Biology subjects

Attie, A. D.

Publications and source records attributed to Attie, A. D..

18 recordsLinked to original sources

bioRxiv2026

Glutamine Metabolism Supports α cell Mass and Glucagon Secretion

The liver- cell axis is a finely tuned biological rheostat that regulates whole body amino acid availability. Pancreatic cells secrete glucagon that regulates amino acid catabolism through gluconeogenesis and ureagenesis, yet the mechanisms linking amino acid levels to cell growth and function are not fully understood. Here, we identify glutaminase, the enzyme that catalyzes glutamine catabolism, as a critical cell regulator. Glutaminase is highly enriched in cells across species. cell expression of glutaminase is required for nutrient-dependent mTORC1 activation, suppression of AMPK signaling, and sustained expression of the glutamine transporter SLC38A5. This establishes a feed-forward loop linking glutamine metabolism to amino acid sensing and growth. Reduced glutaminase activity impairs dynamic glucagon secretion in response to low glucose and amino acids. Together, these findings highlight the importance of glutamine metabolism in cell growth and hormone secretion and suggest it may play a role in cell adaptation to hyperaminoacidemia.

physiology↗

bioRxiv2026

Distinct genetic architecture of gene and isoform level QTL in the Diversity Outbred (DO) mouse population

Genetic association studies of mRNA abundance phenotypes link regulatory gene loci to mRNA abundance (quantitative trait loci; QTL). Here, we used liver RNA-seq data from a large cohort of the Diversity Outbred (DO) mouse population to map QTL for gene abundance (eQTL), isoform abundance (isoQTL), isoform ratios (irQTL), and splicing (sQTL). QTL studies using this heterogeneous mouse population have been limited to gene-level abundance of mRNAs and have not focused on mRNA isoforms or splicing. We show for the first time that genetic effects on expression are distinct from splicing in the DO mouse population. Using allele-effect patterns from local QTL for protein-coding gene-isoform pairs, we show that genetic variation drives allele-specific isoform usage, generating isoforms whose genetic signals diverge from their aggregated gene-level effects. We conducted pathway enrichment on distal eQTL and isoQTL hotspots and uncovered pathways not detected with eQTL. We then applied a composite mediation approach at these distal hotspots that compares gene-gene, isoform-isoform, and isoform-gene mediator models. By contrasting these causal models of transcriptional regulation, we identified unique associations between mRNA isoforms. We identified candidate genes at irQTL and sQTL hotspots that regulate mRNAs primarily through isoform usage and splicing, including Alkbh1 and Dicer1. For these driver genes, we nominated novel candidate targets through mediation analysis and pathway associations with known RNA-binding targets. We integrated our QTL data with human genetic data, prioritizing effector genes in loci associated with metabolically relevant traits. Our data also suggest that sex and diet influence eQTL and isoQTL distinctly and primarily through distal-acting gene loci. Overall, our findings highlight distinctive genetic effects on transcriptional and post-transcriptional mechanisms of gene regulation.

genetics↗

bioRxiv2025

Modeling metabolic disease susceptibility and resilience in genetically diverse mice

Model organisms have provided critical insights into the basic biology of metabolic disorders, however, one of the greatest limitations to translation has been the absence of the genetic heterogeneity that is characteristic of human populations. We examined metabolic health across three genetically diverse mouse strains fed control (low fat, no sucrose) or unhealthy (high fat, high sucrose) diets and observed a wide range of metabolic responses from overt type 2 diabetes in NZO/HlLtJ mice, to obesity and glucose intolerance in C57BL/6J mice, to complete resilience in CAST/EiJ mice. Analysis of multi-tissue gene expression revealed strain- and tissue-specific responses to diet, with strongest responses in white adipose tissue and pancreatic islets. In pancreatic islets, diet response was limited to just NZO mice, which showed high levels of inflammation and associated {beta} cell dysfunction. Adipose tissue was responsive to diet across all three strains and revealed both common and strain-specific changes in inflammatory and metabolic pathways. Using a complementary outbred mouse resource, we mapped genetic loci associated with strain-specific diet responses, including a monocyte regulatory locus on mChr19. This multi-strain approach to modeling metabolic disease revealed a prominent role of white adipose tissue and lipid-associated inflammation in the determination of individual disease risk in response to unhealthy diets.

