Browsing by Author "Graham, Brett H."

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    Aberrant Function of the C-Terminal Tail of HIST1H1E Accelerates Cellular Senescence and Causes Premature Aging
    (Cell Press, 2019-09-05) Flex, Elisabetta; Martinelli, Simone; Van Dijck, Anke; Ciolfi, Andrea; Cecchetti, Serena; Coluzzi, Elisa; Pannone, Luca; Andreoli, Cristina; Radio, Francesca Clementina; Pizzi, Simone; Carpentieri, Giovanna; Bruselles, Alessandro; Catanzaro, Giuseppina; Pedace, Lucia; Miele, Evelina; Carcarino, Elena; Ge, Xiaoyan; Chijiwa, Chieko; Lewis, M.E. Suzanne; Meuwissen, Marije; Kenis, Sandra; Van der Aa, Nathalie; Larson, Austin; Brown, Kathleen; Wasserstein, Melissa P.; Skotko, Brian G.; Begtrup, Amber; Person, Richard; Karayiorgou, Maria; Roos, J. Louw; Van Gassen, Koen L.; Koopmans, Marije; Bijlsma, Emilia K.; Santen, Gijs W.E.; Barge-Schaapveld, Daniela Q.C.M.; Ruivenkamp, Claudia A.L.; Hoffer, Mariette J.V.; Lalani, Seema R.; Streff, Haley; Craigen, William J.; Graham, Brett H.; van den Elzen, Annette P.M.; Kamphuis, Daan J.; Ounap, Katrin; Reinson, Karit; Pajusalu, Sander; Wojcik, Monica H.; Viberti, Clara; Di Gaetano, Cornelia; Bertini, Enrico; Petrucci, Simona; De Luca, Alessandro; Rota, Rossella; Ferretti, Elisabetta; Matullo, Giuseppe; Dallapiccola, Bruno; Sgura, Antonella; Walkiewicz, Magdalena; Kooy, R. Frank; Tartaglia, Marco; Medical and Molecular Genetics, School of Medicine
    Histones mediate dynamic packaging of nuclear DNA in chromatin, a process that is precisely controlled to guarantee efficient compaction of the genome and proper chromosomal segregation during cell division and to accomplish DNA replication, transcription, and repair. Due to the important structural and regulatory roles played by histones, it is not surprising that histone functional dysregulation or aberrant levels of histones can have severe consequences for multiple cellular processes and ultimately might affect development or contribute to cell transformation. Recently, germline frameshift mutations involving the C-terminal tail of HIST1H1E, which is a widely expressed member of the linker histone family and facilitates higher-order chromatin folding, have been causally linked to an as-yet poorly defined syndrome that includes intellectual disability. We report that these mutations result in stable proteins that reside in the nucleus, bind to chromatin, disrupt proper compaction of DNA, and are associated with a specific methylation pattern. Cells expressing these mutant proteins have a dramatically reduced proliferation rate and competence, hardly enter into the S phase, and undergo accelerated senescence. Remarkably, clinical assessment of a relatively large cohort of subjects sharing these mutations revealed a premature aging phenotype as a previously unrecognized feature of the disorder. Our findings identify a direct link between aberrant chromatin remodeling, cellular senescence, and accelerated aging.
