Sindbis viral vector induced apoptosis requires translational inhibition and signaling through Mcl-1 and Bak.
BACKGROUND
Sindbis viral vectors are able to efficiently target and kill tumor cells in vivo, as shown using pancreatic and ovarian cancer models. Infection results in apoptosis both in vitro and in vivo. Sindbis vector uptake is mediated by the LAMR, which is upregulated on a number of different tumor types, thus conferring specificity of the vector to a wide range of cancers. In this study we elucidate the mechanism of apoptosis in two tumor cell lines, MOSEC, derived from the ovarian epithelium and Pan02, derived from a pancreatic adenocarcinoma. A comprehensive understanding of the mechanism of apoptosis would facilitate the design of more effective vectors for cancer therapy.


RESULTS
The initial phase of Sindbis vector induced apoptosis in MOSEC and Pan02 models reconfirms that viral infection is sensed by PKR due to double-stranded RNA intermediates associated with genomic replication. PKR activation results in translation inhibition through eIF2alpha phosphorylation and initiation of the stress response. Our studies indicate that the roles of two proteins, Mcl-1 and JNK, intimately link Sindbis induced translational arrest and cellular stress. Translational arrest inhibits the synthesis of anti-apoptotic Bcl-2 protein, Mcl-1. JNK activation triggers the release of Bad from 14-3-3, which ultimately results in apoptosis. These signals from translational arrest and cellular stress are propagated to the mitochondria where Bad and Bik bind to Bcl-xl and Mcl-1 respectively. Formation of these heterodimers displaces Bak, which results in caspase 9 cleavage and signaling through the mitochondrial pathway of apoptosis.


CONCLUSIONS
The host cell response to Sindbis is triggered through PKR activation. Our studies demonstrate that PKR activation and subsequent translational arrest is linked to both cellular stress and apoptosis. We have also found the linkage point between translational arrest and apoptosis to be Mcl-1, a protein whose constant translation is required for inhibition of apoptosis. With this information vectors can be designed, which express or repress proteins implicated in this study, to enhance their therapeutic potential.
NYU Cancer Institute and the NYU Gene Therapy Center, NYU School of Medicine, 550 First Avenue, New York, NY 10016, USA. lisa.venticinque@gmail.com
Lisa
L
Venticinque
Daniel
D
Meruelo
D018354:Alphavirus Infections / Q000201:enzymology / Q000473:pathology / Q000821:virology; D000818:Animals; D017209:Apoptosis / Q000187:drug effects*; D045744:Cell Line, Tumor; D003594:Cytoplasmic Granules / Q000187:drug effects / Q000201:enzymology / Q000821:virology; D004789:Enzyme Activation / Q000187:drug effects; D005822:Genetic Vectors / Q000494:pharmacology*; D048031:JNK Mitogen-Activated Protein Kinases / Q000378:metabolism; D051379:Mice; D008810:Mice, Inbred C57BL; D008954:Models, Biological; D064549:Myeloid Cell Leukemia Sequence 1 Protein; D014176:Protein Biosynthesis / Q000187:drug effects*; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000378:metabolism*; D012330:RNA, Double-Stranded / Q000378:metabolism; D015398:Signal Transduction / Q000187:drug effects*; D012845:Sindbis Virus / Q000235:genetics*; D013312:Stress, Physiological / Q000187:drug effects; D050998:bcl-2 Homologous Antagonist-Killer Protein / Q000378:metabolism*; D019892:eIF-2 Kinase / Q000378:metabolism
C579094:Mcl1 protein, mouse; D064549:Myeloid Cell Leukemia Sequence 1 Protein; D019253:Proto-Oncogene Proteins c-bcl-2; D012330:RNA, Double-Stranded; D050998:bcl-2 Homologous Antagonist-Killer Protein; D019892:eIF-2 Kinase; D048031:JNK Mitogen-Activated Protein Kinases