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Item Metadata only The impact of the cytoplasmic ubiquitin ligase TNFAIP3 gene variation on transcription factor NF-κB activation in acute kidney injury(Elsevier BV, 2023) Rogers, N.M.; Zammit, N.; Nguyen-Ngo, D.; Souilmi, Y.; Minhas, N.; Meijles, D.N.; Self, E.; Walters, S.N.; Warren, J.; Cultrone, D.; El-Rashid, M.; Li, J.; Chtanova, T.; O'Connell, P.J.; Grey, S.T.Nuclear factor kB (NF-kB) activation is a deleterious molecular mechanism that drives acute kidney injury (AKI) and manifests in transplanted kidneys as delayed graft function. The TNFAIP3 gene encodes A20, a cytoplasmic ubiquitin ligase and a master negative regulator of the NF- kB signaling pathway. Common population-specific TNFAIP3 coding variants that reduce A20’s enzyme function and increase NF- kB activation have been linked to heightened protective immunity and autoimmune disease, but have not been investigated in AKI. Here, we functionally identified a series of unique human TNFAIP3 coding variants linked to the autoimmune genome-wide association studies single nucleotide polymorphisms of F127C; namely F127C;R22Q, F127C;G281E, F127C;W448C and F127C;N449K that reduce A20’s anti-inflammatory function in an NF- kB reporter assay. To investigate the impact of TNFAIP3 hypomorphic coding variants in AKI we tested a mouse Tnfaip3 hypomorph in a model of ischemia reperfusion injury (IRI). The mouse Tnfaip3 coding variant I325N increases NF- kB activation without overt inflammatory disease, providing an immune boost as I325N mice exhibit enhanced innate immunity to a bacterial challenge. Surprisingly, despite exhibiting increased intra-kidney NF- kB activation with inflammation in IRI, the kidney of I325N mice was protected. The I325N variant influenced the outcome of IRI by changing the dynamic expression of multiple cytoprotective mechanisms, particularly by increasing NF- kB-dependent anti-apoptotic factors BCL-2, BCL-XL, c-FLIP and A20, altering the active redox state of the kidney with a reduction of superoxide levels and the enzyme super oxide dismutase-1, and enhancing cellular protective mechanisms including increased Foxp3D T cells. Thus, TNFAIP3 gene variants represent a kidney and population-specific molecular factor that can dictate the course of IRI.Item Metadata only Australian Genomics: Outcomes of a 5-year national program to accelerate the integration of genomics in healthcare(Elsevier (Cell Press), 2023) Stark, Z.; Boughtwood, T.; Haas, M.; Braithwaite, J.; Gaff, C.L.; Goranitis, I.; Spurdle, A.B.; Hansen, D.P.; Hofmann, O.; Laing, N.; Metcalfe, S.; Newson, A.J.; Scott, H.S.; Thorne, N.; Ward, R.L.; Dinger, M.E.; Best, S.; Long, J.C.; Grimmond, S.M.; Pearson, J.; et al.Australian Genomics is a national collaborative partnership of more than 100 organizations piloting a whole-of-system approach to integrating genomics into healthcare, based on federation principles. In the first five years of operation, Australian Genomics has evaluated the outcomes of genomic testing in more than 5,200 individuals across 19 rare disease and cancer flagship studies. Comprehensive analyses of the health economic, policy, ethical, legal, implementation and workforce implications of incorporating genomics in the Australian context have informed evidence-based change in policy and practice, resulting in national government funding and equity of access for a range of genomic tests. Simultaneously, Australian Genomics has built national skills, infrastructure, policy, and data resources to enable effective data sharing to drive discovery research and support improvements in clinical genomic delivery.Item Open Access Introme accurately predicts the impact of coding and noncoding variants on gene splicing, with clinical applications(Springer Nature, 2023) Sullivan, P.J.; Gayevskiy, V.; Davis, R.L.; Wong, M.; Mayoh, C.; Mallawaarachchi, A.; Hort, Y.; McCabe, M.J.; Beecroft, S.; Jackson, M.R.; Arts, P.; Dubowsky, A.; Laing, N.; Dinger, M.E.; Scott, H.S.; Oates, E.; Pinese, M.; Cowley, M.J.Predicting the impact of coding and noncoding variants on splicing is challenging, particularly in non-canonical splice sites, leading to missed diagnoses in patients. Existing splice prediction tools are complementary but knowing which to use for each splicing context remains difficult. Here, we describe Introme, which uses machine learning to integrate predictions from several splice detection tools, additional splicing rules, and gene architecture features to comprehensively evaluate the likelihood of a variant impacting splicing. Through extensive benchmarking across 21,000 splice-altering variants, Introme outperformed all tools (auPRC: 0.98) for the detection of clinically significant splice