Obesity-linked suppression of membrane-bound O-Acyltransferase 7 (MBOAT7) drives non-alcoholic fatty liver disease
- Cleveland Clinic, United States
- Brigham's and Womens Hospital, Harvard Medical School, United States
- University of California, Los Angeles, United States
- Ionis Pharmaceuticals Inc, United States
- Bioinfo, Canada
- Cleveland State University, United States
- University of Virginia, United States
- Wake Forest University School of Medicine, United States
- Brigham's and Women's Hospital, Harvard Medical School, United States
Abstract
Recent studies have identified a genetic variant rs641738 near two genes encoding membrane bound O-acyltransferase domain-containing 7 (MBOAT7) and transmembrane channel-like 4 (TMC4) that associate with increased risk of non-alcoholic fatty liver disease (NAFLD), non-alcoholic steatohepatitis (NASH), alcohol-related cirrhosis, and liver fibrosis in those infected with viral hepatitis1-7. Based on hepatic expression quantitative trait loci analysis it has been suggested that MBOAT7 loss of function promotes liver disease progression1-7, but this has never been formally tested. Here we show that Mboat7 loss, but not Tmc4, in mice is sufficient to promote the progression of NAFLD in the setting of high fat diet. Mboat7 loss of function is associated with accumulation of its substrate lysophosphatidylinositol (LPI) lipids, and direct administration of LPI promotes hepatic inflammatory and fibrotic transcriptional changes in an Mboat7-dependent manner. These studies reveal a novel role for MBOAT7-driven acylation of LPI lipids in suppressing the progression of NAFLD.
Data availability
Data have been deposited to the NCBI Gene Expression Omnibus, under the accession number GSE138947.
Article and author information
Author details
Jonathan Mark Brown
Funding
National Heart, Lung, and Blood Institute (R01-HL122283)
- Jonathan Mark Brown
National Heart, Lung, and Blood Institute (R00-HL12172)
- Mete Civelek
National Heart, Lung, and Blood Institute (R01-HL106173)
- Matthew Spite
National Heart, Lung, and Blood Institute (F32-HL136044)
- Brian E Sansbury
National Institute of Diabetes and Digestive and Kidney Diseases (R01-DK120679)
- Jonathan Mark Brown
National Institute on Alcohol Abuse and Alcoholism (P50-AA-024333)
- Daniela S Allende
- Jonathan Mark Brown
National Heart, Lung, and Blood Institute (P01 HL029582)
- Paul L Fox
National Institute on Alcohol Abuse and Alcoholism (U01-AA021893)
- Daniela S Allende
National Institute of Diabetes and Digestive and Kidney Diseases (U01-DK061732)
- Daniela S Allende
National Institute of Diabetes and Digestive and Kidney Diseases (R01-DK103637)
- Daniela S Allende
National Heart, Lung, and Blood Institute (P01-HL49373)
- Lawrence L Rudel
National Heart, Lung, and Blood Institute (P01-HL30568)
- Aldons J Lusis
All coauthors are responsible for the content of this work, and different aspects of this work was funded by the National Institutes of Health (NIH) and the American Heart Association (AHA).
Ethics
Animal experimentation: Rat Studies of Diet-Induced ObesitySprague Dawley Rats were received at 12 weeks of age and were housed in individual cages, kept at a constant temperature and ambient humidity in a 12-hlight/dark cycle. Animals were then randomly assigned to either a standard chow diet or a high-fat diet (D12492, 60% fat, Research Diets, New Brunswick, NJ, USA) ad libitum to establish diet-induced obesity as previously described20. After 6 months of HFD-feeding, livers were excised for standard qPCR analysis of Mboat7 expression.Hybrid Mouse Diversity Panel92 inbred strains of 8-week-old male mice (180 individual mice) were fed a high fat, high sucrose diet (D12266B, Research Diets, New Brunswick, NJ) for 8 weeks before tissue collection44. Gene expression of Mboat7 in white adipose tissue and liver were measured and correlated with obesity related traits using biweight midcorrelation analysis as previously described4.Mouse Studies of Mboat7 Loss of FunctionTo explore the role of Mboat7 in diet-induced obesity, NAFLD progression, and insulin resistance, we utilized an in vivo knockdown approach in 8-week old adult mice. Selective knockdown of Mboat7 was accomplished using 2'-O-ethyl (cET) modified antisense oligonucleotides (ASO). All ASOs used in this work were synthesized, screened, and purified as described previously45 by Ionis Pharmaceuticals, Inc. (Carlsbad, CA). For Mboat7 knockdown studies, adult (8 week old) male C57BL/6 mice were purchased from Jackson Labs (Bar Harbor, ME USA), and maintained on either a standard rodent chow diet or a high fat diet (HFD, D12492 from Research Diets Inc.) and injected intraperitoneally biweekly with 12.5 mg/kg of either non-targeting control ASO or one of two independent ASOs directed against murine Mboat7 for a period of 20 weeks. Similar results were seen with two independent ASOs targeting different regions of the Mboat7 mRNA, hence key data using one Mboat7 ASO are shown. All rodents were maintained in an Association for the Assessment and Accreditation of Laboratory Animal Care, International-approved animal facility, and all experimental protocols were approved by the Institutional Animal Care and use Committee of the Cleveland Clinic (IACUC protocols # 2015-1519 and # 2018-2053).