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    Association between pre-diagnostic leukocyte mitochondrial DNA copy number and survival among colorectal cancer patients
    (Elsevier, 2020-10) Yang, Keming; Forman, Michele R.; Graham, Brett H.; Monahan, Patrick O.; Giovannucci, Edward L.; De Vivo, Immaculata; Chan, Andrew T.; Nan, Hongmei; Epidemiology, School of Public Health
    Background Mitochondrial DNA copy number (mtDNAcn) is considered a biomarker for mitochondrial function and oxidative stress. Although previous studies have suggested a potential relationship between mtDNAcn at the time of colorectal cancer (CRC) diagnosis and CRC prognosis, findings have been inconsistent, and no study has specifically investigated the association of pre-diagnostic mtDNAcn with CRC survival. Methods We examined the association of pre-diagnostic leukocyte mtDNAcn (measured by qPCR) with overall and CRC-specific survival among 587 patients in Nurses’ Health Study and Health Professionals Follow-Up Study. Cox models were constructed to estimate hazard ratios (HRs) and 95 % confidence intervals (95 % CIs). Results During a mean follow-up of 10.5 years, 395 deaths were identified; 180 were due to CRC. Overall, we did not observe significant associations between mtDNAcn and either overall or CRC-specific survival among all cases or by cancer location, grade, or stage. In an exploratory stratified analysis, a suggestive inverse association of mtDNAcn and overall death risk appeared among current smokers [HR (95 % CI) for 1 SD decrease in mtDNAcn = 1.50 (0.98, 2.32), P-trend = 0.06]. Reduced mtDNAcn and lower CRC-specific death risk was observed among patients aged ≤ 70.5 at diagnosis [HR (95 % CI) for 1 SD decrease of mtDNAcn = 0.71 (0.52, 0.97), P-trend = 0.03], ≤ 5 years from blood collection to diagnosis [HR (95 % CI) for 1 SD decrease in mtDNAcn = 0.65 (0.44, 0.96), P-trend = 0.03] and those consuming a low-inflammatory diet [HR (95 % CI) for 1 SD decrease in mtDNAcn = 0.61 (0.42, 0.88), P-trend = 0.009]. Conclusion no significant associations between pre-diagnostic leukocyte mtDNAcn and either overall or CRC-specific survival appeared but exploratory analysis identified potential sub-group associations.
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    Biallelic variants in COX4I1 associated with a novel phenotype resembling Leigh syndrome with developmental regression, intellectual disability, and seizures
    (Wiley, 2019-10) Pillai, Nishitha R.; AlDhaheri, Noura S.; Gosh, Rajashri; Lim, Jaehyung; Streff, Haley; Nayak, Anuranjita; Graham, Brett H.; Hanchard, Neil; Elsea, Sarah H.; Scaglia, Fernando; Medical and Molecular Genetics, School of Medicine
    Autosomal recessive COX4I1 deficiency has been previously reported in a single individual with a homozygous pathogenic variant in COX4I1, who presented with short stature, poor weight gain, dysmorphic features, and features of Fanconi anemia. COX4I1 encodes subunit 4, isoform 1 of cytochrome c oxidase. Cytochrome c oxidase is a respiratory chain enzyme that plays an important role in mitochondrial electron transport and reduces molecular oxygen to water leading to the formation of ATP. Defective production of cytochrome c oxidase leads to a variable phenotypic spectrum ranging from isolated myopathy to Leigh syndrome. Here, we describe two siblings, born to consanguineous parents, who presented with encephalopathy, developmental regression, hypotonia, pathognomonic brain imaging findings resembling Leigh‐syndrome, and a novel homozygous variant on COX4I1, expanding the known clinical phenotype associated with pathogenic variants in COX4I1.
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    Characterization of a novel deep-intronic variant in DYNC2H1 identified by whole-exome sequencing in a patient with a lethal form of a short-rib thoracic dysplasia type III
    (Cold Spring Harbor Laboratory, 2022-12-28) Buchh, Muqsit; Gillespie, Patrick J.; Treat, Kayla; Abreu, Marco A.; Schwantes-An, Tae-Hwi Linus; Helm, Benjamin M.; Fang, Fang; Xuei, Xiaoling; Mantcheva, Lili; Suhrie, Kristen R.; Graham, Brett H.; Conboy, Erin; Vetrini, Francesco; Medical and Molecular Genetics, School of Medicine
