Overexpression of interferon-gamma-inducible GTPase inhibits coxsackievirus B3-induced apoptosis through the activation of the phosphatidylinositol 3-kinase/Akt pathway and inhibition of viral replication.
Our previous studies using differential mRNA display have shown that interferon-gamma-inducible GTPase (IGTP), was up-regulated in coxsackievirus B3 (CVB3)-infected mouse hearts. In order to explore the effect of IGTP expression on CVB3-induced pathogenesis, we have established a doxycycline-inducible Tet-On HeLa cell line overexpressing IGTP and have analyzed activation of several signaling molecules that are involved in cell survival and death pathways. We found that following IGTP overexpression, protein kinase B/Akt was strongly activated through phosphorylation, which leads to phosphorylation of glycogen synthase kinase-3 (GSK-3). Furthermore, in the presence of CVB3 infection, the intensity of the phosphorylation of Akt was further enhanced and associated with a delayed activation of caspase-9 and caspase-3. These data indicate that IGTP expression appears to confer cell survival in CVB3-infected cells, which was confirmed by 3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium salt cell viability assay. However, the ability of IGTP to induce phosphorylation of Akt and to promote cell survival was attenuated by the phosphotidylinositol-3 kinase (PI3-K) inhibitor LY294002. Transient transfection of the cells with a dominant negative Akt construct followed by doxycycline induction and CVB3 infection reversed Akt phosphorylation to basal levels and returned caspase-3 activity to levels similar to those when the PI3-K inhibitor LY294002 was added. Moreover, IGTP expression inhibited viral replication and delayed CVB3-induced cleavage of eukaryotic translation initiation factor 4G, indicating that IGTP-mediated cell survival relies on not only the activation of PI3-K/Akt, inactivation of GSK-3 and suppression of caspase-9 and caspase-3 but also the inhibition of viral replication.
Zhang
Huifang M
HM
Department of Pathology and Laboratory Medicine, University of British Columbia-MRL/The iCAPTUR4E Centre, St. Paul's Hospital, Vancouver, British Columbia V6Z 1Y6, Canada.
Yuan
Ji
J
Cheung
Paul
P
Luo
Honglin
H
Yanagawa
Bobby
B
Chau
David
D
Stephan-Tozy
Najwan
N
Wong
Brian W
BW
Zhang
Jingchun
J
Wilson
Janet E
JE
McManus
Bruce M
BM
Yang
Decheng
D
D000818:Animals; D017209:Apoptosis; D053148:Caspase 3; D053453:Caspase 9; D020169:Caspases / Q000378:metabolism; D016923:Cell Death; D002470:Cell Survival; D002867:Chromones / Q000494:pharmacology; D004396:Coloring Agents / Q000494:pharmacology; D029822:Enterovirus B, Human / Q000235:genetics*; D004789:Enzyme Activation; D004791:Enzyme Inhibitors / Q000494:pharmacology; D020558:GTP Phosphohydrolases / Q000378:metabolism* / Q000502:physiology*; D005799:Genes, Dominant; D038362:Glycogen Synthase Kinase 3 / Q000378:metabolism; D006367:HeLa Cells; D006801:Humans; D051379:Mice; D009025:Morpholines / Q000494:pharmacology; D009206:Myocardium / Q000201:enzymology / Q000473:pathology*; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism*; D000081082:Phosphoinositide-3 Kinase Inhibitors; D010766:Phosphorylation; D017346:Protein Serine-Threonine Kinases; D011518:Proto-Oncogene Proteins / Q000378:metabolism; D051057:Proto-Oncogene Proteins c-akt; D013778:Tetrazolium Salts / Q000494:pharmacology; D013844:Thiazoles / Q000494:pharmacology; D013997:Time Factors; D014162:Transfection
D053148:Caspase 3; D053453:Caspase 9; D020169:Caspases; D002867:Chromones; D004396:Coloring Agents; D004791:Enzyme Inhibitors; D020558:GTP Phosphohydrolases; D038362:Glycogen Synthase Kinase 3; D009025:Morpholines; D019869:Phosphatidylinositol 3-Kinases; D000081082:Phosphoinositide-3 Kinase Inhibitors; D017346:Protein Serine-Threonine Kinases; D011518:Proto-Oncogene Proteins; D051057:Proto-Oncogene Proteins c-akt; D013778:Tetrazolium Salts; D013844:Thiazoles; C085911:2-(4-morpholinyl)-8-phenyl-4H-1-benzopyran-4-one; C494918:AKT1 protein, human; C505407:CASP3 protein, human; C506433:CASP9 protein, human; C505409:Casp3 protein, mouse; C506434:Casp9 protein, mouse; C101131:Igtp protein, mouse; C022616:thiazolyl blue