Viral oncolysis that targets Raf-1 signaling control of nuclear transport.
The central role of Raf protein kinase isoforms in human cancer demands specific anti-Raf therapeutic inhibitors. Parvoviruses are currently used in experimental cancer therapy due to their natural oncotropism and lytic life cycle. In searching for mechanisms underlying parvovirus oncolysis, we found that trimers of the major structural protein (VP) of the parvovirus minute virus of mice (MVM), which have to be imported into the nucleus for capsid assembly, undergo phosphorylation by the Raf-1 kinase. Purified Raf-1 phosphorylated the capsid subunits in vitro to the two-dimensional pattern found in natural MVM infections. VP trimers isolated from mammalian cells translocated into the nucleus of digitonin-permeabilized human cells. In contrast, VP trimers isolated from insect cells, which are devoid of Raf-1, were neither phosphorylated nor imported into the mammalian nucleus. However, the coexpression of a constitutively active Raf-1 kinase in insect cells restored VP trimer phosphorylation and nuclear transport competence. In MVM-infected normal and transformed cells, Raf-1 inhibition resulted in cytoplasmic retention of capsid proteins, preventing their nuclear assembly and progeny virus maturation. The level of Raf-1 activity in cancer cells was consistent with the extent of VP specific phosphorylation and with the permissiveness to MVM infection. Thus, Raf-1 control of nuclear translocation of MVM capsid assembly intermediates provides a novel target for viral oncolysis. MVM may reinforce specific therapies against frequent human cancers with deregulated Raf signaling.
Riolobos
Laura
L
Centro de Biología Molecular "Severo Ochoa", Universidad Autónoma de Madrid, Cantoblanco, 28049 Madrid, Spain.
Valle
Noelia
N
Hernando
Eva
E
Maroto
Beatriz
B
Kann
Michael
M
Almendral
José M
JM
D021581:Active Transport, Cell Nucleus; D000818:Animals; D036022:Capsid Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D002460:Cell Line; D045744:Cell Line, Tumor; D002472:Cell Transformation, Viral; D006367:HeLa Cells; D006801:Humans; D051379:Mice; D008915:Minute Virus of Mice / Q000235:genetics / Q000502:physiology*; D050130:Oncolytic Virotherapy; D050504:Oncolytic Viruses / Q000235:genetics / Q000502:physiology*; D010766:Phosphorylation; D020836:Protein Structure, Quaternary; D021122:Protein Subunits; D019908:Proto-Oncogene Proteins c-raf / Q000378:metabolism*; D051381:Rats; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D015398:Signal Transduction; D018411:Spodoptera; D019065:Virus Assembly
D036022:Capsid Proteins; D021122:Protein Subunits; D011994:Recombinant Proteins; C442549:VP1 protein, Mice minute virus; D019908:Proto-Oncogene Proteins c-raf