Nucleolin, a novel partner for the Myb transcription factor family that regulates their activity.
To unravel the mechanisms of action of transcriptional regulation by the Myb family of transcription factors, we have set out to isolate their protein partners. We identify nucleolin as one of the nuclear polypeptides that interact specifically with the A-Myb and c-Myb, but not B-Myb DNA-binding domains. We show unambiguously that this interaction is direct and takes place in vivo, as demonstrated by co-immunoprecipitation of the endogenously and exogenously expressed proteins. The minimal DNA-binding domain containing only the R2R3 c-Myb repeats is sufficient for nucleolin binding. Computer analysis of the R2R3 three-dimensional structure, as well as extensive mutational analysis within this region, reveals that the Arg(161) residue, present in c-Myb and A-Myb, but not B-Myb, is crucial for this interaction. We show that the interaction of nucleolin with Myb is functional because co-transfection of nucleolin down-regulates Myb transcriptional activity. Nucleolin is a multifunctional phosphoprotein present in both nucleoplasm and more abundantly in the nucleolus and shows helicase and chromatin decondensing activities. This is the first demonstration of nucleolin binding to a transcription factor.
Ying
G G
GG
Laboratory of Molecular Immunohematology, Department of Immunology and Cell Biology, Istituto Ricerche Farmacologiche "Mario Negri", via Eritrea 62, 20157 Milano, Italy.
Proost
P
P
van Damme
J
J
Bruschi
M
M
Introna
M
M
Golay
J
J
D000595:Amino Acid Sequence; D018797:Cell Cycle Proteins; D002460:Cell Line; D004268:DNA-Binding Proteins / Q000378:metabolism; D005786:Gene Expression Regulation; D006801:Humans; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009687:Nuclear Proteins / Q000378:metabolism; D010750:Phosphoproteins / Q000378:metabolism*; D011233:Precipitin Tests; D011485:Protein Binding; D011518:Proto-Oncogene Proteins / Q000378:metabolism*; D020598:Proto-Oncogene Proteins c-myb / Q000378:metabolism*; D016601:RNA-Binding Proteins / Q000378:metabolism*; D015534:Trans-Activators / Q000378:metabolism*; D014157:Transcription Factors / Q000378:metabolism*; D014162:Transfection; D000097796:Nucleolin
D018797:Cell Cycle Proteins; D004268:DNA-Binding Proteins; D009687:Nuclear Proteins; D010750:Phosphoproteins; D011518:Proto-Oncogene Proteins; D020598:Proto-Oncogene Proteins c-myb; D016601:RNA-Binding Proteins; D015534:Trans-Activators; D014157:Transcription Factors; D000097796:Nucleolin; C088831:MYBL1 protein, human; C064352:MYBL2 protein, human