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From Human to Mouse and Back Again: Genetic and Genomic Ta(i)les of Islet Dysfunction in Type 2 Diabetes.
No full textType 2 diabetes (T2D) is a complex genetic disease with substantial environmental inputs leading to glucose homeostasis defects. Insulin production is central to proper glucose control, and islet cell dysfunction and death lie at the nexus of T2D genetics and pathophysiology. Comprehensive identification of genes and pathways contributing to these processes is essential for mechanistic understanding and therapeutic targeting. Here, we summarize the latest human and mouse T2D genetic and genomic studies and assess how these parallel variant-to-function efforts and associated data contribute convergent or complementary insights and new opportunities to dissect T2D islet (dys)function. We distill mechanistic and phenotypic studies of candidate T2D effector genes into prevailing themes by which these T2D risk genes likely contribute to islet dysfunction. We assess how recent molecular and metabolic studies in genetically diverse mice (i.e., Collabo-rative Cross, Diversity Outbred) help to nominate new putative T2D effector genes and processes for future exploration and provide examples where these studies illuminate potential limitations of studies using inbred mice. Finally, we discuss opportunities to address knowledge gaps and modeling challenges to translate T2D genetic associations into molecular and pathophysiologic understanding
Host-specific bacterial modulation of airway gene expression and alternative splicing.
No full textThe human microbiome varies extensively between individuals. While there are numerous studies investigating the effects of inter-individual differences on microbiome composition, there are few studies investigating inter-individual effects on microbial modulation of the host or host-specific effects. To address this knowledge gap, we colonized human bronchial epithelial air-liquid interface tissue cultures generated from six different adults with one of three phylogenetically diverse bacteria and compared how each microbe differentially modulated host gene expression in each of the six donors. Microbial treatment had the strongest effect on transcription, followed by donor-specific effects. Gene pathways differed markedly in their donor and microbe specificity; interferon expression was highly donor-dependent, while transcription of epithelial barrier and antibacterial innate immunity genes was predominantly microbially driven. Moreover, we evaluated whether microbial regulation of alternative splicing was modulated by the donor. Strikingly, we found significant nonredundant, donor-specific regulation of alternative splicing exclusively in the gram-positive commensal microbes. These findings highlight that microbial effects on the human airway epithelium are not only species-specific but also deeply individualized, underscoring the importance of the host context in shaping microbe-induced transcriptional and splicing responses.IMPORTANCEMicrobiota are integral regulators of host gene expression, utilizing diverse mechanisms that are shaped by the interplay between microbiome composition and inter-individual differences, i.e., host-specific factors. While previous studies have characterized inter-individual variation in microbiome composition and the effects of variable microbiome composition on the host, the extent to which host-specificity itself regulates host-microbe interactions remains poorly understood. In this study, we address this gap by characterizing changes in epithelial gene expression from six different human donors following colonization with one of three phylogenetically diverse bacteria. By systematically comparing donor-specific responses, we demonstrate that host specificity is a key determinant of the host transcriptional response to microbial colonization. Importantly, we demonstrate that the effects of host specificity are not uniform, but instead are dependent on the colonizing microbe. Our findings underscore the complexity of host-microbe relationships and establish host specificity as a significant factor shaping host-microbe interactions
Low-coverage whole-genome sequencing facilitates accurate and cost-effective haplotype reconstruction in complex mouse crosses.
No full textThe search for the underlying genetic contributions to complex traits and diseases relies on accurate genetic data from populations of interest. Outbred populations, like the Diversity Outbred (DO), are commonly genotyped using commercial SNP arrays, such as the Giga Mouse Universal Genotyping Array (GigaMUGA). However, array genotypes are expensive to collect, subject to significant ascertainment bias, and too sparse to capture the genetic structure of highly recombined mouse crosses. We investigated the efficacy of sequencing-based genotyping by comparing genotyping results between the GigaMUGA, double-digest restriction-site associated DNA sequencing (ddRADseq), and low-coverage whole-genome sequencing (lcWGS). We aligned reads at ~ 1× coverage and imputed segregating SNPs from the eight DO founder strains onto 48 DO genomes and reconstructed their haplotypes using R/qtl2. Haplotype reconstructions derived from all three methods were highly concordant. However, lcWGS more faithfully recapitulated crossover counts and identified more small (\u3c 1 Mb) haplotype blocks at as low as 0.1× coverage. Over 90% of local expression quantitative trait loci identified in a set of 183 DO-derived embryoid bodies using the GigaMUGA were recalled by lcWGS at coverages as low as 0.1×. We recommend that lcWGS be adopted as the primary method of genotyping complex crosses, and cell-based resources derived from them because they are as accurate as array-based reconstructions, robust to ultra-low sequencing depths, may more accurately model haplotypes of the mouse genome that are difficult to resolve with dense reference data, and cost-effective
Whole-genome sequencing reveals individual and cohort level insights into chromosome 9p syndromes.
