Protein kinase B/Akt regulates coxsackievirus B3 replication through a mechanism which is not caspase dependent.
The role of signaling pathways including the mitogen-activated protein kinases (MAPKs) and phosphatidylinositol 3-kinase (PI3K) during viral infection has gained much recent attention. Our laboratory reported on an important regulatory role for extracellular signal-regulated kinases (ERK1/2), subfamily members of the MAPKs, during coxsackievirus B3 (CVB3) infection. However, the role of the PI3K pathway in CVB3 infection has not been well characterized. CVB3 is the most common known viral infectant of heart muscle that directly injures and kills infected cardiac myocytes during the myocarditic process. In the present study, we investigated the role of protein kinase B (PKB) (also known as Akt), a general downstream mediator of survival signals through the PI3K cascade, in regulating CVB3 replication and virus-induced apoptosis in a well-established HeLa cell model. We have demonstrated that CVB3 infection leads to phosphorylation of PKB/Akt on both Ser-473 and Thr-308 residues through a PI3K-dependent mechanism. Transfection of HeLa cells with a dominant negative mutant of Akt1 or pretreatment of wild-type HeLa cells with the specific PI3K inhibitor LY294002 significantly suppresses viral RNA expression, as reflected in diminished viral capsid protein expression and viral release. Dominant negative Akt1 and LY294002 also increase apoptosis in infected cells, which can be reversed by addition of the general caspase inhibitor benzyloxycarbonyl-Val-Ala-Asp-fluoromethylketone (zVAD.fmk). Interestingly, blocking of apoptosis by zVAD.fmk does not reverse the viral RNA translation blockade, indicating that the inhibitory effect of dominant negative Akt1 on viral protein expression is not caspase dependent. In addition, we showed that the attachment of virus to its receptor-coreceptor complex is not sufficient for PKB/Akt activation and that postentry viral replication is required for Akt phosphorylation. Taken together, these data illustrate a new and imperative role for Akt in CVB3 infection in HeLa cells and show that the PI3K/Akt signaling is beneficial to CVB3 replication.
Esfandiarei
Mitra
M
The James Hogg iCAPTURE Centre for Cardiovascular and Pulmonary Research, Department of Pathology and Laboratory Medicine, St. Paul's Hospital/Providence Health Care-University of British Columbia, Vancouver, Canada.
Luo
Honglin
H
Yanagawa
Bobby
B
Suarez
Agripina
A
Dabiri
Darya
D
Zhang
Jianchang
J
McManus
Bruce M
BM
D000590:Amino Acid Chloromethyl Ketones / Q000494:pharmacology; D017209:Apoptosis; D001665:Binding Sites / Q000235:genetics; D036022:Capsid Proteins / Q000096:biosynthesis; D020169:Caspases / Q000378:metabolism; D002867:Chromones / Q000494:pharmacology; D015853:Cysteine Proteinase Inhibitors / Q000494:pharmacology; D029822:Enterovirus B, Human / Q000502:physiology*; D004791:Enzyme Inhibitors / Q000494:pharmacology; D006367:HeLa Cells; D006801:Humans; D020935:MAP Kinase Signaling System; D009025:Morpholines / Q000494:pharmacology; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism; D000081082:Phosphoinositide-3 Kinase Inhibitors; D010766:Phosphorylation; D017346:Protein Serine-Threonine Kinases; D011518:Proto-Oncogene Proteins / Q000737:chemistry / Q000235:genetics / Q000502:physiology*; D051057:Proto-Oncogene Proteins c-akt; D012367:RNA, Viral / Q000096:biosynthesis; D012694:Serine / Q000737:chemistry; D013912:Threonine / Q000737:chemistry; D014466:Ultraviolet Rays; D014779:Virus Replication / Q000187:drug effects / Q000502:physiology*
D000590:Amino Acid Chloromethyl Ketones; D036022:Capsid Proteins; D020169:Caspases; D002867:Chromones; D015853:Cysteine Proteinase Inhibitors; D004791:Enzyme Inhibitors; 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; D012367:RNA, Viral; D012694:Serine; D013912:Threonine; C096713:benzyloxycarbonylvalyl-alanyl-aspartyl fluoromethyl ketone; C085911:2-(4-morpholinyl)-8-phenyl-4H-1-benzopyran-4-one; C494918:AKT1 protein, human