Proteome analysis of liver cells expressing a full-length hepatitis C virus (HCV) replicon and biopsy specimens of posttransplantation liver from HCV-infected patients.
The development of a reproducible model system for the study of hepatitis C virus (HCV) infection has the potential to significantly enhance the study of virus-host interactions and provide future direction for modeling the pathogenesis of HCV. While there are studies describing global gene expression changes associated with HCV infection, changes in the proteome have not been characterized. We report the first large-scale proteome analysis of the highly permissive Huh-7.5 cell line containing a full-length HCV replicon. We detected >4,200 proteins in this cell line, including HCV replicon proteins, using multidimensional liquid chromatographic (LC) separations coupled to mass spectrometry. Consistent with the literature, a comparison of HCV replicon-positive and -negative Huh-7.5 cells identified expression changes of proteins involved in lipid metabolism. We extended these analyses to liver biopsy material from HCV-infected patients where a total of >1,500 proteins were detected from only 2 mug of liver biopsy protein digest using the Huh-7.5 protein database and the accurate mass and time tag strategy. These findings demonstrate the utility of multidimensional proteome analysis of the HCV replicon model system for assisting in the determination of proteins/pathways affected by HCV infection. Our ability to extend these analyses to the highly complex proteome of small liver biopsies with limiting protein yields offers the unique opportunity to begin evaluating the clinical significance of protein expression changes associated with HCV infection.
Jacobs
Jon M
JM
Department of Microbiology, University of Washington, Box 358070, Seattle, WA 98195-8070, USA. honey@u.washington.edu
Diamond
Deborah L
DL
Chan
Eric Y
EY
Gritsenko
Marina A
MA
Qian
Weijun
W
Stastna
Miroslava
M
Baas
Tracey
T
Camp
David G
DG
Carithers
Robert L
RL
Smith
Richard D
RD
Katze
Michael G
MG
D000595:Amino Acid Sequence; D001706:Biopsy; D045744:Cell Line, Tumor; D016174:Hepacivirus / Q000235:genetics / Q000378:metabolism / Q000472:pathogenicity*; D006526:Hepatitis C / Q000821:virology; D022781:Hepatocytes / Q000378:metabolism / Q000821:virology; D006801:Humans; D008099:Liver / Q000166:cytology / Q000821:virology; D016031:Liver Transplantation / Q000009:adverse effects; D008969:Molecular Sequence Data; D011506:Proteins / Q000235:genetics / Q000378:metabolism*; D020543:Proteome; D012093:Replicon / Q000235:genetics / Q000502:physiology*; D014764:Viral Proteins / Q000235:genetics / Q000378:metabolism
D011506:Proteins; D020543:Proteome; D014764:Viral Proteins