Mouse rRNA gene transcription factor mUBF requires both HMG-box1 and an acidic tail for nucleolar accumulation: molecular analysis of the nucleolar targeting mechanism.
RNA polymerase I requires at least two nucleolar transcription factors, UBF and SL-1, for ribosomal RNA gene (rDNA) transcription. UBF requires SL-1 for the formation of a stable initiation complex on the rDNA promoter region. We have determined the region of mouse UBF (mUBF) required for nucleolar targeting. Although mUBF has a nuclear localization sequence, this sequence alone is not sufficient for mUBF to accumulate in the nucleolus. Deletion analyses show that mUBF requires a wide region except for the N-terminal 101 amino acids for nucleolar targeting. Deletion of either the HMG-box1, a region crucial for rDNA binding, or the acidic tail, a region that may interact with SL-1, results in the loss of nucleolar targeting. We show by DNA affinity analysis that the HMG-box1 is absolutely necessary for mUBF to bind to the upstream control element of the rDNA. We also show that mUBFs with various internal deletions retain both nucleolar targeting and DNA binding ability. A clear correlation was demonstrated between the DNA binding and nucleolar targeting ability. These results suggest that UBF is transferred to the nucleus by its NLS and is sequestered in the nucleolus by its specific and stable binding to the rDNA promoter via HMG-boxes and the acidic tail.
Maeda
Y
Y
Department of Biochemistry, Faculty of Medicine, University of Tokyo, Japan.
Hisatake
K
K
Kondo
T
T
Hanada
K
K
Song
C Z
CZ
Nishimura
T
T
Muramatsu
M
M
D000818:Animals; D001483:Base Sequence; D002460:Cell Line; D002466:Cell Nucleolus / Q000378:metabolism*; D004275:DNA, Ribosomal / Q000235:genetics*; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D005809:Genes, Regulator; D006801:Humans; D007700:Kinetics; D051379:Mice; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009690:Nucleic Acid Conformation; D009838:Oligodeoxyribonucleotides; D035463:Pol1 Transcription Initiation Complex Proteins; D016133:Polymerase Chain Reaction; D011401:Promoter Regions, Genetic; D012318:RNA Polymerase I / Q000378:metabolism; D012335:RNA, Ribosomal / Q000235:genetics*; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D015183:Restriction Mapping; D017384:Sequence Deletion; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism*; D014158:Transcription, Genetic; D001616:beta-Galactosidase / Q000235:genetics / Q000378:metabolism
D004275:DNA, Ribosomal; D004268:DNA-Binding Proteins; D009838:Oligodeoxyribonucleotides; D035463:Pol1 Transcription Initiation Complex Proteins; D012335:RNA, Ribosomal; D011993:Recombinant Fusion Proteins; D014157:Transcription Factors; C063844:transcription factor UBF; D012318:RNA Polymerase I; D001616:beta-Galactosidase