variants. Introme is available at https://github.com/CCICB/introme .Item Open Access General Practitioners perspectives on infant telomere length screening after a pregnancy complication: a qualitative analysis(Oxford University Press (OUP), 2024) Puglisi, C.J.; McDonough, J.; Bianco-Miotto, T.; A Grieger, J.Background: Pregnancy complications can impact the mother and child’s health in the short and longterm resulting in an increased risk of chronic disease later in life. Telomere length is a biomarker of future cardiometabolic diseases and may offer a novel way of identifying offspring most at risk for future chronic diseases. Objective(s): To qualitatively explore General Practitioners’ (GPs) perspectives on the feasibility and uptake for recommending a telomere screening test in children who were born after a pregnancy complication. Methods:Twelve semi-structured interviews were conducted with GPs within metropolitan Adelaide, South Australia. Interviews were audio recorded, transcribed verbatim, and analysed for codes and themes. Results: Two themes were generated: ethical considerations and practical considerations. Ethically, the GP participants discussed barriers including consenting on behalf of a child, parental guilt, and the impact of health insurance, whereas viewing it for health promotion was a facilitator. For practical considerations, barriers included the difficulty in identifying people eligible for screening, maintaining medical communication between service providers, and time and financial constraints, whereas linking screening for telomere length with existing screening would facilitate uptake. Conclusions: GPs were generally supportive of potential telomere screening in infants, particularly via a saliva test that could be embedded in current antenatal care. However, several challenges, such as lack of knowledge, ethical considerations, and time and financial constraints, need to be overcome before such a test could be implemented into practice.Item Open Access Temporal Exposure to Bt Insecticide Causes Oxidative Stress in Larval Midgut Tissue(MDPI AG, 2023) Muita, B.K.; Baxter, S.W.Bacillus thuringiensis (Bt) three-domain Cry toxins are highly successful biological pesticides; however, the mechanism through which they cause death to targeted larval midgut cells is not fully understood. Herein, we challenged transgenic Bt-susceptible Drosophila melanogaster larvae with moderate doses of activated Cry1Ac toxin and assessed the midgut tissues after one, three, and five hours using transmission electron microscopy and transcriptome sequencing. Larvae treated with Cry1Ac showed dramatic changes to their midgut morphology, including shortened microvilli, enlarged vacuoles, thickened peritrophic membranes, and swelling of the basal labyrinth, suggesting water influx. Transcriptome analysis showed that innate immune responses were repressed, genes involved with cell death pathways were largely unchanged, and mitochondria-related genes were strongly upregulated following toxin exposure. Defective mitochondria produced after toxin exposure were likely to contribute to significant levels of oxidative stress, which represent a common physiological response to a range of toxic chemicals. Significant reductions in both mitochondrial aconitase activity and ATP levels in the midgut tissue supported a rapid increase in reactive oxygen species (ROS) following exposure to Cry1Ac. Overall, these findings support the role of water influx, midgut cell swelling, and ROS activity in response to moderate concentrations of Cry1Ac.Item Metadata only Heritable defects in telomere and mitotic function selectively predispose to sarcomas(American Association for the Advancement of Science (AAAS), 2023) Ballinger, M.L.; Pattnaik, S.; Mundra, P.A.; Zaheed, M.; Rath, E.; Priestley, P.; Baber, J.; Ray-Coquard, I.; Isambert, N.; Causeret, S.; van der Graaf, W.T.A.; Puri, A.; Duffaud, F.; Le Cesne, A.; Seddon, B.; Chandrasekar, C.; Schiffman, J.D.; Brohl, A.S.; James, P.A.; Kurtz, J.-E.; et al.Cancer genetics has to date focused on epithelial malignancies, identifying multiple histotype-specific pathways underlying cancer susceptibility. Sarcomas are rare malignancies predominantly derived from embryonic mesoderm. To identify pathways specific to mesenchymal cancers, we performed whole-genome germline sequencing on 1644 sporadic cases and 3205 matched healthy elderly controls. Using an extreme phenotype design, a combined rare-variant burden and ontologic analysis identified two sarcoma-specific pathways involved in mitotic and telomere functions. Variants in centrosome genes are linked to malignant peripheral nerve sheath and gastrointestinal stromal tumors, whereas heritable defects in the shelterin