    Biallelic pathogenic variants in DYNC2H1 are the cause of short-rib thoracic dysplasia type III with or without polydactyly (OMIM #613091), a skeletal ciliopathy characterized by thoracic hypoplasia due to short ribs. In this report, we review the case of a patient who was admitted to the Neonatal Intensive Care Unit (NICU) of Indiana University Health (IUH) for respiratory support after experiencing respiratory distress secondary to a small, narrow chest causing restrictive lung disease. Additional phenotypic features include postaxial polydactyly, short proximal long bones, and ambiguous genitalia were noted. Exome sequencing (ES) revealed a maternally inherited likely pathogenic variant c.10322C > T p.(Leu3448Pro) in the DYNC2H1 gene. However, there was no variant found on the paternal allele. Microarray analysis to detect deletion or duplication in DYNC2H1 was normal. Therefore, there was insufficient evidence to establish a molecular diagnosis. To further explore the data and perform additional investigations, the patient was subsequently enrolled in the Undiagnosed Rare Disease Clinic (URDC) at Indiana University School of Medicine (IUSM). The investigators at the URDC performed a reanalysis of the ES raw data, which revealed a paternally inherited DYNC2H1 deep-intronic variant c.10606-14A > G predicted to create a strong cryptic acceptor splice site. Additionally, the RNA sequencing of fibroblasts demonstrated partial intron retention predicted to cause a premature stop codon and nonsense-mediated mRNA decay (NMD). Droplet digital RT-PCR (RT-ddPCR) showed a drastic reduction by 74% of DYNCH2H1 mRNA levels. As a result, the intronic variant was subsequently reclassified as likely pathogenic resulting in a definitive clinical and genetic diagnosis for this patient. Reanalysis of ES and fibroblast mRNA experiments confirmed the pathogenicity of the splicing variants to supplement critical information not revealed in original ES or CMA reports. The NICU and URDC collaboration ended the diagnostic odyssey for this family; furthermore, its importance is emphasized by the possibility of prenatally diagnosing the mother's current pregnancy.
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    Deciphering the Role of Mitochondrial Dysfunction in Pulmonary Arterial Hypertension
    (2024-06) Balachandar, Srimmitha; Aldred, Micheala A.; Graham, Brett H.; Zhang, Jie; Geraci, Mark W.; Machado, Roberto F.
    Pulmonary arterial hypertension (PAH) is a life-threatening vasculopathy caused by remodeling of pulmonary arterioles. It is unknown as to why some people are at more risk of developing PAH compared to others. Notably, while germline pathogenic variants in PAH genes are a strong driver of disease susceptibility, less than half of mutation carriers actually develop the disease, suggesting the need for additional triggers. Our previous studies have shown increased DNA damage and total reactive oxygen species (ROS) in cells from PAH patients and unaffected relatives, indicating a potential genetic component, leading to our hypothesis: Mitochondrial dysfunction is an independent genetically determined modifier of PAH susceptibility. Untargeted metabolomics (Metabolon) revealed abnormalities in the antioxidants, glutamate, urea, amino acid, galactose, and phospholipid metabolism pathways in the PAH Lymphoblastoid cells (LCLs) compared to controls. Intriguingly, the healthy relatives also had altered phospholipids, suggesting that it occurs independent of the disease. ROS analysis on LCLs from patients, their relatives and unrelated controls showed that the PAH LCLs had significantly higher levels of all ROS species compared to controls, with the highest in heritable PAH cells. LCLs from relatives clustered into two groups, one with increased mitochondrial (mt) ROS and hydrogen peroxide, the other comparable to controls. Seahorse assays showed that the LCLs with increased mtROS had reduced spare respiratory capacity indicative of dysfunctional electron transport chain (ETC); but no glycolytic switch. Cybrid models generated using the high and low ROS LCLs (H and L-donors) on a 143B nuclear background showed that the H-donors had mt respiration similar to L-donors, suggesting a functional ETC. However, these cells had significantly elevated mtROS, with reduced SOD2 protein (potentially a consequence of increased degradation), passed on from the parental LCLs to the recipient cybrids. PAH is a complex disease, and mutation status alone doesn’t determine disease susceptibility. LCLs from patients recapitulate some of the metabolomic abnormalities in lung vascular cells. Oxidative stress in LCLs extends to some unaffected relatives, suggesting this is an independent genetic trait that modifies PAH risk. Our study highlights the importance of identifying potential modifiers and the second hits in the pathogenesis of PAH.
