Partial purification and characterization of a protein kinase that is activated by nuclear localization signal peptides.
A nuclear localization signal (NLS) is required for the transport of karyophilic proteins from the cytoplasm to the nucleus. In this study, NLS was examined in terms of its effect on diverse cellular functions such as protein phosphorylation reactions. When synthetic peptides containing the NLS of SV40 T-antigen were injected into the cytoplasm of Xenopus oocytes, and the oocytes incubated with [32P]phosphorous-containing medium, a 32 kDa protein was found to be preferentially phosphorylated in an NLS-dependent manner. The incubation of fractionated cytosolic extracts prepared from mouse Ehrlich ascites tumor cells with [gamma-32P]ATP in the presence of the NLS peptides, results in the stimulation of the phosphorylation of several proteins. Similar in vitro stimulation was observed by other functional NLS peptides such as those of polyoma virus T-antigen and nucleoplasmin. Little or no stimulation, however, was detected for peptides of mutant type and reverse type NLS of SV40 T-antigen, and the C-terminal portion of lamin B. Using an in vitro assay, the phosphorylation activity was fractionated chromatographically and a fraction was obtained which contained a high level of activity. The fraction was found to contain three major proteins having molecular masses of 95, 70, and 43 kDa. The in vivo and in vitro results are consistent with the existence of a protein kinase, called NLS kinase, that is specifically activated by NLS peptides.
Kurihara
T
T
Department of Anatomy and Cell Biology, Osaka University Medical School, Japan.
Hori
M
M
Takeda
H
H
Inoue
M
M
Yoneda
Y
Y
D000255:Adenosine Triphosphate / Q000378:metabolism; D000595:Amino Acid Sequence; D000818:Animals; D000952:Antigens, Polyomavirus Transforming / Q000737:chemistry; D002286:Carcinoma, Ehrlich Tumor / Q000378:metabolism; D003593:Cytoplasm / Q000378:metabolism; D004789:Enzyme Activation / Q000187:drug effects; D005260:Female; D051379:Mice; D008969:Molecular Sequence Data; D009865:Oocytes / Q000378:metabolism; D010750:Phosphoproteins / Q000378:metabolism; D010758:Phosphorus / Q000378:metabolism; D010766:Phosphorylation; D011494:Protein Kinases / Q000302:isolation & purification / Q000378:metabolism*; D021382:Protein Sorting Signals / Q000494:pharmacology*; D014981:Xenopus
D000952:Antigens, Polyomavirus Transforming; D010750:Phosphoproteins; D021382:Protein Sorting Signals; D010758:Phosphorus; D000255:Adenosine Triphosphate; D011494:Protein Kinases