Identification of a signal for nuclear targeting in platelet-derived-growth-factor-related molecules.
The v-vis gene encodes p28sis, the transforming protein of simian sarcoma virus. This gene resulted from a fusion of the env gene of simian sarcoma-associated virus and the woolly monkey gene for the B chain of platelet-derived growth factor (PDGF). Previous work has shown that the v-sis gene product undergoes signal sequence cleavage, glycosylation, dimerization, and proteolytic processing to yield a secreted form of the protein. It transport across the endoplasmic reticulum is blocked by the introduction of a charged amino acid residue within the signal sequence, the protein does not dimerize, is not secreted, and is no longer transforming as assayed by focus-forming ability in NIH 3T3 cells. Instead, this mutant protein localizes to the nucleus as demonstrated by both indirect immunofluorescence and cell fractionation. Using a series of deletion mutations, we delimited an amino acid sequence within this protein which is responsible for nuclear localization. This region is completely conserved in the predicted human c-sis protein, although it lies outside of regions required for transformation by the v-sis gene product. This nuclear transport signal is contained within amino acid residues 237 to 255, RVTIRTVRVRRPPKGKHRK. An amino acid sequence containing these residues is capable of directing cytoplasmic v-sis mutant proteins to the nucleus. This sequence is also capable of directing less efficient nuclear transport of a normally cytoplasmic protein, pyruvate kinase. Pulse-chase experiments indicate that the half-lives of nuclear and cytoplasmic v-sis mutant proteins are approximately 35 min. Using the heat-inducible hsp70 promoter from Drosophila melanogaster, we showed that the nuclear v-sis protein accumulates in the nucleus within 30 min of induction. The identification of a nuclear transport signal in the v-sis gene product raises interesting questions regarding the possibility of some function for PDGF or PDGF-related molecules in the nucleus.
Lee
B A
BA
Department of Biology, University of California, San Diego, La Jolla 92093.
Maher
D W
DW
Hannink
M
M
Donoghue
D J
DJ
D000818:Animals; D002451:Cell Compartmentation; D002460:Cell Line; D002467:Cell Nucleus / Q000378:metabolism; D002522:Chlorocebus aethiops; D003593:Cytoplasm / Q000378:metabolism; D004252:DNA Mutational Analysis; D005455:Fluorescent Antibody Technique; D008970:Molecular Weight; D009687:Nuclear Proteins / Q000502:physiology*; D009856:Oncogene Proteins, Viral / Q000502:physiology*; D009857:Oncogenes; D010982:Platelet-Derived Growth Factor / Q000502:physiology*
D009687:Nuclear Proteins; D009856:Oncogene Proteins, Viral; D010982:Platelet-Derived Growth Factor