complex link susceptibility to sarcoma, melanoma, and thyroid cancers. These studies indicate a specific role for heritable defects in mitotic and telomere biology in risk of sarcomas.Item Metadata only A custom hybridisation enrichment forensic intelligence panel to infer biogeographic ancestry, hair and eye colour, and Y chromosome lineage(Elsevier, 2022) Bardan, F.; Higgins, D.; Austin, J.J.Massively parallel sequencing can provide genetic data for hundreds to thousands of loci in a single assay for various types of forensic testing. However, available commercial kits require an initial PCR amplification of short-to-medium sized targets which limits their application for highly degraded DNA. Development and optimisation of large PCR multiplexes also prevents creation of custom panels that target different suites of markers for identity, biogeographic ancestry, phenotype, and lineage markers (Y-chromosome and mtDNA). Hybridisation enrichment, an alternative approach for target enrichment prior to sequencing, uses biotinylated probes to bind to target DNA and has proven successful on degraded and ancient DNA. We developed a customisable hybridisation capture method, that uses individually mixed baits to allow tailored and targeted enrichment to specific forensic questions of interest. To allow collection of forensic intelligence data, we assembled and tested a custom panel of hybridisation baits to infer biogeographic ancestry, hair and eye colour, and paternal lineage (and sex) on modern male and female samples with a range of self-declared ancestries and hair/eye colour combinations. The panel correctly estimated biogeographic ancestry in 9/12 samples (75%) but detected European admixture in three individuals from regions with admixed demographic history. Hair and eye colour were predicted correctly in 83% and 92% of samples respectively, where intermediate eye colour and blond hair were problematic to predict. Analysis of Y-chromosome SNPs correctly assigned sex and paternal haplogroups, the latter complementing and supporting biogeographic ancestry predictions. Overall, we demonstrate the utility of this hybridisation enrichment approach to forensic intelligence testing using a combined suite of biogeographic ancestry, phenotype, and Y-chromosome SNPs for comprehensive biological profiling.Item Open Access Human Genetic Research in Wallacea and Sahul: Recent Findings and Future Prospects(MDPI AG, 2023) Taufik, L.; Teixeira, J.C.; Llamas, B.; Sudoyo, H.; Tobler, R.; Purnomo, G.A.Genomic sequence data from worldwide human populations have provided a range of novel insights into our shared ancestry and the historical migrations that have shaped our global genetic diversity. However, a comprehensive understanding of these fundamental questions has been impeded by the lack of inclusion of many Indigenous populations in genomic surveys, including those from the Wallacean archipelago (which comprises islands of present-day Indonesia located east and west of Wallace’s and Lydekker’s Lines, respectively) and the former continent of Sahul (which once combined New Guinea and Australia during lower sea levels in the Pleistocene). Notably, these regions have been important areas of human evolution throughout the Late Pleistocene, as documented by diverse fossil and archaeological records which attest to the regional presence of multiple hominin species prior to the arrival of anatomically modern human (AMH) migrants. In this review, we collate and discuss key findings from the past decade of population genetic and phylogeographic literature focussed on the hominin history in Wallacea and Sahul. Specifically, we examine the evidence for the timing and direction of the ancient AMH migratory movements and subsequent hominin mixing events, emphasising several novel but consistent results that have important implications for addressing these questions. Finally, we suggest potentially lucrative directions for future genetic research in this key region of human evolution.Item Open Access BAF complex-mediated chromatin relaxation is required for establishment of X chromosome inactivation(Springer Nature, 2022) Keniry, A.; Jansz, N.; Gearing, L.J.; Wanigasuriya, I.; Chen, J.; Nefzger, C.M.; Hickey, P.F.; Gouil, Q.; Liu, J.; Breslin, K.A.; Iminitoff, M.; Beck, T.; Tapia Del Fierro, A.; Whitehead, L.; Jarratt, A.; Kinkel, S.A.; Taberlay, P.C.; Willson, T.; Pakusch, M.; Ritchie, M.E.; et al.The process of epigenetic silencing, while fundamentally important, is not yet completely understood. Here we report a replenishable female mouse embryonic stem cell (mESC) system, Xmas, that allows rapid assessment of X chromosome inactivation (XCI), the epigenetic silencing mechanism of one of the two X chromosomes that enables