physiology↗

bioRxiv2024

Target deconvolution of an insulin hypersecretion-inducer acting through VDAC1 with a distinct transcriptomic signature in beta-cells

Obesity, insulin resistance, and a host of environmental and genetic factors can drive hyperglycemia, causing {beta}-cells to compensate by increasing insulin production and secretion. In type 2 diabetes (T2D), {beta}-cells under these conditions eventually fail. Rare {beta}-cell diseases like congenital hyperinsulinism (HI) also cause inappropriate insulin secretion, and some HI patients develop diabetes. However, the mechanisms of insulin hypersecretion and how it causes {beta}-cell dysfunction are not fully understood. We previously discovered small molecules (e.g. SW016789) that cause insulin hypersecretion and lead to a loss in {beta}-cell function without cell death. Here, we uncover the protein target of SW016789 and provide the first time-course transcriptomic analysis of hypersecretory responses versus thapsigargin-mediated ER stress in {beta}-cells. In mouse MIN6 and human EndoC-{beta}H1 {beta}-cells, we identified and validated VDAC1 as a SW016789 target using photoaffinity proteomics, cellular thermal shift assays, siRNA, and small molecule inhibitors. SW016789 raises membrane potential to enhance Ca2+ influx, potentially through VDAC1. Chronically elevated intracellular Ca2+ appears to underpin the negative impacts of hypersecretion, as nifedipine protected against each small molecule hypersecretion inducer we tested. Using time- course RNAseq, we discovered that hypersecretion induced a distinct transcriptional pattern compared to ER stress. Clustering analyses led us to focus on ER-associated degradation (ERAD) as a potential mediator of the adaptive response. SW016789 reduced the abundance of ERAD substrate OS-9 and pharmacological inhibition of ERAD worsened {beta}-cell survival in response to hypersecretory stress. Changes in other ERAD components in MIN6 and EndoC-{beta}H1 at the protein level were minor with either SW016789 or thapsigargin. However, immunostaining for core ERAD components SEL1L, HRD1, and DERL3 in non-diabetic and T2D human pancreas revealed altered distributions of SEL1L/HRD1 and SEL1L/DERL3 rations in {beta}-cells of T2D islets, in alignment with altered ERAD in stressed {beta}-cells. We conclude that hypersecretory stimuli, including SW016789- mediated VDAC1 activation, cause enhanced Ca2+ influx and insulin release. Subsequent differential gene expression represents a {beta}-cell hypersecretory response signature that is reflected at the protein level for some, but not all genes. A better understanding of how {beta}-cells induce hypersecretion and the mechanisms of negative feedback on secretory rate may lead to the discovery of novel therapeutic targets for T2D and HI.

cell biology↗

bioRxiv2024

Transcripts with high distal heritability mediate genetic effects on complex metabolic traits

Although many genes are subject to local regulation, recent evidence suggests that complex distal regulation may be more important in mediating phenotypic variability. To assess the role of distal gene regulation in complex traits, we combined multi-tissue transcriptomes with physiological outcomes to model diet-induced obesity and metabolic disease in a population of Diversity Outbred mice. Using a novel high-dimensional mediation analysis, we identified a composite transcriptome signature that summarized genetic effects on gene expression and explained 30% of the variation across all metabolic traits. The signature was heritable, interpretable in biological terms, and predicted obesity status from gene expression in an independently derived mouse cohort and multiple human studies. Transcripts contributing most strongly to this composite mediator frequently had complex, distal regulation distributed throughout the genome. These results suggest that trait-relevant variation in transcription is largely distally regulated, but is nonetheless identifiable, interpretable, and translatable across species.

genetics↗

bioRxiv2024

Systems genetics approach uncovers associations between host amylase locus, gut microbiome and metabolic traits in hyperlipidemic mice

