mutations in two other RECQ helicases, BLM and WRN, are responsible for the cancer predisposition conditions Bloom and Werner syndromes, respectively. Previous studies have shown that BLM and WRN-deficient cells demonstrate increased sensitivity to hydroxyurea (HU), camptothecin (CPT), and 4-nitroquinoline 1-oxide (4NQO). Little is known about the sensitivity of RECQL4-deficient cells to these and other genotoxic agents. The purpose of this study was to determine if RTS cells display any distinct cellular phenotypes in response to DNA damaging agents or replication blocks that could provide insight into the molecular function of the RECQL4 protein. Our results show that primary fibroblasts from RTS patients carrying two deleterious RECQL4 mutations, compared to wild type (WT) fibroblasts, have increased sensitivity to HU, CPT, and doxorubicin (DOX), modest sensitivity to other DNA damaging agents including ultraviolet (UV) irradiation, ionizing radiation (IR), and cisplatin (CDDP), and relative resistance to 4NQO. The RECQ family of DNA helicases has been implicated in the regulation of DNA replication, recombination, and repair. Because HU, CPT, and DOX exert their effects primarily during S phase, these results support a greater role for the RECQL4 protein in DNA replication as opposed to repair of exogenous damage.
| S-EPMC2585174 | biostudies-literature
Project description:Rothmund-Thomson syndrome (RTS) is a genodermatosis presenting with a characteristic facial rash (poikiloderma) associated with short stature, sparse scalp hair, sparse or absent eyelashes and/or eyebrows, juvenile cataracts, skeletal abnormalities, radial ray defects, premature aging and a predisposition to cancer. The prevalence is unknown but around 300 cases have been reported in the literature so far. The diagnostic hallmark is facial erythema, which spreads to the extremities but spares the trunk, and which manifests itself within the first year and then develops into poikiloderma. Two clinical subforms of RTS have been defined: RTSI characterised by poikiloderma, ectodermal dysplasia and juvenile cataracts, and RTSII characterised by poikiloderma, congenital bone defects and an increased risk of osteosarcoma in childhood and skin cancer later in life. The skeletal abnormalities may be overt (frontal bossing, saddle nose and congenital radial ray defects), and/or subtle (visible only by radiographic analysis). Gastrointestinal, respiratory and haematological signs have been reported in a few patients. RTS is transmitted in an autosomal recessive manner and is genetically heterogeneous: RTSII is caused by homozygous or compound heterozygous mutations in the RECQL4 helicase gene (detected in 60-65% of RTS patients), whereas the aetiology in RTSI remains unknown. Diagnosis is based on clinical findings (primarily on the age of onset, spreading and appearance of the poikiloderma) and molecular analysis for RECQL4 mutations. Missense mutations are rare, while frameshift, nonsense mutations and splice-site mutations prevail. A fully informative test requires transcript analysis not to overlook intronic deletions causing missplicing. The diagnosis of RTS should be considered in all patients with osteosarcoma, particularly if associated with skin changes. The differential diagnosis should include other causes of childhood poikiloderma (including dyskeratosis congenita, Kindler syndrome and Poikiloderma with Neutropaenia), other rare genodermatoses with prominent telangiectasias (including Bloom syndrome, Werner syndrome and Ataxia-telangiectasia) and the allelic disorders, RAPADILINO syndrome and Baller-Gerold syndrome, which also share some clinical features. A few mutations recur in all three RECQL4 diseases. Genetic counselling should be provided for RTS patients and their families, together with a recommendation for cancer surveillance for all patients with RTSII. Patients should be managed by a multidisciplinary team and offered long term follow-up. Treatment includes the use of pulsed dye laser photocoagulation to improve the telangiectatic component of the rash, surgical removal of the cataracts and standard treatment for individuals who develop cancer. Although some clinical signs suggest precocious aging, life expectancy is not impaired in RTS patients if they do not develop cancer. Outcomes in patients with osteosarcoma are similar in RTS and non-RTS patients, with a five-year survival rate of 60-70%. The sensitivity of RTS cells to genotoxic agents exploiting cells with a known RECQL4 status is being elucidated and is aimed at optimizing the chemotherapeutic regimen for osteosarcoma.
| S-EPMC2826297 | biostudies-literature
Project description:Rothmund-Thomson syndrome (RTS) is a rare autosomal-recessive disorder with clinical features consisting of rash, poikiloderma, sparse hair, short stature, juvenile cataracts, skeletal abnormalities, and cancer predisposition. Genetic studies involving detection of pathogenic RECQL4 variants provide the diagnostic certitude. Osteosarcoma was found in two-thirds RECQL4-mutated RTS patients, while hematological malignancies were rarely reported. The variant diversity of RECQL4 gene has not been fully identified and mutations associated with hematologic malignancies are not well described. In this study, we presented a pedigree of RTS from a Chinese family, among which the proband was diagnosed with de novo myelodysplastic syndrome (MDS). Comprehensive medical examination and chromosome karyotyping were performed on the proband. Whole exome sequencing (WES) was performed on the proband, his sister and his mother. The familial cosegregation of sequence variants derived from WES was conducted by polymerase chain reaction-based Sanger sequencing. Structures of candidate RECQL4 mutants were done by in silico analysis to assess pathogenicity. Three novel RECQL4 germline variants, including c.T274C, c.G3014A, and c.G801C, were identified by WES and validated by Sanger sequencing. Prediction of conformation indicated that the structural stability of human RECQL4 protein was largely affected with these variants. The co-occurring U2AF1 p.S34F and TP53 p.Y220C mutations might contribute to the development of MDS. Our study expands the mutational spectrum of RECQL4 and provides underlying molecular mechanism for the development of MDS in RTS patients.
| S-EPMC10205365 | biostudies-literature
Project description:Rothmund-Thomson syndrome (RTS) is a rare autosomal recessive disorder characterized by poikiloderma, small stature, sparse hair, skeletal abnormalities, increased risk of osteosarcoma, and decreased bone mass. To date, there has not been a comprehensive evaluation of the prevalence and extent of metabolic bone disease in RTS. Furthermore, the mechanisms that result in this phenotype are largely unknown. In this report, we provide a detailed evaluation of 29 individuals with RTS with respect to their metabolic bone status including bone mineral density, calcium kinetics studies, and markers of bone remodeling. We show that individuals with RTS have decreased areal bone mineral density. Additionally, we demonstrate that the presence of pathogenic variants in RECQL4 and low bone mineral density correlate with the history of increased risk of fractures. Using a RECQL4-deficient mouse model that recapitulates skeletal abnormalities seen in individuals with RTS, we demonstrate that generalized skeletal involvement is likely due to decreased osteogenesis. Our findings are clinically relevant as they may help in the risk stratification of patients with RTS and also in the identification of individuals who may benefit from additional surveillance and management of metabolic bone disease.
| S-EPMC5886079 | biostudies-literature