title: Identification of a putative nuclear export signal motif in human NANOG homeobox domain.
abstract: NANOG is a homeobox-containing transcription factor that plays an important role in pluripotent stem cells and tumorigenic cells. To understand how nuclear localization of human NANOG is regulated, the NANOG sequence was examined and a leucine-rich nuclear export signal (NES) motif ((125)MQELSNILNL(134)) was found in the homeodomain (HD). To functionally validate the putative NES motif, deletion and site-directed mutants were fused to an EGFP expression vector and transfected into COS-7 cells, and the localization of the proteins was examined. While hNANOG HD exclusively localized to the nucleus, a mutant with both NLSs deleted and only the putative NES motif contained (hNANOG HD-ΔNLSs) was predominantly cytoplasmic, as observed by nucleo/cytoplasmic fractionation and Western blot analysis as well as confocal microscopy. Furthermore, site-directed mutagenesis of the putative NES motif in a partial hNANOG HD only containing either one of the two NLS motifs led to localization in the nucleus, suggesting that the NES motif may play a functional role in nuclear export. Furthermore, CRM1-specific nuclear export inhibitor LMB blocked the hNANOG potent NES-mediated export, suggesting that the leucine-rich motif may function in CRM1-mediated nuclear export of hNANOG. Collectively, a NES motif is present in the hNANOG HD and may be functionally involved in CRM1-mediated nuclear export pathway.
authors.affiliation: Department of Biomedical Science, College of Life Science, CHA University, Seongnam-si, Gyeonggi-Do, South Korea.
authors.forename: Sung-Won
authors.initials: SW
authors.lastname: Park
authors.forename: Hyun-Jin
authors.initials: HJ
authors.lastname: Do
authors.forename: Sun-Hyung
authors.initials: SH
authors.lastname: Huh
authors.forename: Boreum
authors.initials: B
authors.lastname: Sung
authors.forename: Sang-Jun
authors.initials: SJ
authors.lastname: Uhm
authors.forename: Hyuk
authors.initials: H
authors.lastname: Song
authors.forename: Nam-Hyung
authors.initials: NH
authors.lastname: Kim
authors.forename: Jae-Hwan
authors.initials: JH
authors.lastname: Kim
mesh_terms: D020816:Amino Acid Motifs; D000595:Amino Acid Sequence; D000818:Animals; D019556:COS Cells; D002467:Cell Nucleus / Q000378:metabolism; D002522:Chlorocebus aethiops; D018398:Homeodomain Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D028884:Karyopherins / Q000378:metabolism*; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D000071317:Nanog Homeobox Protein; D049790:Nuclear Export Signals; D017434:Protein Structure, Tertiary; D018160:Receptors, Cytoplasmic and Nuclear / Q000378:metabolism*; D017422:Sequence Analysis, DNA; D000097606:Exportin 1 Protein
chemical_list: D018398:Homeodomain Proteins; D028884:Karyopherins; D000071317:Nanog Homeobox Protein; D049790:Nuclear Export Signals; D018160:Receptors, Cytoplasmic and Nuclear; D000097606:Exportin 1 Protein; C486780:NANOG protein, human