No full textBACKGROUND: Previous genomic efforts on chromosome 9p deletion and duplication syndromes have utilized low-resolution strategies (i.e., karyotypes, chromosome microarrays). These studies have provided important initial insights into these syndromes. This current study is the first large-scale whole-genome sequencing (WGS) study of 100 individuals from families with chromosome 9p syndromes.
METHODS: Through the newly formed 9P-ARCH (Advanced Research in Chromosomal Health: Genomic, Phenotypic, and Functional Aspects of 9p-Related syndromes) research network, we assembled a cohort of individuals from families with chromosome 9p syndromes. WGS was applied to 100 individuals, and other genomic technologies were applied to a subset of individuals. To prioritize genes on 9p, we utilized two independent approaches: statistical analyses of genomic data and spatial transcriptomic profiling of embryonic mouse tissue. To assess the enrichment of DNVs within genomic regions, we developed a computational tool, DiamondsDenovo ( https://github.com/TNTurnerLab/DiamondsDenovo ).
RESULTS: Unlike previous low-resolution studies, we analyzed the genomic architecture of chromosome 9p syndromes, highlighting fundamental features and their commonalities and differences across individuals. A machine-learning model was developed to predict 9p deletion syndrome based on gene copy number estimates using WGS data. We identified two late-replicating regions containing most structural variant breakpoints in 9p deletion syndrome, pointing to replication-based issues as a potential cause of structural variant formation in most individuals and structural rearrangements in some individuals. Genes on 9p were prioritized based on statistical assessment of human genomic variation and through spatial transcriptomics, with 24 genes (AK3, BRD10, CD274, CDC37L1, DMRT1, DMRT2, DMRT3, DOCK8, GLIS3, JAK2, KANK1, KDM4C, PLPP6, PTPRD, PUM3, RANBP6, RCL1, RFX3, RIC1, SLC1A1, SMARCA2, UHRF2, VLDLR, and ZNG1A) identified as important for the majority (83%) of individuals with 9p deletion syndrome. Testing of the mitochondrial genome revealed excess copy number in individuals with 9p deletion syndrome.
CONCLUSIONS: This study introduces the 9P-ARCH research network that is actively pursuing genomic, phenotypic, and functional aspects of 9p-related syndromes. We advanced the study of 9p-related syndromes both at the individual level and across the cohort through the largest, most comprehensive genomic analysis of 9p-related syndromes to date
Gut dysbiosis patterns in CVID patients with noninfectious complications observed in a germ-free mouse model through fecal microbiota transplantation.
No full textPatients with common variable immunodeficiency (CVID) who develop noninfectious complications (NIC) have worse clinical outcomes than those with infections only (INF). While gut microbiome aberrations have been linked to NIC, reductionist animal models that accurately recapitulate CVID are lacking. Our aim in this study was to uncover potential microbiome roles in the development of NIC in CVID. We performed whole-genome shotgun sequencing on fecal samples from CVID patients with NIC, INF, and their household controls. We also performed fecal microbiota transplants from CVID patients to germ-free mice. We found potentially pathogenic microbe
Young KRAB-zinc finger gene clusters are highly dynamic incubators of ERV-driven genetic heterogeneity in mice.
No full textKRAB-zinc finger proteins (KZFPs) comprise the largest family of mammalian transcription factors, rapidly evolving within and between species. Most KZFPs in human and mice have been found to repress endogenous retroviruses (ERVs) and other retrotransposons, with KZFP gene numbers correlating with the ERV load across species, suggesting coevolution. Whether new KZFPs emerge in response to ERV invasions is currently unknown. Using a combination of long-read sequencing technologies and genome assembly, we present a detailed comparative analysis of young KZFP gene clusters in the mouse lineage, which has undergone recent KZFP gene expansion and ERV infiltration. Detailed annotation of KZFP genes in a cluster on Mus musculus Chromosome 4 reveals parallel expansion and diversification of this locus in different mouse strains (C57BL/6 J, 129S1/SvImJ and CAST/EiJ) and species (Mus spretus and Mus pahari). Our data supports a model by which new ERV integrations within young KZFP gene clusters likely promoted recombination events leading to the emergence of new KZFPs that repress them. At the same time, ERVs also increased their numbers by duplication instead of retrotransposition alone, unraveling a new mechanism for ERV enrichment at these loci
Retention of lysosomal acid sphingomyelinase protects from Niemann-Pick Disease.
