Nip21 gene expression reduces coxsackievirus B3 replication by promoting apoptotic cell death via a mitochondria-dependent pathway.
Our previous studies, using differential mRNA display, suggested that the mouse Nip21 gene may be involved in myocarditis development in the coxsackievirus B3 (CVB3)-infected mouse heart. Sequence comparison indicated that the mouse Nip21 gene shares high sequence homology to human Nip2. This human protein is known to interact with both the apoptosis inhibitor Bcl-2 and a homologous protein, the adenovirus E1B 19-kDa protein. Such interactions implicate Nip21 gene in cell death pathways. To study the function of this gene, we have cloned Nip21 from mouse hearts and established a Tet-On doxycycline-inducible HeLa cell line and a cardiomyocyte H9c2 cell line expressing Nip21 to characterize gene function in relation to apoptosis. We demonstrated that Nip21 expression could induce apoptosis via caspase-depended mitochondria activation. To further determine the function of Nip21 in CVB3-induced apoptosis, the Tet-On/Nip21 HeLa cell line was induced by doxycycline followed by CVB3 infection. We found that activation of caspase-3 and cleavage of poly-(ADP-ribose) polymerase occurred 2 hours earlier than in vector-transfected control cells, suggesting that Nip21 expression enhances CVB3-induced apoptosis. We also demonstrated a significant decrease in HeLa cell and H9c2 cell viability. Particularly, as illustrated by viral plaque assay, CVB3 replication was dramatically reduced in Tet-On HeLa cells, due at least in part to the earlier killing of the host cells by Nip21 overexpression.
Zhang
Huifang M
HM
Department of Pathology and Laboratory Medicine, MRL/The iCAPTUR E Center, University of British Columbia, Vancouver, Canada.
Yanagawa
Bobby
B
Cheung
Paul
P
Luo
Honglin
H
Yuan
Ji
J
Chau
David
D
Wang
Aikun
A
Bohunek
Lubos
L
Wilson
Janet E
JE
McManus
Bruce M
BM
Yang
Decheng
D
D000595:Amino Acid Sequence; D000818:Animals; D017209:Apoptosis; D051023:BH3 Interacting Domain Death Agonist Protein; D002135:Calcium-Binding Proteins / Q000235:genetics; D002352:Carrier Proteins / Q000378:metabolism; D053148:Caspase 3; D020169:Caspases / Q000378:metabolism; D002460:Cell Line; D002470:Cell Survival; D003001:Cloning, Molecular; D003574:Cytochrome c Group / Q000378:metabolism; D029822:Enterovirus B, Human / Q000254:growth & development*; D015870:Gene Expression; D006367:HeLa Cells; D006801:Humans; D051379:Mice; D008928:Mitochondria / Q000502:physiology*; D008969:Molecular Sequence Data; D009206:Myocardium / Q000166:cytology / Q000378:metabolism*; D011506:Proteins / Q000235:genetics / Q000378:metabolism / Q000502:physiology*; D016415:Sequence Alignment; D015398:Signal Transduction; D014779:Virus Replication
D051023:BH3 Interacting Domain Death Agonist Protein; C494814:BID protein, human; C494815:Bid protein, mouse; D002135:Calcium-Binding Proteins; D002352:Carrier Proteins; D003574:Cytochrome c Group; C459664:Nip21 protein, mouse; D011506:Proteins; C505407:CASP3 protein, human; C505409:Casp3 protein, mouse; D053148:Caspase 3; D020169:Caspases