Temporal Proteome and Lipidome Profiles Reveal Hepatitis C Virus-Associated Reprogramming of Hepatocellular Metabolism and Bioenergetics

Proteomic and lipidomic profiling was performed over a time course of acute hepatitis C virus (HCV) infection in cultured Huh-7.5 cells to gain new insights into the intracellular processes influenced by this virus. Our proteomic data suggest that HCV induces early perturbations in glycolysis, the pentose phosphate pathway, and the citric acid cycle, which favor host biosynthetic activities supporting viral replication and propagation. This is followed by a compensatory shift in metabolism aimed at maintaining energy homeostasis and cell viability during elevated viral replication and increasing cellular stress. Complementary lipidomic analyses identified numerous temporal perturbations in select lipid species (e.g. phospholipids and sphingomyelins) predicted to play important roles in viral replication and downstream assembly and secretion events. The elevation of lipotoxic ceramide species suggests a potential link between HCV-associated biochemical alterations and the direct cytopathic effect observed in this in vitro system. Using innovative computational modeling approaches, we further identified mitochondrial fatty acid oxidation enzymes, which are comparably regulated during in vitro infection and in patients with histological evidence of fibrosis, as possible targets through which HCV regulates temporal alterations in cellular metabolic homeostasis.

[1]  T. Veenstra,et al.  Packed capillary reversed-phase liquid chromatography with high-performance electrospray ionization Fourier transform ion cyclotron resonance mass spectrometry for proteomics. , 2001, Analytical chemistry.

[2]  P. Sarnow,et al.  Modulation of Hepatitis C Virus RNA Abundance by a Liver-Specific MicroRNA , 2005, Science.

[3]  K. Chayama,et al.  Serine palmitoyltransferase inhibitor suppresses HCV replication in a mouse model. , 2006, Biochemical and biophysical research communications.

[4]  Ronald J Moore,et al.  Fully automated four-column capillary LC-MS system for maximizing throughput in proteomic analyses. , 2008, Analytical chemistry.

[5]  Nobuyuki Kato,et al.  Modulation of host metabolism as a target of new antivirals☆ , 2007, Advanced Drug Delivery Reviews.

[6]  M A Gerber,et al.  The natural history of community-acquired hepatitis C in the United States. The Sentinel Counties Chronic non-A, non-B Hepatitis Study Team. , 1992, The New England journal of medicine.

[7]  Y. Inoue,et al.  Critical Role of Virion-Associated Cholesterol and Sphingolipid in Hepatitis C Virus Infection , 2008, Journal of Virology.

[8]  Nikola Tolić,et al.  PRISM: A data management system for high‐throughput proteomics , 2006, Proteomics.

[9]  Filippo Conti,et al.  NMR-based metabolic profiling of human hepatoma cells in relation to cell growth by culture media analysis. , 2006, Biochimica et biophysica acta.

[10]  N. Rajewsky,et al.  Silencing of microRNAs in vivo with ‘antagomirs’ , 2005, Nature.

[11]  R. Bartenschlager,et al.  The lipid droplet is an important organelle for hepatitis C virus production , 2007, Nature Cell Biology.

[12]  Soon B. Hwang,et al.  Hepatitis C Virus RNA Replication Occurs on a Detergent-Resistant Membrane That Cofractionates with Caveolin-2 , 2003, Journal of Virology.

[13]  Raymond T Chung,et al.  Apolipoprotein B–dependent hepatitis C virus secretion is inhibited by the grapefruit flavonoid naringenin , 2008, Hepatology.

[14]  Michael G. Katze,et al.  Genomic Analysis Reveals a Potential Role for Cell Cycle Perturbation in HCV-Mediated Apoptosis of Cultured Hepatocytes , 2009, PLoS pathogens.

[15]  Martin M. Matzuk,et al.  Continuous Fatty Acid Oxidation and Reduced Fat Storage in Mice Lacking Acetyl-CoA Carboxylase 2 , 2001, Science.

[16]  C. Voisset,et al.  Ceramide enrichment of the plasma membrane induces CD81 internalization and inhibits hepatitis C virus entry , 2008, Cellular microbiology.

[17]  Ronald J Moore,et al.  Quantitative Proteome Analysis of Human Plasma following in Vivo Lipopolysaccharide Administration Using 16O/18O Labeling and the Accurate Mass and Time Tag Approach*S , 2005, Molecular & Cellular Proteomics.

[18]  S. Kalhan,et al.  The Key Role of Anaplerosis and Cataplerosis for Citric Acid Cycle Function* , 2002, Journal of Biological Chemistry.

