Nuclear import of the BTB/POZ transcriptional regulator Kaiso.
Kaiso is a BTB/POZ transcription factor that functions in vitro as a transcriptional repressor of the matrix metalloproteinase gene matrilysin and the non-canonical Wnt signaling gene Wnt-11, and as an activator of the acetylcholine-receptor-clustering gene rapsyn. Similar to other BTB/POZ proteins (e.g. Bcl-6, PLZF, HIC-1), endogenous Kaiso localizes predominantly to the nuclei of mammalian cells. To date, however, the mechanism of nuclear import for most POZ transcription factors, including Kaiso, remain unknown. Here, we report the identification and characterization of a highly basic nuclear localization signal (NLS) in Kaiso. The functionality of this NLS was verified by its ability to target a heterologous beta-galactosidase/green-fluorescent-protein fusion protein to nuclei. The mutation of one positively charged lysine to alanine in the NLS of full-length Kaiso significantly inhibited its nuclear localization in various cell types. In addition, wild-type Kaiso, but not NLS-defective Kaiso, interacted directly with the nuclear import receptor Importin-alpha2 both in vitro and in vivo. Finally, minimal promoter assays using a sequence-specific Kaiso-binding-site fusion with luciferase as reporter demonstrated that the identified NLS was crucial for Kaiso-mediated transcriptional repression. The identification of a Kaiso NLS thus clarifies the mechanism by which Kaiso translocates to the nucleus to regulate transcription of genes with diverse roles in cell growth and development.
Kelly
Kevin F
KF
Department of Biology, McMaster University, 1280 Main Street West, Hamilton, ON, L8S 4K1, Canada.
Otchere
Abena A
AA
Graham
Monica
M
Daniel
Juliet M
JM
D021581:Active Transport, Cell Nucleus; D000409:Alanine / Q000737:chemistry; D000595:Amino Acid Sequence; D000818:Animals; D051177:Catenins; D015815:Cell Adhesion Molecules / Q000378:metabolism; D002467:Cell Nucleus / Q000378:metabolism; D005982:Glutathione Transferase / Q000378:metabolism; D049452:Green Fluorescent Proteins / Q000378:metabolism; D006367:HeLa Cells; D006801:Humans; D015151:Immunoblotting; D047468:Immunoprecipitation; D008156:Luciferases / Q000378:metabolism; D008239:Lysine / Q000737:chemistry; D051379:Mice; D008856:Microscopy, Fluorescence; D008969:Molecular Sequence Data; D005810:Multigene Family; D016297:Mutagenesis, Site-Directed; D009154:Mutation; D041681:NIH 3T3 Cells; D019913:Nuclear Localization Signals; D009687:Nuclear Proteins / Q000378:metabolism; D010750:Phosphoproteins / Q000378:metabolism; D010957:Plasmids / Q000378:metabolism; D017354:Point Mutation; D011401:Promoter Regions, Genetic; D011485:Protein Binding; D014176:Protein Biosynthesis; D017434:Protein Structure, Tertiary; D017386:Sequence Homology, Amino Acid; D015398:Signal Transduction; D014157:Transcription Factors / Q000096:biosynthesis*; D014158:Transcription, Genetic; D014162:Transfection; D016335:Zinc Fingers; D028901:alpha Karyopherins; D000095002:Delta Catenin
D051177:Catenins; D015815:Cell Adhesion Molecules; C490630:KPNA2 protein, human; C490631:Kpna2 protein, mouse; D019913:Nuclear Localization Signals; D009687:Nuclear Proteins; D010750:Phosphoproteins; D014157:Transcription Factors; C492254:ZBTB33 protein, human; C118822:Zbtb33 protein, mouse; D028901:alpha Karyopherins; D049452:Green Fluorescent Proteins; D008156:Luciferases; D005982:Glutathione Transferase; D008239:Lysine; D000409:Alanine; D000095002:Delta Catenin