Class III phosphatidylinositol 4-kinase alpha and beta are novel host factor regulators of hepatitis C virus replication.
Host factor pathways are known to be essential for hepatitis C virus (HCV) infection and replication in human liver cells. To search for novel host factor proteins required for HCV replication, we screened a subgenomic genotype 1b replicon cell line (Luc-1b) with a kinome and druggable collection of 20,779 siRNAs. We identified and validated several enzymes required for HCV replication, including class III phosphatidylinositol 4-kinases (PI4KA and PI4KB), carbamoyl-phosphate synthetase 2, aspartate transcarbamylase, and dihydroorotase (CAD), and mevalonate (diphospho) decarboxylase. Knockdown of PI4KA could inhibit the replication and/or HCV RNA levels of the two subgenomic genotype 1b clones (SG-1b and Luc-1b), two subgenomic genotype 1a clones (SG-1a and Luc-1a), JFH-1 genotype 2a infectious virus (JFH1-2a), and the genomic genotype 1a (FL-1a) replicon. In contrast, PI4KB knockdown inhibited replication and/or HCV RNA levels of Luc-1b, SG-1b, and Luc-1a replicons. The small molecule inhibitor, PIK93, was found to block subgenomic genotype 1b (Luc-1b), subgenomic genotype 1a (Luc-1a), and genomic genotype 2a (JFH1-2a) infectious virus replication in the nanomolar range. PIK93 was characterized by using quantitative chemical proteomics and in vitro biochemical assays to demonstrate PIK93 is a bone fide PI4KA and PI4KB inhibitor. Our data demonstrate that genetic or pharmacological modulation of PI4KA and PI4KB inhibits multiple genotypes of HCV and represents a novel druggable class of therapeutic targets for HCV infection.
Borawski
Jason
J
Novartis Institutes for Biomedical Research, 250 Massachusetts Ave., Cambridge, Massachusetts 02139, USA.
Troke
Philip
P
Puyang
Xiaoling
X
Gibaja
Veronica
V
Zhao
Shanchaun
S
Mickanin
Craig
C
Leighton-Davies
Juliet
J
Wilson
Christopher J
CJ
Myer
Vic
V
Cornellataracido
Ivan
I
Baryza
Jeremy
J
Tallarico
John
J
Joberty
Gerard
G
Bantscheff
Marcus
M
Schirle
Markus
M
Bouwmeester
Tewis
T
Mathy
Joanna E
JE
Lin
Kai
K
Compton
Teresa
T
Labow
Mark
M
Wiedmann
Brigitte
B
Gaither
L Alex
LA
D019870:1-Phosphatidylinositol 4-Kinase / Q000737:chemistry / Q000378:metabolism*; D000998:Antiviral Agents / Q000494:pharmacology; D001667:Binding, Competitive; D002460:Cell Line; D020868:Gene Silencing; D005838:Genotype; D016174:Hepacivirus / Q000235:genetics* / Q000378:metabolism*; D006801:Humans; D020128:Inhibitory Concentration 50; D008099:Liver / Q000821:virology*; D013058:Mass Spectrometry / Q000379:methods; D040901:Proteomics / Q000379:methods; D034741:RNA, Small Interfering / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013844:Thiazoles / Q000494:pharmacology; D014779:Virus Replication
D000998:Antiviral Agents; D034741:RNA, Small Interfering; D013844:Thiazoles; D019870:1-Phosphatidylinositol 4-Kinase