title: Genomic Characterization of Metformin Hepatic Response.
abstract: Metformin is used as a first-line therapy for type 2 diabetes (T2D) and prescribed for numerous other diseases. However, its mechanism of action in the liver has yet to be characterized in a systematic manner. To comprehensively identify genes and regulatory elements associated with metformin treatment, we carried out RNA-seq and ChIP-seq (H3K27ac, H3K27me3) on primary human hepatocytes from the same donor treated with vehicle control, metformin or metformin and compound C, an AMP-activated protein kinase (AMPK) inhibitor (allowing to identify AMPK-independent pathways). We identified thousands of metformin responsive AMPK-dependent and AMPK-independent differentially expressed genes and regulatory elements. We functionally validated several elements for metformin-induced promoter and enhancer activity. These include an enhancer in an ataxia telangiectasia mutated (ATM) intron that has SNPs in linkage disequilibrium with a metformin treatment response GWAS lead SNP (rs11212617) that showed increased enhancer activity for the associated haplotype. Expression quantitative trait locus (eQTL) liver analysis and CRISPR activation suggest that this enhancer could be regulating ATM, which has a known role in AMPK activation, and potentially also EXPH5 and DDX10, its neighboring genes. Using ChIP-seq and siRNA knockdown, we further show that activating transcription factor 3 (ATF3), our top metformin upregulated AMPK-dependent gene, could have an important role in gluconeogenesis repression. Our findings provide a genome-wide representation of metformin hepatic response, highlight important sequences that could be associated with interindividual variability in glycemic response to metformin and identify novel T2D treatment candidates.
authors.affiliation: Department of Bioengineering and Therapeutic Sciences, University of California San Francisco, San Francisco, California, United States of America.|Institute for Human Genetics, University of California San Francisco, San Francisco, California, United States of America.|Department of General Biology, Institute of Biological Sciences, Federal University of Minas Gerais, Belo Horizonte, Minas Gerais, Brazil.
authors.forename: Marcelo R
authors.initials: MR
authors.lastname: Luizon
authors.affiliation: Department of Bioengineering and Therapeutic Sciences, University of California San Francisco, San Francisco, California, United States of America.|Institute for Human Genetics, University of California San Francisco, San Francisco, California, United States of America.
authors.forename: Walter L
authors.initials: WL
authors.lastname: Eckalbar
authors.affiliation: Department of Bioengineering and Therapeutic Sciences, University of California San Francisco, San Francisco, California, United States of America.|Institute for Human Genetics, University of California San Francisco, San Francisco, California, United States of America.|School of Pharmaceutical Sciences, Tsinghua University, Beijing, China.
authors.forename: Yao
authors.initials: Y
authors.lastname: Wang
authors.affiliation: Department of Bioengineering and Therapeutic Sciences, University of California San Francisco, San Francisco, California, United States of America.|Institute for Human Genetics, University of California San Francisco, San Francisco, California, United States of America.
authors.forename: Stacy L
authors.initials: SL
authors.lastname: Jones
authors.affiliation: Department of Bioengineering and Therapeutic Sciences, University of California San Francisco, San Francisco, California, United States of America.|Institute for Human Genetics, University of California San Francisco, San Francisco, California, United States of America.
authors.forename: Robin P
authors.initials: RP
authors.lastname: Smith
authors.affiliation: Library and Center for Knowledge Management, University of California San Francisco, San Francisco, California, United States of America.
authors.forename: Megan
authors.identifier: http://orcid.org/0000-0001-8219-8660
authors.initials: M
authors.lastname: Laurance
authors.affiliation: Department of Bioengineering and Therapeutic Sciences, University of California San Francisco, San Francisco, California, United States of America.
authors.forename: Lawrence
authors.initials: L
authors.lastname: Lin
authors.affiliation: Bioinformatics Research Center, North Carolina State University, Raleigh, North Carolina, United States of America.
authors.forename: Paul J
authors.initials: PJ
authors.lastname: Gallins
authors.affiliation: Eshelman School of Pharmacy, University of North Carolina at Chapel Hill, Chapel Hill, North Carolina, United States of America.
authors.forename: Amy S
authors.initials: AS
authors.lastname: Etheridge
authors.affiliation: Bioinformatics Research Center, North Carolina State University, Raleigh, North Carolina, United States of America.
authors.forename: Fred
authors.initials: F
authors.lastname: Wright
authors.affiliation: Bioinformatics Research Center, North Carolina State University, Raleigh, North Carolina, United States of America.
authors.forename: Yihui
authors.initials: Y
authors.lastname: Zhou
authors.affiliation: Merck Research Labs, Merck & Co. Inc., Kenilworth, New Jersey, United States of America.
authors.forename: Cliona
authors.identifier: http://orcid.org/0000-0003-4578-3910
authors.initials: C
authors.lastname: Molony
authors.affiliation: Eshelman School of Pharmacy, University of North Carolina at Chapel Hill, Chapel Hill, North Carolina, United States of America.
authors.forename: Federico
authors.initials: F
authors.lastname: Innocenti
authors.affiliation: Department of Bioengineering and Therapeutic Sciences, University of California San Francisco, San Francisco, California, United States of America.
authors.forename: Sook Wah
authors.initials: SW
authors.lastname: Yee
authors.affiliation: Department of Bioengineering and Therapeutic Sciences, University of California San Francisco, San Francisco, California, United States of America.|Institute for Human Genetics, University of California San Francisco, San Francisco, California, United States of America.
authors.forename: Kathleen M
authors.initials: KM
authors.lastname: Giacomini
authors.affiliation: Department of Bioengineering and Therapeutic Sciences, University of California San Francisco, San Francisco, California, United States of America.|Institute for Human Genetics, University of California San Francisco, San Francisco, California, United States of America.
authors.forename: Nadav
authors.initials: N
authors.lastname: Ahituv
mesh_terms: D055372:AMP-Activated Protein Kinases / Q000096:biosynthesis* / Q000235:genetics; D051700:Activating Transcription Factor 3 / Q000235:genetics*; D048868:Adaptor Proteins, Signal Transducing / Q000235:genetics; D064007:Ataxia Telangiectasia Mutated Proteins / Q000096:biosynthesis* / Q000235:genetics; D053487:DEAD-box RNA Helicases / Q000235:genetics; D003924:Diabetes Mellitus, Type 2 / Q000188:drug therapy* / Q000235:genetics / Q000473:pathology; D004742:Enhancer Elements, Genetic; D055785:Gene Knockdown Techniques; D005943:Gluconeogenesis / Q000235:genetics; D006239:Haplotypes; D022781:Hepatocytes / Q000187:drug effects / Q000378:metabolism; D006801:Humans; D015810:Linkage Disequilibrium; D008099:Liver / Q000187:drug effects / Q000378:metabolism*; D008687:Metformin / Q000009:adverse effects / Q000627:therapeutic use; D020641:Polymorphism, Single Nucleotide
chemical_list: C490449:ATF3 protein, human; D051700:Activating Transcription Factor 3; D048868:Adaptor Proteins, Signal Transducing; C514892:EXPH5 protein, human; D008687:Metformin; C576324:ATM protein, human; D064007:Ataxia Telangiectasia Mutated Proteins; D055372:AMP-Activated Protein Kinases; C499217:DDX10 protein, human; D053487:DEAD-box RNA Helicases