The role of the phosphatidylinositol 4-kinase PI4KA in hepatitis C virus-induced host membrane rearrangement.
BACKGROUND
Hepatitis C virus (HCV), like other positive-sense RNA viruses, replicates on an altered host membrane compartment that has been called the "membranous web." The mechanisms by which the membranous web are formed from cellular membranes are poorly understood. Several recent RNA interference screens have demonstrated a critical role for the host phosphatidylinositol 4-kinase PI4KA in HCV replication. We have sought to define the function of PI4KA in viral replication.


RESULTS
Using a nonreplicative model of membranous web formation, we show that PI4KA silencing leads to aberrant web morphology. Furthermore, we find that PI4KA and its product, phosphatidylinositol 4-phosphate, are enriched on membranous webs and that PI4KA is found in association with NS5A in HCV-infected cells. While the related lipid kinase PI4KB also appears to support HCV replication, it does not interact with NS5A. Silencing of PI4KB does not overtly impair membranous web morphology or phosphatidylinositol 4-phosphate enrichment at webs, suggesting that it acts at a different point in viral replication. Finally, we demonstrate that the aberrant webs induced by PI4KA silencing require the activity of the viral NS3-4A serine protease but not integrity of the host secretory pathway.


CONCLUSIONS
PI4KA is necessary for the local enrichment of PI 4-phosphate at the HCV membranous web and for the generation of morphologically normal webs. We also show that nonreplicative systems of web formation can be used to order molecular events that drive web assembly.
Division of Gastroenterology, Department of Internal Medicine, University of Michigan, Ann Arbor, Michigan, United States of America. andrewwt@med.umich.edu
Andrew W
AW
Tai
Shadi
S
Salloum
D019870:1-Phosphatidylinositol 4-Kinase / Q000378:metabolism*; D045744:Cell Line, Tumor; D002462:Cell Membrane / Q000378:metabolism* / Q000821:virology*; D016000:Cluster Analysis; D004721:Endoplasmic Reticulum / Q000378:metabolism; D020868:Gene Silencing; D006056:Golgi Apparatus / Q000378:metabolism; D016174:Hepacivirus / Q000502:physiology*; D006526:Hepatitis C / Q000201:enzymology / Q000821:virology; D054884:Host-Pathogen Interactions; D006801:Humans; D018129:Phosphatidylinositol Phosphates / Q000378:metabolism; D020815:Polyproteins / Q000378:metabolism; D011485:Protein Binding; D021381:Protein Transport; D015398:Signal Transduction; D017361:Viral Nonstructural Proteins / Q000378:metabolism; D012324:RNA-Dependent RNA Polymerase
D019870:1-Phosphatidylinositol 4-Kinase; D018129:Phosphatidylinositol Phosphates; D020815:Polyproteins; D017361:Viral Nonstructural Proteins; C037178:phosphatidylinositol 4-phosphate; C087036:NS-5 protein, hepatitis C virus; D012324:RNA-Dependent RNA Polymerase