The human A-myb protein is a strong activator of transcription.
The A-myb gene belongs to the family of the c-myb proto-oncogene. We report here the cloning from a B lymphocyte cDNA library of the previously missing 3' half of the human A-myb cDNA, thus closing the previously still incomplete open reading frame. Analysis of the homologies between the different myb proteins reveals four domains of high conservation. We show, using a polyclonal rabbit antibody, that the 90 kd human A-myb protein is nuclear and that it activates transcription from the KHK-CAT reporter 6-10 times more strongly than c-myb in NIH3T3 cells. The transactivating function of A-myb depends on the presence of the myb binding site in the reporter, and on both the DNA binding and acidic domains of the A-myb protein. The bacterially expressed protein protects the myb binding sites of the reporter in footprint experiments. Binding of the A-myb protein is shown in gel retardation assays to be specific for the classical c-myb recognition sequence PyAACG/TG. In addition, like c-myb, A-myb binds more strongly to the MIM-A synthetic oligonucleotide that carries the TAACGG sequence than to the MBS-I oligonucleotide containing TAAGTG. Finally, DNA binding activity is demonstrated to require the N-terminal portion of the protein containing the three tandem repeats of amino acids conserved in all myb proteins. We have thus shown that the A-myb protein is a strong activator of transcription and that this activity depends on both the DNA-binding and acidic domains.
Golay
J
J
Istituto di Ricerche Farmacologiche Mario Negri, Milan, Italy.
Loffarelli
L
L
Luppi
M
M
Castellano
M
M
Introna
M
M
D016475:3T3 Cells; D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D001665:Binding Sites; D002467:Cell Nucleus / Q000737:chemistry; D002478:Cells, Cultured; D003001:Cloning, Molecular; D004247:DNA / Q000378:metabolism; D006801:Humans; D051379:Mice; D008969:Molecular Sequence Data; D009857:Oncogenes; D000090063:Proto-Oncogene Mas; D011518:Proto-Oncogene Proteins / Q000235:genetics / Q000502:physiology*; D020598:Proto-Oncogene Proteins c-myb; D017386:Sequence Homology, Amino Acid; D015534:Trans-Activators / Q000235:genetics / Q000502:physiology*
C000715470:MAS1 protein, human; C088831:MYBL1 protein, human; C490283:Mybl1 protein, mouse; D000090063:Proto-Oncogene Mas; D011518:Proto-Oncogene Proteins; D020598:Proto-Oncogene Proteins c-myb; D015534:Trans-Activators; D004247:DNA