[19]  Jin-cui Ye.  Reliance of Host Cholesterol Metabolic Pathways for the Life Cycle of Hepatitis C Virus , 2007, PLoS pathogens.

[20]  Richard D. Smith,et al.  Quantitative Analysis of Human Immunodeficiency Virus Type 1-Infected CD4+ Cell Proteome: Dysregulated Cell Cycle Progression and Nuclear Transport Coincide with Robust Virus Production , 2007, Journal of Virology.

[21]  M. Selbach,et al.  Global analysis of cellular protein translation by pulsed SILAC , 2009, Proteomics.

[22]  Toshiaki Maruyama,et al.  Complete Replication of Hepatitis C Virus in Cell Culture , 2005, Science.

[23]  Matthew D. Dyer,et al.  The Landscape of Human Proteins Interacting with Viruses and Other Pathogens , 2008, PLoS pathogens.

[24]  M. Ripoli,et al.  Hepatitis C virus protein expression causes calcium‐mediated mitochondrial bioenergetic dysfunction and nitro‐oxidative stress , 2007, Hepatology.

[25]  F. Negro,et al.  Hepatitis C virus, steatosis and lipid abnormalities: clinical and pathogenic data , 2009, Liver international : official journal of the International Association for the Study of the Liver.

[26]  Gregory Stephanopoulos,et al.  Quantifying Reductive Carboxylation Flux of Glutamine to Lipid in a Brown Adipocyte Cell Line* , 2008, Journal of Biological Chemistry.

[27]  Thomas Shenk,et al.  Dynamics of the Cellular Metabolome during Human Cytomegalovirus Infection , 2006, PLoS pathogens.

[28]  P. Romano,et al.  Protein synthesis and endoplasmic reticulum stress can be modulated by the hepatitis C virus envelope protein E2 through the eukaryotic initiation factor 2alpha kinase PERK. , 2003, Journal of Virology.

[29]  M. Otto,et al.  Dynamics of Subgenomic Hepatitis C Virus Replicon RNA Levels in Huh-7 Cells after Exposure to Nucleoside Antimetabolites , 2003, Journal of Virology.

[30]  J. Montani,et al.  Substrate cycling between de novo lipogenesis and lipid oxidation: a thermogenic mechanism against skeletal muscle lipotoxicity and glucolipotoxicity , 2004, International Journal of Obesity.

[31]  Mark Gerstein,et al.  The Importance of Bottlenecks in Protein Networks: Correlation with Gene Essentiality and Expression Dynamics , 2007, PLoS Comput. Biol..

[32]  M. Gale,et al.  Hepatitis C virus production by human hepatocytes dependent on assembly and secretion of very low-density lipoproteins , 2007, Proceedings of the National Academy of Sciences.

[33]  F. Chisari,et al.  Hepatitis C virus RNA replication is regulated by host geranylgeranylation and fatty acids. , 2005, Proceedings of the National Academy of Sciences of the United States of America.

[34]  M. Katze,et al.  Translational Control of Viral Gene Expression in Eukaryotes , 2000, Microbiology and Molecular Biology Reviews.

[35]  Michael G. Katze,et al.  Proteome Analysis of Liver Cells Expressing a Full-Length Hepatitis C Virus (HCV) Replicon and Biopsy Specimens of Posttransplantation Liver from HCV-Infected Patients , 2005, Journal of Virology.

[36]  Jin-cui Ye,et al.  Long Chain Acyl-CoA Synthetase 3-mediated Phosphatidylcholine Synthesis Is Required for Assembly of Very Low Density Lipoproteins in Human Hepatoma Huh7 Cells* , 2008, Journal of Biological Chemistry.

[37]  R. Deberardinis,et al.  Beyond aerobic glycolysis: Transformed cells can engage in glutamine metabolism that exceeds the requirement for protein and nucleotide synthesis , 2007, Proceedings of the National Academy of Sciences.

[38]  R. Deberardinis,et al.  The biology of cancer: metabolic reprogramming fuels cell growth and proliferation. , 2008, Cell metabolism.

[39]  H. Hefter,et al.  Liver cell death and anemia in Wilson disease involve acid sphingomyelinase and ceramide , 2007, Nature Medicine.

[40]  笠井 大介.  HCV replication suppresses cellular glucose uptake through down-regulation of cell surface expression of glucose transporters , 2009 .

[41]  Mark Graham,et al.  miR-122 regulation of lipid metabolism revealed by in vivo antisense targeting. , 2006, Cell metabolism.

[42]  J. Yates,et al.  Adaptation of energy metabolism in breast cancer brain metastases. , 2007, Cancer research.