dosage compensation in female mammals. Through a targeted genetic screen in differentiating Xmas mESCs, we reveal that the BAF complex is required to create nucleosome-depleted regions at promoters on the inactive X chromosome during the earliest stages of establishment of XCI. Without this action gene silencing fails. Xmas mESCs provide a tractable model for screen-based approaches that enable the discovery of unknown facets of the female-specific process of XCI and epigenetic silencing more broadly.Item Open Access Antisense oligonucleotide therapy for KCNT1 encephalopathy(American Society for Clinical Investigation (ASCI), 2022) Burbano, L.E.; Li, M.; Jancovski, N.; Jafar-Nejad, P.; Richards, K.; Sedo, A.; Soriano, A.; Rollo, B.; Jia, L.; Gazina, E.V.; Piltz, S.; Adikusuma, F.; Thomas, P.Q.; Kopsidas, H.; Rigo, F.; Reid, C.A.; Maljevic, S.; Petrou, S.Developmental and epileptic encephalopathies (DEE) are characterized by pharmacoresistant seizures with concomitant intellectual disability. Epilepsy of infancy with migrating focal seizures (EIMFS) is one of the most severe of these syndromes. De novo variants in ion channels, including gain-of-function variants in KCNT1, have been found to play a major role in the etiology of EIMFS. Here, we test a potential precision therapeutic approach in KCNT1-associated DEE using a gene silencing antisense oligonucleotide (ASO) approach. We generated a mouse model carrying the KCNT1 p.P924L pathogenic variant; only the homozygous animals presented with the frequent, debilitating seizures and developmental compromise that are seen in patients. After a single intracerebroventricular bolus injection of a Kcnt1 gapmer ASO in symptomatic mice at postnatal day 40, seizure frequency was significantly reduced, behavioral abnormalities improved, and overall survival was extended compared to mice treated with a control ASO (non-hybridizing sequence). ASO administration at neonatal age was also well-tolerated and effective in controlling seizures and extending the lifespan of treated animals. The data presented here provide proof of concept for ASO-based gene silencing as a promising therapeutic approach in KCNT1-associated epilepsies.Item Open Access A nearly complete skeleton of a new eusphenodontian from the Upper Jurassic Morrison Formation, Wyoming, USA, provides insight into the evolution and diversity of Rhynchocephalia (Reptilia: Lepidosauria)(Informa UK Limited, 2022) DeMar, D.G.; Jones, M.E.H.; Carrano, M.T.We describe a new, small-bodied rhynchocephalian reptile, Opisthiamimus gregori gen. et sp. nov., from the Upper Jurassic Morrison Formation of Wyoming, USA. Whereas many fossil rhynchocephalians are based on isolated incomplete jaws, the holotype of O. gregori includes most of the skull and postcranium and therefore represents one of the most complete specimens of Rhynchocephalia known from North America. We used micro-computed tomography to examine its skeletal anatomy in detail and to develop a three-dimensional reconstruction of the skull. The skull of O. gregori is similar to that of several non-neosphenodontian rhynchocephalians such as Planocephalosaurus (e.g. large orbits) and Clevosaurus (e.g. parietal parasagittal crests) yet exhibits a suite of other features related to the proal shearing mechanism that becomes increasingly elaborated among more phylogenetically nested taxa such as Sphenodon (e.g. lateral palatine tooth row parallels maxillary tooth row along its entire length, pyramidal dentary teeth with mesial shearing crests). The postcranial skeleton of O. gregori exhibits characteristics typical of a terrestrial rhynchocephalian. Our phylogenetic analyses use a substantially updated data set of 118 characters and 46 taxa, and both maximum parsimony and Bayesian frameworks. Results place O. gregori inside Eusphenodontia but outside Neosphenodontia, and therefore in a key position for contributing to character polarity for more deeply nested clades such as Clevosauridae, Sphenodontidae and Pleurosauridae. We also erect Leptorhynchia taxon nov., composed primarily of aquatically adapted taxa (e.g. Pleurosaurus, Sapheosaurus), which is supported by both cranial and postcranial characters. Because O. gregori is not particularly closely related to the other named Morrison rhynchocephalians (e.g. Opisthias rarus), it increases both the alpha and beta taxonomic diversities within the formation. Similarly, major differences in body size and inferred diet of the Morrison taxa imply considerable concomitant palaeoecological diversity just prior to a major global decline in rhynchocephalian diversity around the close of the Jurassic.Item Open Access CRISPR applications for Duchenne muscular dystrophy: From animal models to potential