The molecular basis for how host genetic variation impacts gut microbial community and bacterial metabolic niches remain largely unknown. We leveraged 90 inbred hyperlipidemic mouse strains from the Hybrid Mouse Diversity Panel (HMDP), previously studied for a variety of cardio-metabolic traits. Metagenomic analysis of cecal DNA followed by genome-wide association analysis identified genomic loci that were associated with microbial enterotypes in the gut. Among these we detected a genetic locus surrounding multiple amylase genes that was associated with abundances of Firmicutes (Lachnospiraceae family) and Bacteroidetes (Muribaculaceae family) taxa encoding distinct starch and sugar metabolism functions. We also found that lower amylase gene number in the mouse genome was associated with higher gut Muribaculaceae levels. Previous work suggests that modulation of host amylase activity impacts the availability of carbohydrates to the host and potentially to gut bacteria. The genetic variants described above were associated with distinct gut microbial communities (enterotypes) with different predicted metabolic capacities for carbohydrate degradation. Mendelian randomization analysis revealed host phenotypes, including liver fibrosis and plasma HDL-cholesterol levels, that were associated with gut microbiome enterotypes. This work reveals novel relationships between host genetic variation, gut microbial enterotypes and host physiology/disease phenotypes in mice.

genetics↗

bioRxiv2023

Heme biosynthesis regulates BCAA catabolism and thermogenesis in brown adipose tissue

With age, people tend to accumulate body fat and reduce energy expenditure1. Brown (BAT) and beige adipose tissue dissipate heat and increase energy expenditure via the activity of the uncoupling protein UCP1 and other thermogenic futile cycles2,3. The activity of brown and beige depots inversely correlates with BMI and age4-11, suggesting that promoting thermogenesis may be an effective approach for combating age-related metabolic disease12-15. Heme is an enzyme cofactor and signaling molecule that we recently showed to regulate BAT function16. Here, we show that heme biosynthesis is the primary contributor to intracellular heme levels in brown adipocytes. Inhibition of heme biosynthesis leads to mitochondrial dysfunction and reduction in UCP1. Although supplementing heme can restore mitochondrial function in heme-synthesis-deficient cells, the downregulation of UCP1 persists due to the accumulation of the heme precursors, particularly propionyl-CoA, which is a product of branched-chain amino acids (BCAA) catabolism. Cold exposure promotes BCAA uptake in BAT, and defects in BCAA catabolism in this tissue hinder thermogenesis17. However, BCAAs contribution to the TCA cycle in BAT and WAT never exceeds 2% of total TCA flux18. Our work offers a way to integrate current literature by describing heme biosynthesis as an important metabolic sink for BCAAs.

cell biology↗

bioRxiv2023

Identification of genetic drivers of plasma lipoproteins in the Diversity Outbred mouse population

Despite great progress in understanding lipoprotein physiology, there is still much to be learned about the genetic drivers of lipoprotein abundance, composition, and function. We used ion mobility spectrometry to survey 16 plasma lipoprotein subfractions in 500 Diversity Outbred (DO) mice maintained on a Western-style diet. We identified 21 quantitative trait loci (QTL) affecting lipoprotein abundance. To refine the QTL and link them to disease risk in humans, we asked if the human homologues of genes located at each QTL were associated with lipid traits in human genome-wide association studies (GWAS). Integration of mouse QTL with human GWAS yielded candidate gene drivers for 18 of the 21 QTL. This approach enabled us to nominate the gene encoding the neutral ceramidase, Asah2, as a novel candidate driver at a QTL on chromosome 19 for large HDL particles (HDL-2b). To experimentally validate Asah2, we surveyed lipoproteins in Asah2-/-mice. Compared to wild-type mice, female Asah2-/- mice showed an increase in several lipoproteins, including HDL. Our results provide insights into the genetic regulation of circulating lipoproteins, as well as mechanisms by which lipoprotein subfractions may affect cardiovascular disease risk in humans.

genetics↗

bioRxiv2023

Pancreatic islet α cell function and proliferation requires the arginine transporter SLC7A2