Human subjects: Human MBOAT7 Expression Levels in Lean and Obese SubjectsThe majority of subjects recruited to examine MBOAT7 expression levels were morbidly obese bariatric surgery patients, but we were able to obtain liver biopsies from 10 subjects with a BMI under 30 as normal weight controls. For recruitment, adult patients undergoing gastric bypass surgery at Wake Forest School of Medicine were consented via written consent and enrolled by a member of the study staff following institutionally approved IRB protocols as previously described41. Exclusion criteria included: positive hepatitis C antibody, positive hepatitis B surface antigen, history of liver disease other than NAFLD, Childs A, B, or C cirrhosis, past or present diagnosis/treatment of malignancy other than non-melanocytic skin cancer, INR greater than 1.8 at baseline or need for chronic anticoagulation with warfarin or heparin products, use of immunomodulation for or history of inflammatory diseases including but not limited to malignancy, rheumatoid arthritis, psoriasis, lupus, sarcoidosis and inflammatory bowel disease, and greater or equal to 7 alcohol drinks per week or 3 alcoholic drinks in a given day each week. In addition to bariatric surgery patients, a small number of non-obese subjects (body mass index <30.0) consented to liver biopsy during elective gall bladder removal surgery (n=10). Each subject was assigned a unique identifier which was used throughout the study and did not include any identifiable information about the patient such as name, address, telephone number, social security number, medical record number or any of the identifiers outlined in the HIPAA Privacy Rule regulations. Only the principal investigator had access to the code linking the unique identifier to the study subject. Basic clinical information was obtained via self-reporting and a 15 ml baseline blood sample was obtained at the time of enrollment. A subset of this cohort has been previously described20,41. At the time of surgery, the surgeon collected a roughly 1-gram sample from the lateral left lobe. Wedge biopsies were rinsed with saline and immediately snap frozen in liquid nitrogen in the operating room before subsequent storage at -80{degree sign}C. For data shown in Figure 7 showing levels of MBOAT7 substrate and product lipids, de-identified patient samples from the Cleveland Clinic hepatology clinic (IRB # 10-947) were analyzed. These patients had biopsy proven Ishak fibrosis scores42 of 0 (normal) or 4 (advanced fibrosis). For analysis of hepatic MBOAT7 expression, RNA isolated from liver biopsies were used for quantitative real time PCR (qPCR) as described below.
Copyright
© 2019, Helsley et al.
This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.
Metrics
-
- 4,885
- views
-
- 739
- downloads
-
- 100
- citations
Views, downloads and citations are aggregated across all versions of this paper published by eLife.
Download links
A two-part list of links to download the article, or parts of the article, in various formats.
Downloads (link to download the article as PDF)
Open citations (links to open the citations from this article in various online reference manager services)
Cite this article (links to download the citations from this article in formats compatible with various reference manager tools)
Obesity-linked suppression of membrane-bound O-Acyltransferase 7 (MBOAT7) drives non-alcoholic fatty liver disease
eLife 8:e49882.
https://doi.org/10.7554/eLife.49882
Further reading
-
- Chromosomes and Gene Expression
- Genetics and Genomics
Among the major classes of RNAs in the cell, tRNAs remain the most difficult to characterize via deep sequencing approaches, as tRNA structure and nucleotide modifications can each interfere with cDNA synthesis by commonly-used reverse transcriptases (RTs). Here, we benchmark a recently-developed RNA cloning protocol, termed Ordered Two-Template Relay (OTTR), to characterize intact tRNAs and tRNA fragments in budding yeast and in mouse tissues. We show that OTTR successfully captures both full-length tRNAs and tRNA fragments in budding yeast and in mouse reproductive tissues without any prior enzymatic treatment, and that tRNA cloning efficiency can be further enhanced via AlkB-mediated demethylation of modified nucleotides. As with other recent tRNA cloning protocols, we find that a subset of nucleotide modifications leave misincorporation signatures in OTTR datasets, enabling their detection without any additional protocol steps. Focusing on tRNA cleavage products, we compare OTTR with several standard small RNA-Seq protocols, finding that OTTR provides the most accurate picture of tRNA fragment levels by comparison to "ground truth" Northern blots. Applying this protocol to mature mouse spermatozoa, our data dramatically alter our understanding of the small RNA cargo of mature mammalian sperm, revealing a far more complex population of tRNA fragments - including both 5′ and 3′ tRNA halves derived from the majority of tRNAs – than previously appreciated. Taken together, our data confirm the superior performance of OTTR to commercial protocols in analysis of tRNA fragments, and force a reappraisal of potential epigenetic functions of the sperm small RNA payload.
-
- Chromosomes and Gene Expression
- Genetics and Genomics
Models of nuclear genome organization often propose a binary division into active versus inactive compartments yet typically overlook nuclear bodies. Here, we integrated analysis of sequencing and image-based data to compare genome organization in four human cell types relative to three different nuclear locales: the nuclear lamina, nuclear speckles, and nucleoli. Although gene expression correlates mostly with nuclear speckle proximity, DNA replication timing correlates with proximity to multiple nuclear locales. Speckle attachment regions emerge as DNA replication initiation zones whose replication timing and gene composition vary with their attachment frequency. Most facultative LADs retain a partially repressed state as iLADs, despite their positioning in the nuclear interior. Knock out of two lamina proteins, Lamin A and LBR, causes a shift of H3K9me3-enriched LADs from lamina to nucleolus, and a reciprocal relocation of H3K27me3-enriched partially repressed iLADs from nucleolus to lamina. Thus, these partially repressed iLADs appear to compete with LADs for nuclear lamina attachment with consequences for replication timing. The nuclear organization in adherent cells is polarized with nuclear bodies and genomic regions segregating both radially and relative to the equatorial plane. Together, our results underscore the importance of considering genome organization relative to nuclear locales for a more complete understanding of the spatial and functional organization of the human genome.