therapies(Wiley, 2023) Chey, Y.C.J.; Arudkumar, J.; Aartsma‐Rus, A.; Adikusuma, F.; Thomas, P.Q.CRISPR gene-editing technology creates precise and permanent modifications to DNA. It has significantly advanced our ability to generate animal disease models for use in biomedical research and also has potential to revolutionize the treatment of genetic disorders. Duchenne muscular dystrophy (DMD) is a monogenic muscle-wasting disease that could potentially benefit from the development of CRISPR therapy. It is commonly associated with mutations that disrupt the reading frame of the DMD gene that encodes dystrophin, an essential scaffolding protein that stabilizes striated muscles and protects them from contractile-induced damage. CRISPR enables the rapid generation of various animal models harboring mutations that closely simulates the wide variety of mutations observed in DMD patients. These models provide a platform for the testing of sequence-specific interventions like CRISPR therapy that aim to reframe or skip DMD mutations to restore functional dystrophin expression.Item Open Access The biomechanical role of the chondrocranium and the material properties of cartilage(Senckenberg Gesellschaft für Naturforschung, 2020) Jones, M.E.H.; Gröning, F.; Aspden, R.M.; Dutel, H.; Sharp, A.; Moazen, M.; Fagan, M.J.; Evans, S.E.The chondrocranium is the cartilage component of the vertebrate braincase. Among jawed vertebrates it varies greatly in structure, mineralisation, and in the extent to which it is replaced by bone during development. In mammals, birds, and some bony fsh, most of the chondrocranium is replaced by bone whereas in lizards, amphibians, and chondrichthyan fsh it may remain a signifcant part of the braincase complex in adulthood. To what extent this variation relates to differences in skull biomechanics is poorly understood. However, there have been examinations of chondrocranium histology, in vivo strain, and impact on rostrum growth following partial removal of the chondrocranium. These studies have led to suggestions that the chondrocranium may provide structural support or serve to dampen external loads. Advances in computing-power have also facilitated an increase in the number of three-dimensional computer-based models. These models can be analysed (in silico) to test specifc biomechanical hypotheses under specifed loading conditions. However, representing the material properties of cartilage is still problematic because these properties differ according to the speed and direction of loading. The relationship between stress and strain is also non-linear. Nevertheless, analyses to date suggest that the chondrocranium does not provide a vertical support in lizards but it may serve to absorb some loads in humans. We anticipate that future models will include ever more detailed representations of the loading, anatomy, and material properties, in tandem with rigorous forms of model validation. However, comparison among a wider range of vertebrate subjects should also be pursued, in particular larvae, juveniles, and very small adult animals.Item Open Access The Nestin neural enhancer is essential for normal levels of endogenous Nestin in neuroprogenitors but is not required for embryo development(Public Library of Science, 2021) Thomson, E.; Dawson, R.; H'ng, C.H.; Adikusuma, F.; Piltz, S.; Thomas, P.Q.; Varshney, G.Enhancers are vitally important during embryonic development to control the spatial and temporal expression of genes. Recently, large scale genome projects have identified a vast number of putative developmental regulatory elements. However, the proportion of these that have been functionally assessed is relatively low. While enhancers have traditionally been studied using reporter assays, this approach does not characterise their contribution to endogenous gene expression. We have studied the murine Nestin (Nes) intron 2 enhancer, which is widely used to direct exogenous gene expression within neural progenitor cells in cultured cells and in vivo. We generated CRISPR deletions of the enhancer region in mice and assessed their impact on Nes expression during embryonic development. Loss of the Nes neural enhancer significantly reduced Nes expression in the developing CNS by as much as 82%. By assessing NES protein localization, we also show that this enhancer region contains repressor element(s) that inhibit Nes expression within the vasculature. Previous reports have stated that Nes is an essential gene, and its loss causes embryonic lethality. We also generated 2 independent Nes null lines and show that both develop without any obvious phenotypic effects. Finally, through crossing of null and enhancer deletion mice we provide evidence of trans-chromosomal interaction of the Nes