Interrupting glucagon signaling decreases gluconeogenesis and the fractional extraction of amino acids by liver from blood resulting in lower glycemia. The resulting hyperaminoacidemia stimulates cell proliferation and glucagon secretion via a liver- cell axis. We hypothesized that cells detect and respond to circulating amino acids levels via a unique amino acid transporter repertoire. We found that Slc7a2ISLC7A2 is the most highly expressed cationic amino acid transporter in cells with its expression being three-fold greater in than {beta} cells in both mouse and human. Employing cell culture, zebrafish, and knockout mouse models, we found that the cationic amino acid arginine and SLC7A2 are required for cell proliferation in response to interrupted glucagon signaling. Ex vivo and in vivo assessment of islet function in Slc7a2-/- mice showed decreased arginine-stimulated glucagon and insulin secretion. We found that arginine activation of mTOR signaling and induction of the glutamine transporter SLC38A5 was dependent on SLC7A2, showing that boths role in cell proliferation is dependent on arginine transport and SLC7A2. Finally, we identified single nucleotide polymorphisms in SLC7A2 associated with HbA1c. Together, these data indicate a central role for SLC7A2 in amino acid-stimulated cell proliferation and islet hormone secretion.

physiology↗

bioRxiv2023

HNF4a regulates acyl chain remodeling and ether lipid accumulation in hepatic steatosis

Hepatocyte nuclear factor 4 (HNF4) is an established transcriptional master regulator of differentiation, maintenance, and metabolism. Polymorphisms in HNF4 are linked to several diseases in humans including diabetes and nonalcoholic fatty liver disease (NAFLD). Identifying novel regulation of lipid metabolism by HNF4 would inform on NAFLD development and progression. We directly assessed HNF4 activity through chromatin immunoprecipitation (ChIP)-sequencing and integration of untargeted lipidomics. Direct regulation by HNF4 can be difficult to assess due to the role of HNF4 in liver homeostasis; to rapidly disrupt activity, mice were exposed to cold stress which induces hepatic steatosis in several hours. Cold exposure shifted HNF4 activity with differential genome occupancy of more than 50% of HNF4 binding sites. Focusing on HNF4 binding to promoter with active transcription determined that HNF4 directly regulates fatty acid desaturation, ether lipid synthesis, and peroxisomal biogenesis in response to cold exposure. Integration of lipidomics found that cold exposure increases the very long chain polyunsaturated fatty acid composition of the hepatic lipid pool, including ether lipids, in an HNF4 dependent manner. Because portions of ether lipid synthesis are in the peroxisome and peroxisomal biogenesis is directly HNF4 regulated, we analyzed peroxisomal abundance and found increases with cold exposure that are ablated with loss of HNF4. This peroxisomal regulation was independent of PPAR-- a known regulator of peroxisomes and lipid metabolism--since loss of HNF4 was not rescued by PPAR overexpression. These data determined that regulation of hepatic steatosis by HNF4 is more complex than triglyceride accumulation and includes acyl chain modifications, ether lipid synthesis, and peroxisomal oxidation.

molecular biology↗

bioRxiv2023

Pptc7 maintains mitochondrial protein content by suppressing receptor-mediated mitophagy

Pptc7 is a resident mitochondrial phosphatase essential for maintaining proper mitochondrial content and function. Newborn mice lacking Pptc7 exhibit aberrant mitochondrial protein phosphorylation, suffer from a range of metabolic defects, and fail to survive beyond one day after birth. Using an inducible knockout model, we reveal that loss of Pptc7 in adult mice causes marked reduction in mitochondrial mass concomitant with elevation of the mitophagy receptors Bnip3 and Nix. Consistently, Pptc7-/- mouse embryonic fibroblasts (MEFs) exhibit a major increase in mitophagy that is reversed upon deletion of these receptors. Our phosphoproteomics analyses reveal a common set of elevated phosphosites between perinatal tissues, adult liver, and MEFs-- including multiple sites on Bnip3 and Nix. These data suggest that Pptc7 deletion causes mitochondrial dysfunction via dysregulation of several metabolic pathways and that Pptc7 may directly regulate mitophagy receptor function or stability. Overall, our work reveals a significant role for Pptc7 in the mitophagic response and furthers the growing notion that management of mitochondrial protein phosphorylation is essential for ensuring proper organelle content and function.

biochemistry↗

bioRxiv2022

Genetic variation in mouse islet Ca2+ oscillations reveals novel regulators of islet function