[43]  X. Yao,et al.  Proteolytic 18O labeling for comparative proteomics: model studies with two serotypes of adenovirus. , 2001, Analytical chemistry.

[44]  B. Berkhout,et al.  Proteomic Studies Reveal Coordinated Changes in T-Cell Expression Patterns upon Infection with Human Immunodeficiency Virus Type 1 , 2008, Journal of Virology.

[45]  Navdeep Jaitly,et al.  Decon2LS: An open-source software package for automated processing and visualization of high resolution mass spectrometry data , 2009, BMC Bioinformatics.

[46]  Richard D. Smith,et al.  Robust algorithm for alignment of liquid chromatography-mass spectrometry analyses in an accurate mass and time tag data analysis pipeline. , 2006, Analytical chemistry.

[47]  M. Katze,et al.  HepatoProteomics: Applying proteomic technologies to the study of liver function and disease , 2006, Hepatology.

[48]  Hyunjin Yoon,et al.  Bottlenecks and Hubs in Inferred Networks Are Important for Virulence in Salmonella typhimurium , 2009, J. Comput. Biol..

[49]  Hans-Georg Kräusslich,et al.  The HIV lipidome: a raft with an unusual composition. , 2006, Proceedings of the National Academy of Sciences of the United States of America.

[50]  Xinxia Peng,et al.  Virus-host interactions: from systems biology to translational research. , 2009, Current opinion in microbiology.

[51]  A. Siddiqui,et al.  Hepatitis C Virus Subgenomic Replicons Induce Endoplasmic Reticulum Stress Activating an Intracellular Signaling Pathway , 2002, Journal of Virology.

[52]  Johan Trygg,et al.  Chemometrics in metabonomics. , 2007, Journal of proteome research.

[53]  P. Romano,et al.  Protein Synthesis and Endoplasmic Reticulum Stress Can Be Modulated by the Hepatitis C Virus Envelope Protein E2 through the Eukaryotic Initiation Factor 2α Kinase PERK , 2003, Journal of Virology.

[54]  Michael G. Katze,et al.  Dynamic Host Energetics and Cytoskeletal Proteomes in Human Immunodeficiency Virus Type 1-Infected Human Primary CD4 Cells: Analysis by Multiplexed Label-Free Mass Spectrometry , 2009, Journal of Virology.

[55]  D. Lewin,et al.  Cold elicits the simultaneous induction of fatty acid synthesis and β‐oxidation in murine brown adipose tissue: prediction from differential gene expression and confirmation in vivo , 2002, FASEB journal : official publication of the Federation of American Societies for Experimental Biology.

[56]  Seng-Lai Tan,et al.  Emerging host cell targets for hepatitis C therapy. , 2007, Drug discovery today.

[57]  M. Aoki,et al.  Host sphingolipid biosynthesis as a target for hepatitis C virus therapy , 2005, Nature chemical biology.

[58]  M. Vidal,et al.  Hepatitis C virus infection protein network , 2008, Molecular systems biology.

[59]  R. Aebersold,et al.  A statistical model for identifying proteins by tandem mass spectrometry. , 2003, Analytical chemistry.

[60]  F. Chisari,et al.  Cellular Determinants of Hepatitis C Virus Assembly, Maturation, Degradation, and Secretion , 2007, Journal of Virology.

[61]  Marina A Gritsenko,et al.  Proteomic profiling of human liver biopsies: Hepatitis C virus–induced fibrosis and mitochondrial dysfunction , 2007, Hepatology.

[62]  Navdeep Jaitly,et al.  VIPER: an advanced software package to support high-throughput LC-MS peptide identification , 2007, Bioinform..

[63]  M. Gale,et al.  Identification of FBL2 as a geranylgeranylated cellular protein required for hepatitis C virus RNA replication. , 2005, Molecular cell.

[64]  Xiao-Jiang Feng,et al.  Systems-level metabolic flux profiling identifies fatty acid synthesis as a target for antiviral therapy , 2008, Nature Biotechnology.

[65]  E. Shinya,et al.  Suppression of virus replication via down-modulation of mitochondrial short chain enoyl-CoA hydratase in human glioblastoma cells. , 2007, Antiviral research.

[66]  Qibin Zhang,et al.  Capillary LC coupled with high-mass measurement accuracy mass spectrometry for metabolic profiling. , 2007, Analytical chemistry.

[67]  J. Mullins,et al.  Nef Induces Multiple Genes Involved in Cholesterol Synthesis and Uptake in Human Immunodeficiency Virus Type 1-Infected T Cells , 2005, Journal of Virology.