enhancer and promoter.Item Open Access Brain transcriptomes of zebrafish and mouse Alzheimer's disease knock-in models imply early disrupted energy metabolism(Company of Biologists, 2022) Barthelson, K.; Newman, M.; Lardelli, M.Energy production is the most fundamentally important cellular activity supporting all other functions, particularly in highly active organs such as brains, Here we summarise transcriptome analyses of young adult (pre-disease) brains from a collection of eleven early-onset familial Alzheimer’s disease (EOfAD)-like and non-EOfAD-like mutations in three zebrafish genes. The one cellular activity consistently predicted as affected by only the EOfAD-like mutations is oxidative phosphorylation that produces most of the brain’s energy. All the mutations were predicted to affect protein synthesis. We extended our analysis to knock-in mouse models of APOE alleles and found the same effect for the late onset Alzheimer’s disease risk allele 4. Our results support a common molecular basis for initiation of the pathological processes leading to both early and late onset forms of Alzheimer’s disease and illustrate the utility of zebrafish and of knock-in, single EOfAD mutation models for understanding the causes of this disease.Item Open Access Microchromosomes are building blocks of bird, reptile, and mammal chromosomes(Proceedings of the National Academy of Sciences, 2021) Waters, P.D.; Patel, H.R.; Ruiz-Herrera, A.; Álvarez-González, L.; Lister, N.C.; Simakov, O.; Ezaz, T.; Kaur, P.; Frere, C.; Grutzner, F.; Georges, A.; Graves, J.A.M.Microchromosomes, once considered unimportant shreds of the chicken genome, are gene-rich elements with a high GC content and few transposable elements. Their origin has been debated for decades. We used cytological and whole-genome sequence comparisons, and chromosome conformation capture, to trace their origin and fate in genomes of reptiles, birds, and mammals. We find that microchromosomes as well as macrochromosomes are highly conserved across birds and share synteny with single small chromosomes of the chordate amphioxus, attesting to their origin as elements of an ancient animal genome. Turtles and squamates (snakes and lizards) share different subsets of ancestral microchromosomes, having independently lost microchromosomes by fusion with other microchromosomes or macrochromosomes. Patterns of fusions were quite different in different lineages. Cytological observations show that microchromosomes in all lineages are spatially separated into a central compartment at interphase and during mitosis and meiosis. This reflects higher interaction between microchromosomes than with macrochromosomes, as observed by chromosome conformation capture, and suggests some functional coherence. In highly rearranged genomes fused microchromosomes retain most ancestral characteristics, but these may erode over evolutionary time; surprisingly, de novo microchromosomes have rapidly adopted high interaction. Some chromosomes of early-branching monotreme mammals align to several bird microchromosomes, suggesting multiple microchromosome fusions in a mammalian ancestor. Subsequently, multiple rearrangements fueled the extraordinary karyotypic diversity of therian mammals. Thus, microchromosomes, far from being aberrant genetic elements, represent fundamental building blocks of amniote chromosomes, and it is mammals, rather than reptiles and birds, that are atypical.Item Metadata only Mapping complex traits in a diversity outbred f1 mouse population identifies germline modifiers of metastasis in human prostate cancer(Elsevier, 2017) Winter, J.; Gildea, D.; Andreas, J.; Gatti, D.; Williams, K.; Lee, M.; Hu, Y.; Zhang, S.; Mullikin, J.; Wolfsberg, T.; McDonnell, S.; Fogarty, Z.; Larson, M.; French, A.; Schaid, D.; Thibodeau, S.; Churchill, G.; Crawford, N.It is unclear how standing genetic variation affects the prognosis of prostate cancer patients. To provide one controlled answer to this problem, we crossed a dominant, penetrant mouse model of prostate cancer to Diversity Outbred mice, a collection of animals that carries over 40 million SNPs. Integration of disease phenotype and SNP variation data in 493 F1 males identified a metastasis modifier locus on Chromosome 8 (LOD = 8.42); further analysis identified the genes Rwdd4, Cenpu, and Casp3 as functional effectors of this locus. Accordingly, analysis of over 5,300 prostate cancer patient samples revealed correlations between the presence of genetic variants at these loci, their expression levels, cancer aggressiveness, and patient survival. We also observed that ectopic overexpression of RWDD4 and CENPU increased the aggressiveness of two human prostate cancer cell lines. In aggregate, our approach demonstrates