An intact homeobox domain is required for complete nuclear localization of human Nanog.
Nanog is a homeobox-containing transcriptional factor required for maintaining the pluripotent state of stem cells. We investigated the nuclear localization signal (NLS) motif required for human Nanog (hNanog) nuclear import. Mutation analysis revealed that a mutant containing only the homeodomain (HD) was exclusively localized to the nucleus, while other mutants containing either the N- or C-terminal region (NR or CR) had impaired nuclear localization. In addition, NR and CR were exclusively localized to the nucleus when they were fused to the HD, indicating that complete nuclear localization is only driven by functional NLS motif(s) within the HD. Furthermore, partial loss of HD led to the incomplete localization of hNanog, suggesting that the intact HD is required for hNanog nuclear import. A series of deletion and site-directed mutagenesis within the HD revealed that two basic NLS motifs are located at the N-terminus and C-terminus of the HD and that both motifs are required for complete hNanog nuclear localization.
Do
Hyun-Jin
HJ
ChaBiotech Co. Ltd., Seoul 135-907, Republic of Korea.
Lim
Hye-Young
HY
Kim
Jin-Hoi
JH
Song
Hyuk
H
Chung
Hyung-Min
HM
Kim
Jae-Hwan
JH
D000595:Amino Acid Sequence; D002467:Cell Nucleus / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D005801:Genes, Homeobox / Q000502:physiology*; D018398:Homeodomain Proteins / Q000235:genetics / Q000378:metabolism*; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D000071317:Nanog Homeobox Protein; D019913:Nuclear Localization Signals / Q000502:physiology*; D021381:Protein Transport / Q000502:physiology*; D011993:Recombinant Fusion Proteins / Q000378:metabolism
D004268:DNA-Binding Proteins; D018398:Homeodomain Proteins; C486780:NANOG protein, human; D000071317:Nanog Homeobox Protein; D019913:Nuclear Localization Signals; D011993:Recombinant Fusion Proteins