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    Impaired mitochondrial complex I function as a candidate driver in the biological stress response and a concomitant stress-induced brain metabolic reprogramming in male mice
    (Nature, 2020-06-01) Emmerzaal, Tim L.; Preston, Graeme; Geenen, Bram; Verweij, Vivienne; Wiesmann, Maximilian; Vasileiou, Elisavet; Grüter, Femke; de Groot, Corné; Schoorl, Jeroen; de Veer, Renske; Roelofs, Monica; Arts, Martijn; Hendriksen, Yara; Klimars, Eva; Donti, Taraka R.; Graham, Brett H.; Morava, Eva; Rodenburg, Richard J.; Kozicz, Tamas; Medical and Molecular Genetics, School of Medicine
    Mitochondria play a critical role in bioenergetics, enabling stress adaptation, and therefore, are central in biological stress responses and stress-related complex psychopathologies. To investigate the effect of mitochondrial dysfunction on the stress response and the impact on various biological domains linked to the pathobiology of depression, a novel mouse model was created. These mice harbor a gene trap in the first intron of the Ndufs4 gene (Ndufs4GT/GT mice), encoding the NDUFS4 protein, a structural component of complex I (CI), the first enzyme of the mitochondrial electron transport chain. We performed a comprehensive behavioral screening with a broad range of behavioral, physiological, and endocrine markers, high-resolution ex vivo brain imaging, brain immunohistochemistry, and multi-platform targeted mass spectrometry-based metabolomics. Ndufs4GT/GT mice presented with a 25% reduction of CI activity in the hippocampus, resulting in a relatively mild phenotype of reduced body weight, increased physical activity, decreased neurogenesis and neuroinflammation compared to WT littermates. Brain metabolite profiling revealed characteristic biosignatures discriminating Ndufs4GT/GT from WT mice. Specifically, we observed a reversed TCA cycle flux and rewiring of amino acid metabolism in the prefrontal cortex. Next, exposing mice to chronic variable stress (a model for depression-like behavior), we found that Ndufs4GT/GT mice showed altered stress response and coping strategies with a robust stress-associated reprogramming of amino acid metabolism. Our data suggest that impaired mitochondrial CI function is a candidate driver for altered stress reactivity and stress-induced brain metabolic reprogramming. These changes result in unique phenomic and metabolomic signatures distinguishing groups based on their mitochondrial genotype.
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    Increased COUP-TFII expression in adult hearts induces mitochondrial dysfunction resulting in heart failure
    (Springer Nature, 2015-09-10) Wu, San-Pin; Kao, Chung-Yang; Wang, Leiming; Creighton, Chad J.; Yang, Jin; Donti, Taraka R.; Harmancey, Romain; Vasquez, Hernan G.; Graham, Brett H.; Bellen, Hugo J.; Taegtmeyer, Heinrich; Chang, Ching-Pin; Tsai, Ming-Jer; Tsai, Sophia Y.; Department of Medicine, IU School of Medicine
    Mitochondrial dysfunction and metabolic remodelling are pivotal in the development of cardiomyopathy. Here, we show that myocardial COUP-TFII overexpression causes heart failure in mice, suggesting a causal effect of elevated COUP-TFII levels on development of dilated cardiomyopathy. COUP-TFII represses genes critical for mitochondrial electron transport chain enzyme activity, oxidative stress detoxification and mitochondrial dynamics, resulting in increased levels of reactive oxygen species and lower rates of oxygen consumption in mitochondria. COUP-TFII also suppresses the metabolic regulator PGC-1 network and decreases the expression of key glucose and lipid utilization genes, leading to a reduction in both glucose and oleate oxidation in the hearts. These data suggest that COUP-TFII affects mitochondrial function, impairs metabolic remodelling and has a key role in dilated cardiomyopathy. Last, COUP-TFII haploinsufficiency attenuates the progression of cardiac dilation and improves survival in a calcineurin transgenic mouse model, indicating that COUP-TFII may serve as a therapeutic target for the treatment of dilated cardiomyopathy.
