Apoptin nucleocytoplasmic shuttling is required for cell type-specific localization, apoptosis, and recruitment of the anaphase-promoting complex/cyclosome to PML bodies.
The chicken anemia virus protein Apoptin selectively induces apoptosis in transformed cells while leaving normal cells intact. This selectivity is thought to be largely due to cell type-specific localization: Apoptin is cytoplasmic in primary cells and nuclear in transformed cells. The basis of Apoptin cell type-specific localization and activity remains to be determined. Here we show that Apoptin is a nucleocytoplasmic shuttling protein whose localization is mediated by an N-terminal nuclear export signal (NES) and a C-terminal nuclear localization signal (NLS). Both signals are required for cell type-specific localization, since Apoptin fragments containing either the NES or the NLS fail to differentially localize in transformed and primary cells. Significantly, cell type-specific localization can be conferred in trans by coexpression of the two separate fragments, which interact through an Apoptin multimerization domain. We have previously shown that Apoptin interacts with the APC1 subunit of the anaphase-promoting complex/cyclosome (APC/C), resulting in G(2)/M cell cycle arrest and apoptosis in transformed cells. We found that the nucleocytoplasmic shuttling activity is critical for efficient APC1 association and induction of apoptosis in transformed cells. Interestingly, both Apoptin multimerization and APC1 interaction are mediated by domains that overlap with the NES and NLS sequences, respectively. Apoptin expression in transformed cells induces the formation of PML nuclear bodies and recruits APC/C to these subnuclear structures. Our results reveal a mechanism for the selective killing of transformed cells by Apoptin.
Heilman
Destin W
DW
Howard Hughes Medical Institute, and Program in Gene Function and Expression, University of Massachusetts Medical School, 364 Plantation Street, Worcester, MA 01605, USA.
Teodoro
Jose G
JG
Green
Michael R
MR
D021581:Active Transport, Cell Nucleus; D064173:Anaphase-Promoting Complex-Cyclosome; D064189:Apc1 Subunit, Anaphase-Promoting Complex-Cyclosome; D017209:Apoptosis; D036022:Capsid Proteins / Q000235:genetics / Q000378:metabolism*; D002289:Carcinoma, Non-Small-Cell Lung / Q000378:metabolism / Q000473:pathology; D002461:Cell Line, Transformed / Q000378:metabolism; D002467:Cell Nucleus / Q000378:metabolism* / Q000648:ultrastructure; D017727:Chicken anemia virus / Q000502:physiology; D003593:Cytoplasm / Q000378:metabolism; D005347:Fibroblasts / Q000378:metabolism; D006801:Humans; D045586:Intranuclear Inclusion Bodies / Q000378:metabolism*; D008175:Lung Neoplasms / Q000378:metabolism / Q000473:pathology; D008856:Microscopy, Fluorescence; D049790:Nuclear Export Signals; D019913:Nuclear Localization Signals; D012867:Skin / Q000378:metabolism; D014407:Tumor Cells, Cultured; D043743:Ubiquitin-Protein Ligase Complexes / Q000378:metabolism*
C577335:ANAPC1 protein, human; D064189:Apc1 Subunit, Anaphase-Promoting Complex-Cyclosome; D036022:Capsid Proteins; D049790:Nuclear Export Signals; D019913:Nuclear Localization Signals; C091717:VP3 protein, Chicken anemia virus; D043743:Ubiquitin-Protein Ligase Complexes; D064173:Anaphase-Promoting Complex-Cyclosome