title: Involvement of DNA binding domain in the cellular stability and importin affinity of NF-κB component RelB.
abstract: NF-κB is a transcription factor for the immune activation and tissue stability, but excess activation of NF-κB often causes inflammation and cancer. An NF-κB component RelB is involved in B-cell maturation and autoimmunity. In the present research we studied the role of the RelB DNA binding domain on cellular stability and importin affinity. We prepared a RelB protein mutated at Arg141 to Ala and Tyr142 to Ala (AA mutant) having no DNA binding activity. The stability of this mutant protein was greatly reduced compared with that of the wild-type protein. We also constructed a nuclear localization signal-inactivated mutant of RelB, and found that this mutant was also unstable in the cells. Thus, RelB destabilization was caused by the loss of DNA binding possibly because of the change in cellular localization. The mutation also decreased the affinity to importin-α5 decreasing the nuclear localization. Our newly discovered NF-κB inhibitor (-)-DHMEQ binds to a specific Cys residue in RelB to inhibit DNA binding and also decreased the stability and importin affinity. These findings would indicate that the DNA binding activity of this transcription factor is a crucial for its stability and intracellular localization.
authors.affiliation: Department of Applied Chemistry, Faculty of Science and Technology, Keio University, 3-14-1 Hiyoshi, Yokohama 223-0061, Japan.
authors.forename: Masatoshi
authors.initials: M
authors.lastname: Takeiri
authors.forename: Kana
authors.initials: K
authors.lastname: Horie
authors.forename: Daisuke
authors.initials: D
authors.lastname: Takahashi
authors.forename: Mariko
authors.initials: M
authors.lastname: Watanabe
authors.forename: Ryoichi
authors.initials: R
authors.lastname: Horie
authors.forename: Siro
authors.initials: S
authors.lastname: Simizu
authors.forename: Kazuo
authors.initials: K
authors.lastname: Umezawa
mesh_terms: D000409:Alanine / Q000737:chemistry / Q000235:genetics; D000595:Amino Acid Sequence; D001120:Arginine / Q000737:chemistry / Q000235:genetics; D001549:Benzamides / Q000494:pharmacology; D001665:Binding Sites; D002467:Cell Nucleus / Q000378:metabolism; D003512:Cyclohexanones / Q000494:pharmacology; D006367:HeLa Cells; D006801:Humans; D028884:Karyopherins / Q000235:genetics / Q000378:metabolism*; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009154:Mutation; D019913:Nuclear Localization Signals; D055550:Protein Stability; D017434:Protein Structure, Tertiary; D021381:Protein Transport; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D051841:Transcription Factor RelB / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D014443:Tyrosine / Q000737:chemistry / Q000235:genetics
chemical_list: D001549:Benzamides; D003512:Cyclohexanones; D028884:Karyopherins; D019913:Nuclear Localization Signals; D011994:Recombinant Proteins; C464444:dehydroxymethylepoxyquinomicin; D051841:Transcription Factor RelB; D014443:Tyrosine; D001120:Arginine; D000409:Alanine