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    Insulinemic Potential of Lifestyle Is Inversely Associated with Leukocyte Mitochondrial DNA Copy Number in US White Adults
    (Elsevier, 2020-08-01) Yang, Keming; Forman, Michele R.; Monahan, Patrick O.; Graham, Brett H.; Chan, Andrew T.; Zhang, Xuehong; De Vivo, Immaculata; Giovannucci, Edward L.; Tabung, Fred K.; Nan, Hongmei; Epidemiology, School of Public Health
    Background: Poor lifestyles have been linked to insulin insensitivity/hyperinsulinemia, which may contribute to downstream changes such as inflammation and oxidative damage and the development of chronic diseases. As a biomarker of intracellular oxidative stress, leukocyte mitochondrial DNA copy number (mtDNA-CN) has been related to lifestyle factors including diet and weight. No epidemiologic study has examined the relation between combined insulinemic potential of lifestyle and mtDNA-CN. Objectives: Our aim was to examine the association between Empirical Lifestyle Index for Hyperinsulinemia (ELIH) and leukocyte mtDNA-CN in US men and women. Methods: This cross-sectional analysis included 2835 white adults without cancers, diabetes, or cardiovascular disease at blood collection, including 2160 women from the Nurses' Health Study and 675 men from the Health Professionals Follow-Up Study. ELIH is an index based on plasma C-peptide that characterizes the insulinemic potential of lifestyle (diet, body weight, and physical activity). Relative mtDNA-CN in peripheral blood leukocytes was measured by qPCR-based assay. Results: We found a significant inverse association between ELIH and mtDNA-CN. In multivariable-adjusted linear models, absolute least squares means ± SDs of mtDNA-CN z score across ELIH quintiles in women were as follows: Q1: 0.14 ± 0.05; Q2: 0.04 ± 0.06; Q3: 0.008 ± 0.05; Q4: 0.01 ± 0.05; and Q5: -0.06 ± 0.05 (P-trend = 0.006). Means ± SDs in men were as follows: Q1: 0.25 ± 0.09; Q2: 0.23 ± 0.09; Q3: 0.07 ± 0.09; Q4: 0.02 ± 0.09; and Q5: -0.04 ± 0.09 (P-trend = 0.007). Means ± SDs in all participants were as follows: Q1: 0.16 ± 0.05; Q2: 0.07 ± 0.05; Q3: 0.01 ± 0.05; Q4: 0.01 ± 0.05; and Q5: -0.05 ± 0.05 (P-trend = 0.0004). Conclusions: Hyperinsulinemic lifestyles (i.e., higher ELIH) were associated with lower leukocyte mtDNA-CN among subjects without major diseases, suggesting that the difference in lifestyle insulinemic potential may be related to excessive oxidative stress damage.
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    Investigation into Tissue-Specific Mechanisms of Mitochondrial Dysfunction: Models of SUCLA2 Deficiency and a Screen for Potential Genetic Modifiers
    (2023-11) Lancaster, Makayla S.; Graham, Brett H.; Kim, Jungsu; Hoffman-Longtin, Krista; White, Kenneth E.
    With no currently effective treatments available, mitochondrial diseases are one of the most common forms of inherited multisystem disease. Primary disorders of the mitochondria affect an estimated 1 in 4,300 people with typical onset in early childhood. Mitochondrial disorders are classically defined by defects in the mitochondrial powerhouse, or respiratory chain (RC). Therefore, they are uniquely complex as the proteins within the RC are encoded by two separate genomes – nuclear DNA (nDNA) and mitochondrial DNA (mtDNA). The mitochondrial genome encodes 13 protein genes within the RC, with the remaining mitochondrial proteome being nuclear encoded. Therefore, mitochondrial disorders result from pathogenic variants within either genome. While mitochondrial disorders can affect multiple tissue symptoms, organs with high energy demand, such as the brain and skeletal muscle, are most typically affected; thus, mitochondrial disease typically manifests as an encephalomyopathy. A wide range of symptoms, including developmental delay, seizures, strokes, and sensorineural hearing loss have been associated with mitochondrial dysfunction. In short, however, investigation into the pathogenic mechanisms of mitochondrial disorders has proven difficult due to the wide clinical and genetic heterogeneity associated with the disorders. Therefore, this project seeks to investigate pathways of mitochondrial dysfunction using two genetic approaches. First, reverse genetics tools are used to generate tissue-specific mouse models of succinyl-CoA synthetase deficiency, which is a known cause of mitochondrial disease in humans. In parallel, forward genetics is used to screen for variation in mitochondrial phenotypes in a genetically diverse population of mice to identify potential genetic modifiers of mitochondrial function and health. Using both forward and reverse genetics approaches, these studies will allow for the investigation into tissue-specific mitochondrial pathogenesis in novel mouse models, as well as broadly characterize tissue-specific mitochondrial function in vivo. Taken together, both genetic approaches are used to broaden understanding of tissue-specific mitochondrial function in health and disease.
