177 research outputs found
DAG1 haploinsufficiency is associated with sporadic and familial isolated or pauci-symptomatic hyperCKemia
No full textDAG1 encodes for dystroglycan, a key component of the dystrophin-glycoprotein complex (DGC) with a pivotal role in skeletal muscle function and maintenance. Biallelic loss-of-function DAG1 variants cause severe muscular dystrophy and muscle-eye-brain disease. A possible contribution of DAG1 deficiency to milder muscular phenotypes has been suggested. We investigated the genetic background of twelve subjects with persistent mild-to-severe hyperCKemia to dissect the role of DAG1 in this condition. Genetic testing was performed through exome sequencing (ES) or custom NGS panels including various genes involved in a spectrum of muscular disorders. Histopathological and Western blot analyses were performed on muscle biopsy samples obtained from three patients. We identified seven novel heterozygous truncating variants in DAG1 segregating with isolated or pauci-symptomatic hyperCKemia in all families. The variants were rare and predicted to lead to nonsense-mediated mRNA decay or the formation of a truncated transcript. In four cases, DAG1 variants were inherited from similarly affected parents. Histopathological analysis revealed a decreased expression of dystroglycan subunits and Western blot confirmed a significantly reduced expression of beta-dystroglycan in muscle samples. This study supports the pathogenic role of DAG1 haploinsufficiency in isolated or pauci-symptomatic hyperCKemia, with implications for clinical management and genetic counseling
Using Genome Sequencing and Advanced Technologies to Diagnose Individuals with Rare Diseases
No full textAlthough individually rare, rare diseases (RDs) are estimated to impact up to 1 in 12 Canadians, and approximately 80% are estimated to be genetic. While the clinical adoption of exome sequencing has seen great success, over 50% of individuals with an RD remain undiagnosed after standard of care genetic testing. Short-read genome sequencing (GS) and additional technologies such as long-read whole-genome sequencing (LR-WGS) and RNA sequencing (RNAseq) offer improved detection of all forms of genetic variants across the genome, however they are currently primarily limited to translational research. The primary goal of this thesis was to develop and optimize an approach to using GS, and subsequent RNAseq and LR-WGS, to diagnose individuals with RDs. I specifically focused on the diagnosis of a cohort of 18 individuals for whom previous genetic testing identified a single heterozygous, known pathogenic variant in an autosomal recessive disease gene, felt to be a fit for their clinical presentation. Experimental validation and additional clinical testing were also performed when necessary to prove the pathogenicity of suspected disease causing variants. In this thesis, I discuss the successful diagnosis of seven individuals in our cohort. Chapters 3-6 describe how four of these molecular diagnoses were achieved and how they contribute to our understanding of disease mechanisms in the respective gene. These chapters specifically discuss SCLT1-related ciliopathies, SPG7 hereditary spastic paraplegia, ALG1 congenital disorder of glycosylation, and SLC39A8 congenital disorder of glycosylation. Chapter 7 outlines the larger results of our study and highlights the key strengths of our approach to using GS and other technologies specifically for providing molecular diagnoses for individuals with a previously identified heterozygous pathogenic variant in an AR disease gene. In Chapter 8, I outline the broader limitations of current approaches to the diagnosis of RDs and how the findings of this thesis have informed our holistic perspective towards using available technologies to improve diagnostic success
A diagnosis for all rare genetic diseases: the horizon and the next frontiers
No full textThe introduction of exome sequencing in the clinic has sparked tremendous optimism for the future of rare disease diagnosis, and there is exciting opportunity to further leverage these advances. To provide diagnostic clarity to all of these patients, however, there is a critical need for the field to develop and implement strategies to understand the mechanisms underlying all rare diseases and translate these to clinical care.</p
Clinical study of ferredoxin-reductase-related mitochondriopathy: Genotype-phenotype correlation and proposal of ancestry-based carrier screening in the Mexican population
No full textPurpose: Ferredoxin reductase (FDXR) is a flavoprotein that functions in both iron sulfur cluster biogenesis and steroid biosynthesis pathways in the mitochondria. Not surprisingly, loss of FDXR function causes severe mitochondrial diseases in humans. Although several FDXR-related mitochondriopathy (FRM) cohorts have been reported in the literature, further characterization of the natural history of FRM is warranted. Methods: To better understand the spectrum of FRM, a natural history study of FRM was performed. New cases were added to previously reported FRM cases for analysis (n = 62 cases). Results: Optic atrophy, movement disorder, and developmental delay were frequent findings. Mortality is high, with 18% of patients, often infants, passing from complications. Notably, 25% of cases were homozygous or compound heterozygous for the previously reported p.Arg386Trp “hotspot” variant. Of the obtained ancestry, all but 1 individual heterozygous for the p.Arg386Trp variant was Hispanic, with many reporting Mexican heritage. Utilizing recent large-scale genome sequencing surveys, the carrier frequency of the p.Arg386Trp variant was estimated as 1 of 185 in the Mexican population. Conclusion: Given the high mortality of FRM and carrier frequency of the common variant, consideration of a new approach for population carrier screening and development of therapeutics for affected individuals is needed
Investigating the role of splicing in disorders of craniofacial development