Insufficient insulin secretion to meet metabolic demand results in diabetes. The intracellular flux of Ca2+ into {beta}-cells triggers insulin release. Since genetics strongly influences variation in islet secretory responses, we surveyed islet Ca2+ dynamics in eight genetically diverse mouse strains. We found high strain variation in response to four conditions: 1) 8 mM glucose; 2) 8 mM glucose plus amino acids; 3) 8 mM glucose, amino acids, plus 10 nM GIP; and 4) 2 mM glucose. These stimuli interrogate {beta}-cell function, -cell to {beta}-cell signaling, and incretin responses. We then correlated components of the Ca2+ waveforms to islet protein abundances in the same strains used for the Ca2+ measurements. To focus on proteins relevant to human islet function, we identified human orthologues of correlated mouse proteins that are proximal to glycemic-associated SNPs in human GWAS. Several orthologues have previously been shown to regulate insulin secretion (e.g. ABCC8, PCSK1, and GCK), supporting our mouse-to-human integration as a discovery platform. By integrating these data, we nominated novel regulators of islet Ca2+ oscillations and insulin secretion with potential relevance for human islet function. We also provide a resource for identifying appropriate mouse strains in which to study these regulators.

biochemistry↗

bioRxiv2021

Systems genetics uncovers microbe-lipid-host connections in the murine gut

The molecular bases of how host genetic variation impact gut microbiome remain largely unknown. Here, we used a genetically diverse mouse population and systems genetics strategies to identify interactions between molecular phenotypes, including microbial functions, intestinal transcripts and cecal lipids that influence microbe-host dynamics. Quantitative trait loci (QTL) analysis identified genomic regions associated with variations in bacterial taxa, bacterial functions, including motility, sporulation and lipopolysaccharide production, and levels of bacterial- and host-derived lipids. We found overlapping QTL for the abundance of Akkermansia muciniphila and cecal levels of ornithine lipids (OL). Follow-up studies revealed that A. muciniphila is a major source of these lipids in the gut, provided evidence that OL have immunomodulatory effects and identified intestinal transcripts co-regulated with these traits. Collectively, these results suggest that OL are key players in A. muciniphila-host interactions and support the role of host genetics as a determinant of responses to gut microbes.

genetics↗

bioRxiv2021

INFIMA leverages multi-omics model organism data to identify effector genes of human GWAS variants

Genome-wide association studies have revealed many non-coding variants associated with complex traits. However, model organism studies have largely remained as an untapped resource for unveiling the effector genes of non-coding variants. We develop INFIMA, Integrative Fine-Mapping, to pinpoint causal SNPs for Diversity Outbred (DO) mice eQTL by integrating founder mice multi-omics data including ATAC-seq, RNA-seq, footprinting, and in silico mutation analysis. We demonstrate INFIMAs superior performance compared to alternatives with human and mouse chromatin conformation capture datasets. We apply INFIMA to identify novel effector genes for GWAS variants associated with diabetes. The results of the application are available at http://www.statlab.wisc.edu/shiny/INFIMA/

bioinformatics↗

bioRxiv2020

Coding variants identified in diabetic patients alter PICK1 BAR domain function in insulin granule biogenesis

Bin/amphiphysin/Rvs (BAR) domains are positively charged crescent-shaped modules that shape negatively charged curved lipid membranes during membrane remodeling processes. The BAR domain proteins ICA69, PICK1 and arfaptins have recently been demonstrated to coordinate the budding and formation of immature secretory granules (ISGs) at the trans-Golgi network. Here, we identify four coding variants in the PICK1 gene from a Danish whole-exome screening of diabetic patients, that all involve change of positively charged residues in the PICK1 BAR domain. All four coding variants failed to rescue the insulin content in INS-1E cells upon KD of endogenous PICK1. Moreover, two variants showed dominant negative properties. Interestingly, in vitro assays addressing the BAR domain function suggest that the coding variants compromised membrane binding capacity but increased capacity to cause fission of liposomes. Live confocal microscopy and super-resolution microscopy further revealed that PICK1 resides transiently on ISGs before egress via vesicular budding events. Interestingly, this egress of PICK1 was accelerated in the coding variants. We propose that PICK1 assists or complements the removal of excess membrane and generic membrane trafficking proteins, and possibly also insulin from ISGs during the maturation process and that the coding variants may cause premature budding possibly explaining their dominant negative function.