No full textNiemann-Pick Disease (NPD) types A and B are lysosomal storage disorders resulting from dysfunction or loss of acid sphingomyelinase (aSMase), which hydrolyzes sphingomyelin (SM) to ceramide and phosphocholine. Patients with NPD-A develop severe neurologic and visceral pathology and rarely live beyond the age of 3 years, while patients with NPD-B typically live to adolescence/early adulthood without neurologic involvement. There are currently no therapies for NPD-A. SMPD1, the gene that encodes aSMase, gives rise to two distinct enzymes - lysosomal sphingomyelinase (L-SMase) and secretory sphingomyelinase (S-SMase), with the latter being associated with inflammation and chemokine amplification. This study sought to define the role of secretory-deficient aSMase variants in NPD. Human NPD fibroblasts transfected with wildtype or two aSMase variants demonstrated that serine residues at either position 507 or 508 in human aSMase retained L-SMase yet lacked S-SMase activity, basally and in response to inflammatory stimuli. We next generated a novel mouse model harboring the S505A variant (aSMas
Visual detection of seizures in mice using supervised machine learning.
No full textSeizures are caused by abnormal synchronous brain activity. The resulting changes in muscle tone, such as twitching, stiffness, or jerking, are used in visual scoring systems such as the Racine scale to quantify seizure intensity. However, visual inspection is time consuming, low throughput, and partially subjective, and there is a need for scalable and rigorous quantitative approaches. We used supervised machine learning approaches to develop automated classifiers to predict seizure severity directly from non-invasive video data. Using the pentylenetetrazole (PTZ)-induced seizure model in mice, we trained video-only classifiers to predict ictal events and combined these events to predict composite seizure intensity for a recording session, as well as time-localized seizure intensity scores. Our results show that seizure events and overall intensity can be rigorously quantified directly from overhead video of mice in a standard open field using supervised approaches. These results enable high-throughput, non-invasive, and standardized seizure scoring for neurogenetics and therapeutic discovery
Effective integration of multi-omics with prior knowledge to identify biomarkers via explainable graph neural networks.
Get PDFThe rapid growth of multi-omics datasets and the wealth of biological knowledge necessitates the development of effective methods for their integration. Such methods are essential for building predictive models and identifying drug targets based on a limited number of samples. We propose a framework called GNNRAI for the supervised integration of multi-omics data with biological priors represented as knowledge graphs. Our framework leverages graph neural networks (GNNs) to model the correlation structures among features from high-dimensional \u27omics data, which reduces the effective dimensions in data and enables us to analyze thousands of genes simultaneously using hundreds of samples. Furthermore, our framework incorporates explainability methods to elucidate informative biomarkers. We apply our framework to Alzheimer\u27s disease (AD) multi-omics data, showing that the integration of transcriptomics and proteomics data with prior AD knowledge is effective, improving the prediction accuracy of AD status over single-omics analyses and highlighting both known and novel AD-predictive biomarkers
High-Throughput Echocardiography-Guided Induction of Myocardial Ischemia/Reperfusion in Mice.
Get PDFBACKGROUND: Mouse models of myocardial ischemia with subsequent heart failure are common approaches to examine heart failure pathology and possible treatment strategies. We sought to establish a high-throughput approach for echocardiography-guided induction of myocardial ischemia/reperfusion (IR) in mice.
METHODS: After visualization of the left coronary artery with high-resolution ultrasound imaging and echocardiographic definition of the level of coronary occlusion, the left anterior descending artery was temporarily occluded with 2 micromanipulator-controlled needles. Functional and molecular changes were assessed and compared with commonly performed surgical techniques.
RESULTS: Echocardiography-guided induction of myocardial IR enabled standardized induction of myocardial IR injury with subsequent left ventricular remodeling. Incorporation of various quality control measures throughout the procedure ensured a high success rate and the absence of relevant postinterventional mortality in experienced hands. Compared with surgical approaches, echocardiography-guided induction of myocardial IR showed a quicker recovery time and induced a less pronounced inflammatory response characterized by decreased local and systemic neutrophil counts. Notably, infarct size and subsequent post-myocardial infarction cardiac dysfunction were comparable between methods. The novel procedure was successfully implemented at different academic institutions with imaging expertise and demonstrated high interinstitutional reproducibility.
CONCLUSIONS: Echocardiography-guided induction of myocardial IR enables high-throughput induction of myocardial IR injury with precise echocardiographic definition of the occlusion level and immediate evaluation of cardiac function during ischemia. The method provides a more clinically relevant assessment of IR sequelae and offers notable animal welfare advantages by eliminating the need for ventilation and thoracotomy, thereby mitigating potential surgery-related confounders