No full textFollowing clinical delineation of a rare disease, identification of the causative gene(s) is a crucial first step towards providing enhanced patient care and understanding disease aetiology. Disease gene discovery, especially when considered in the context of related disorders, can also provide new insight regarding normal development and physiology. Herein, I present two novel disease-causing genes: the ribosomal gene RPLP2 in Nager syndrome (NS) and the spliceosomal gene SNRPB in cerebro-costo-mandibular syndrome (CCMS). The mutations identified in SNRPB are the first example of de-regulated alternative splicing-coupled nonsense-mediated decay as a mechanism for human disease. NS and CCMS are both disorders of first and second pharyngeal arch development, with the jaw and ears being affected. Over the past five years, the association between disorders of craniofacial development and mutations in spliceosomal genes has become apparent with the discovery of SF3B4 in NS, EFTUD2 in mandibulofacial dysostosis type Guion-Almeida, and TXNL4A in Burn McKeown syndrome. The specificity of the phenotypes resulting from mutations in such ubiquitous genes has perplexed the scientific community, but it is at least clear that spliceosomal genes play a more prominent yet subtle role in development than previously thought. The association between Treacher Collins syndrome, which overlaps phenotypically with CCMS and NS, and the ribosomal genes TCOF1, POLR1C, and POLR1D has been known for longer. This work establishes NS as having both spliceosomal and ribosomal defects as a cause. In this thesis I discuss potential links between the mechanisms underlying ribosomal and spliceosomal defects in disorders of craniofacial development, particularly increased sensitivity to reactive oxygen species (ROS)
Using Next-Generation Sequencing to Identify Genes Mutated in Human Disorders
No full textNext-generation sequencing – and, in particular, exome sequencing, the targeted application of next-generation sequencing to the coding portion of the genome – appeared on the scene, in the last decade, as a promising new way of efficiently searching for and successfully identifying causative mutations in human disease. I have applied exome sequencing to the study of two human traits. The first is ROHHAD (Rapid-Onset Obesity with Hypoventilation, Hypothalamic Dysfunction and Autonomic Dysregulation), a rare, complex, and potentially fatal pediatric syndrome. The second is DTD (Developmental Topographical Disorientation), a novel behavioural phenotype in which individuals are unable to effectively navigate their surroundings. Though the two phenotypes themselves have little in common, they are united in the fact that both are believed to have a genetic basis, and are therefore candidates for the application of exome sequencing to search for causative mutations. The research described in this thesis has applied exome sequencing to both ROHHAD and DTD to search for causative mutations. In neither case were we able to identify causative genes, but in both cases we were able to rule out several genetic hypotheses (rare coding mutations across most of the genome), and, perhaps most importantly, we were able to rule out exome sequencing as a means of identifying disease-causing mutations for these traits. In addition to ruling out major hypotheses as to the underlying bases of these two traits, thus clearing the way for the exploration of additional hypotheses, the ROHHAD and DTD studies also provided informative illustrations of next-generation sequencing studies that do not identify causative genes. Together, they provided a framework for reporting negative results from next-generation sequencing studies, highlighting the importance of clearly reporting the data-quality, as well as ways to limit the number of false positive variant calls and considerations for the effective filtering of DNA sequence variants to ensure that the search for disease-causing candidates has been as thorough as possible
A rational approach to the child with mental retardation for the paediatrician
No full textMental Retardation (MR) is a problem encountered in almost all paediatric clinical settings. The assessment of a child with MR is a common diagnostic and management dilemma for paediatricians. The field of MR research is currently in a state of flux regarding not just our understanding of the condition, but also in the language and the processes we use in naming, defining and describing MR. This article will provide a better understanding and a rational approach toward MR. Prevalence rates for MR are variable in the literature and may be attributable to the variation in major classification systems and the diversity in study operation definitions and methodologies. Etiologies of MR are diverse and include many different influences. MR most often presents during infancy or preschool years as developmental delay. There is no universally accepted approach to the etiological work-up of mental retardation. The number of medical conditions associated with MR that are completely treatable by medical means remains small. The paediatrician plays a key role establishing short and long term treatment goals, as well as providing support to families who have children with MR
Institutional oblivion, social memory. A Tibilisi Printing House-Museum Case Study
No full textW roku 2003 Micheil Saakaszwili zaproponował Gruzinom nowy projekt tożsamościowy oparty na rekonstrukcji pamięci we wszelkich jej postaciach. Celem było odrzucenie pozytywnej pamięci o okresie komunizmu. Z przestrzeni publicznej
usunięto pomniki Lenina i Stalina, zmieniły się nazwy ulic. Odgórnie narzucone zmiany dotknęły instytucję pamięci – pojawiła się próba likwidacji muzeum Stalina w Gori, a w Tbilisi powstało Muzeum Sowieckiej Okupacji. Narzucone instytucjonalnie prze-pisanie pamięci nie w pełni obrazuje oddolny stan pamięci kulturowej i kolektywnej. Na przykładzie muzeum przy tbiliskiej ulicy Kaspi upamiętniającego nielegalną podziemną komunistyczną drukarnię działającą w latach 1903–1906, autor przedstawia narracyjne zderzenie między
instytucjonalnie narzuconym politycznym zapomnieniem oraz społecznym zapotrzebowaniem.
Antropologicznie badane są polityczne i dyskursywne uwarunkowania rekonstrukcji pamięci instytucjonalnej w kontekście zagadnienia tożsamości.In 2003, Micheil Saakaszwili suggested to Georgians a new identity project based on reconstructing memory in all its forms. The aim was the reject the positive memory of the communist period. The statues of Lenin and Stalin were removed from the public space, the names of the streets were changed. The governmentsteered transformation touched the institutional memory – there was an attempt at removing the Stalin museum in Gori, and in Tibilisi was created the Museum of Soviet Occupation. The rewriting of memory, institutionally enforced, does not fully reflect the grassroots state of cultural and collective memory. Taking as an example the Tibilisi museum at Kaspi street, which commemorates an illegal underground
printing house of communists in the years 1903–1906, the author shows the narrative clash between the institutionally enforced political oblivion and the social need. Political and discursive conditions of reconstructing institutional memory in the context of identity concept are subject to anthropological
analysis
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