cell biology↗

bioRxiv2019

Genetic determinants of gut microbiota composition and bile acid profiles in mice

The microbial communities that inhabit the distal gut of humans and other mammals exhibit large inter-individual variation. While host genetics is a known factor that influences gut microbiota composition, the mechanisms underlying this variation remain largely unknown. Bile acids (BAs) are hormones that are produced by the host and chemically modified by gut bacteria. BAs serve as environmental cues and nutrients to microbes, but they can also have antibacterial effects. We hypothesized that host genetic variation in BA metabolism and homeostasis influence gut microbiota composition. To address this, we used the Diversity Outbred (DO) stock, a population of genetically distinct mice derived from eight founder strains. We characterized the fecal microbiota composition and plasma and cecal BA profiles from 400 DO mice maintained on a high-fat high-sucrose diet for [~]22 weeks. Using quantitative trait locus (QTL) analysis, we identified several genomic regions associated with variations in both bacterial and BA profiles. Notably, we found overlapping QTL for Turicibacter sp. and plasma cholic acid, which mapped to a locus containing the gene for the ileal bile acid transporter, Slc10a2. Mediation analysis and subsequent follow-up validation experiments suggest that differences in Slc10a2 gene expression associated with the different strains influences levels of both traits and revealed novel interactions between Turicibacter and BAs. This work illustrates how systems genetics can be utilized to generate testable hypotheses and provide insight into host-microbe interactions.\n\nAuthor summaryInter-individual variation in the composition of the intestinal microbiota can in part be attributed to host genetics. However, the specific genes and genetic variants underlying differences in the microbiota remain largely unknown. To address this, we profiled the fecal microbiota composition of 400 genetically distinct mice, for which genotypic data is available. We identified many loci of the mouse genome associated with changes in abundance of bacterial taxa. One of these loci is also associated with changes in the abundance of plasma bile acids--metabolites generated by the host that influence both microbiota composition and host physiology. Follow up validation experiments provide mechanistic insights linking host genetic differences, with changes in ileum gene expression, bile acid-bacteria interactions and bile acid homeostasis. Together, this work demonstrates how genetic approaches can be used to generate testable hypothesis to yield novel insight into how host genetics shape gut microbiota composition.

systems biology↗

bioRxiv2019

Identification of sample mix-ups and mixtures in microbiome data in Diversity Outbred mice

In a Diversity Outbred mouse project with genotype data on 500 mice, including 297 with microbiome data, we identified three sets of sample mix-ups (two pairs and one trio) as well as at least 15 microbiome samples that appear to be mixtures of pairs of mice. The microbiome data consisted of shotgun sequencing reads from fecal DNA, used to characterize the gut microbial communities present in these mice. These sequence reads included sufficient reads derived from the host mouse to identify the individual. A number of microbiome samples appeared to contain a mixture of DNA from two mice. We describe a method for identifying sample mix-ups in such microbiome data, as well as a method for evaluating sample mixtures in this context.

genetics↗

bioRxiv2018

Pptc7 is an essential phosphatase for promoting mammalian mitochondrial metabolism and biogenesis

Mitochondrial proteins are replete with phosphorylation; however, the origin, abundance, and functional relevance of these modifications are largely unclear. Nonetheless, mitochondria possess multiple resident phosphatases, suggesting that protein dephosphorylation may be broadly important for mitochondrial activities. To explore this, we deleted the poorly characterized matrix phosphatase Pptc7 from mice using CRISPR-Cas9 technology. Strikingly, Pptc7-/- mice exhibited marked hypoketotic hypoglycemia, elevated acylcarnitines, and lactic acidosis, and died soon after birth. Pptc7-/- tissues had significantly diminished mitochondrial size and protein content despite normal transcript levels, but consistently elevated phosphorylation on select mitochondrial proteins. These putative Pptc7 substrates include the protein translocase complex subunit Timm50, whose phosphorylation reduced import activity. We further find that phosphorylation in or near the mitochondrial targeting sequences of multiple proteins can disrupt their import rates and matrix processing. Overall, our data define Pptc7 as a protein phosphatase essential for proper mitochondrial function and biogenesis during the extrauterine transition.

biochemistry↗