how well-characterized genetic variation in mice can be harnessed in conjunction with systems genetics approaches to identify and characterize germline modifiers of human disease processes.Item Metadata only A predictive computational framework for direct reprogramming between human cell types(NATURE PUBLISHING GROUP, 2016) Rackham, O.J.L.; Firas, J.; Fang, H.; Oates, M.E.; Holmes, M.L.; Knaupp, A.S.; Suzuki, H.; Nefzger, C.M.; Daub, C.O.; Shin, J.W.; Petretto, E.; Forrest, A.R.R.; Hayashizaki, Y.; Polo, J.M.; Gough, J.Transdifferentiation, the process of converting from one cell type to another without going through a pluripotent state, has great promise for regenerative medicine. The identification of key transcription factors for reprogramming is currently limited by the cost of exhaustive experimental testing of plausible sets of factors, an approach that is inefficient and unscalable. Here we present a predictive system (Mogrify) that combines gene expression data with regulatory network information to predict the reprogramming factors necessary to induce cell conversion. We have applied Mogrify to 173 human cell types and 134 tissues, defining an atlas of cellular reprogramming. Mogrify correctly predicts the transcription factors used in known transdifferentiations. Furthermore, we validated two new transdifferentiations predicted by Mogrify. We provide a practical and efficient mechanism for systematically implementing novel cell conversions, facilitating the generalization of reprogramming of human cells. Predictions are made available to help rapidly further the field of cell conversion.Item Metadata only DNA hypermethylation encroachment at CpG island borders in cancer is predisposed by H3K4 monomethylation patterns(Elsevier BV, 2019) Skvortsova, K.; Masle-Farquhar, E.; Luu, P.L.; Song, J.Z.; Qu, W.; Zotenko, E.; Gould, C.M.; Du, Q.; Peters, T.J.; Colino-Sanguino, Y.; Pidsley, R.; Nair, S.S.; Khoury, A.; Smith, G.C.; Miosge, L.A.; Reed, J.H.; Kench, J.G.; Rubin, M.A.; Horvath, L.; Bogdanovic, O.; et al.Promoter CpG islands are typically unmethylated in normal cells, but in cancer a proportion are subject to hypermethylation. Using methylome sequencing we identified CpG islands that display partial methylation encroachment across the 5' or 3' CpG island borders. CpG island methylation encroachment is widespread in prostate and breast cancer and commonly associates with gene suppression. We show that the pattern of H3K4me1 at CpG island borders in normal cells predicts the different modes of cancer CpG island hypermethylation. Notably, genetic manipulation of Kmt2d results in concordant alterations in H3K4me1 levels and CpG island border DNA methylation encroachment. Our findings suggest a role for H3K4me1 in the demarcation of CpG island methylation borders in normal cells, which become eroded in cancer.Item Metadata only Enhanced cardiac phosphoinositide 3-kinase (p110α) using gene therapy attenuates cardiac remodeling in type 2 diabetic mice(American Physiological Society, 2020) Prakoso, D.; De Blasio, M.J.; Tate, M.; Kiriazis, H.; Donner, D.G.; Qian, H.; Nash, D.; Deo, M.; Weeks, K.L.; Parry, L.J.; Gregorevic, P.; McMullen, J.R.; Ritchie, R.H.Diabetic cardiomyopathy is a distinct form of heart disease that represents a major cause of death and disability in diabetic patients, particularly the more prevalent type-2 diabetic population. In the current study, we investigated administration of recombinant adeno-associated viral vectors carrying a constitutively-active PI3K(p110α) construct (rAAV6-caPI3K) at a clinically-relevant time point attenuates diabetic cardiomyopathy in a pre-clinical type-2 diabetes (T2D) model. T2D was induced by a combination of high-fat diet and low-dose streptozotocin, and confirmed by increased body weight, hyperglycemia, and impaired glucose tolerance. After 18 weeks of untreated diabetes, impaired left ventricular (LV) systolic dysfunction was evident, as confirmed by echocardiography. A single tail vein injection of rAAV6-caPI3K gene therapy was then administered. Mice were followed for an additional 8 weeks before end-point. Administration of cardiac targeted rAAV6-caPI3K attenuates diabetes-induced cardiac remodeling by limiting cardiac fibrosis and cardiomyocyte hypertrophy. The diabetes-induced LV systolic dysfunction was reversed with rAAV6-caPI3K as demonstrated by improved fractional shortening and velocity of circumferential fiber shortening. This cardioprotection occurred in combination with reduced LV ROS levels and an associated decrease in markers of endoplasmic reticulum stress. Together, the findings demonstrate that cardiac-selective increases in PI3K(p110α), via rAAV6-caPI3K, attenuates diabetic cardiomyopathy in a mouse model of T2D.