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    Lessons learned from additional research analyses of unsolved clinical exome cases
    (BioMed Central, 2017-03-21) Eldomery, Mohammad K.; Coban-Akdemir, Zeynep; Harel, Tamar; Rosenfeld, Jill A.; Gambin, Tomasz; Stray-Pedersen, Asbjørg; Küry, Sébastien; Mercier, Sandra; Lessel, Davor; Denecke, Jonas; Wiszniewski, Wojciech; Penney, Samantha; Liu, Pengfei; Bi, Weimin; Lalani, Seema R.; Schaaf, Christian P.; Wangler, Michael F.; Bacino, Carlos A.; Lewis, Richard Alan; Potocki, Lorraine; Graham, Brett H.; Belmont, John W.; Scaglia, Fernando; Orange, Jordan S.; Jhangiani, Shalini N.; Chiang, Theodore; Doddapaneni, Harsha; Hu, Jianhong; Muzny, Donna M.; Xia, Fan; Beaudet, Arthur L.; Boerwinkle, Eric; Eng, Christine M.; Plon, Sharon E.; Sutton, V. Reid; Gibbs, Richard A.; Posey, Jennifer E.; Yang, Yaping; Lupski, James R.; Department of Pathology and Laboratory Medicine, IU School of Medicine
    BACKGROUND: Given the rarity of most single-gene Mendelian disorders, concerted efforts of data exchange between clinical and scientific communities are critical to optimize molecular diagnosis and novel disease gene discovery. METHODS: We designed and implemented protocols for the study of cases for which a plausible molecular diagnosis was not achieved in a clinical genomics diagnostic laboratory (i.e. unsolved clinical exomes). Such cases were recruited to a research laboratory for further analyses, in order to potentially: (1) accelerate novel disease gene discovery; (2) increase the molecular diagnostic yield of whole exome sequencing (WES); and (3) gain insight into the genetic mechanisms of disease. Pilot project data included 74 families, consisting mostly of parent-offspring trios. Analyses performed on a research basis employed both WES from additional family members and complementary bioinformatics approaches and protocols. RESULTS: Analysis of all possible modes of Mendelian inheritance, focusing on both single nucleotide variants (SNV) and copy number variant (CNV) alleles, yielded a likely contributory variant in 36% (27/74) of cases. If one includes candidate genes with variants identified within a single family, a potential contributory variant was identified in a total of ~51% (38/74) of cases enrolled in this pilot study. The molecular diagnosis was achieved in 30/63 trios (47.6%). Besides this, the analysis workflow yielded evidence for pathogenic variants in disease-associated genes in 4/6 singleton cases (66.6%), 1/1 multiplex family involving three affected siblings, and 3/4 (75%) quartet families. Both the analytical pipeline and the collaborative efforts between the diagnostic and research laboratories provided insights that allowed recent disease gene discoveries (PURA, TANGO2, EMC1, GNB5, ATAD3A, and MIPEP) and increased the number of novel genes, defined in this study as genes identified in more than one family (DHX30 and EBF3). CONCLUSION: An efficient genomics pipeline in which clinical sequencing in a diagnostic laboratory is followed by the detailed reanalysis of unsolved cases in a research environment, supplemented with WES data from additional family members, and subject to adjuvant bioinformatics analyses including relaxed variant filtering parameters in informatics pipelines, can enhance the molecular diagnostic yield and provide mechanistic insights into Mendelian disorders. Implementing these approaches requires collaborative clinical molecular diagnostic and research efforts.