Modulation of c-Myb-induced transcription activation by a phosphorylation site near the negative regulatory domain.
The c-myb protooncogene encodes a highly conserved transcription factor that functions as both an activator and a repressor of transcription. The v-myb oncogenes of E26 leukemia virus and avian myeloblastosis virus encode proteins that are truncated at both the amino and the carboxyl terminus, deleting portions of the c-Myb DNA-binding and negative regulatory domains. This has led to speculation that the deleted regions contain important regulatory sequences. We previously reported that the 42-kDa mitogen-activated protein kinase (p42mapk) phosphorylates chicken and murine c-Myb at multiple sites in the negative regulatory domain in vitro, suggesting that phosphorylation might provide a mechanism to regulate c-Myb function. We now report that three tryptic phosphopeptides derived from in vitro phosphorylated c-Myb comigrate with three tryptic phosphopeptides derived from metabolically labeled c-Myb immunoprecipitated from murine erythroleukemia cells. At least two of these peptides are phosphorylated on serine-528. Replacement of serine-528 with alanine results in a 2- to 7-fold increase in the ability of c-Myb to transactivate a Myb-responsive promoter/reporter gene construct. These findings suggest that phosphorylation serves to regulate c-Myb activity and that loss of this phosphorylation site from the v-Myb proteins may contribute to their transforming potential.
Aziz
N
N
Department of Pharmacology, University of Virginia, Charlottesville 22908, USA.
Miglarese
M R
MR
Hendrickson
R C
RC
Shabanowitz
J
J
Sturgill
T W
TW
Hunt
D F
DF
Bender
T P
TP
D000595:Amino Acid Sequence; D000818:Animals; D009189:Avian Myeloblastosis Virus / Q000235:genetics; D002460:Cell Line; D002522:Chlorocebus aethiops; D004268:DNA-Binding Proteins / Q000378:metabolism; D015972:Gene Expression Regulation, Neoplastic; D005982:Glutathione Transferase / Q000096:biosynthesis / Q000302:isolation & purification; D007668:Kidney; D009052:Leukemia Virus, Murine / Q000235:genetics; D004915:Leukemia, Erythroblastic, Acute; D051379:Mice; D008969:Molecular Sequence Data; D009857:Oncogenes; D010449:Peptide Mapping; D010748:Phosphopeptides / Q000737:chemistry / Q000302:isolation & purification; D010750:Phosphoproteins / Q000302:isolation & purification / Q000378:metabolism; D011505:Protein-Tyrosine Kinases / Q000378:metabolism; D011518:Proto-Oncogene Proteins / Q000096:biosynthesis / Q000302:isolation & purification / Q000378:metabolism*; D020598:Proto-Oncogene Proteins c-myb; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis / Q000302:isolation & purification; D012045:Regulatory Sequences, Nucleic Acid; D012694:Serine; D014158:Transcription, Genetic; D015533:Transcriptional Activation; D014162:Transfection; D014407:Tumor Cells, Cultured
D004268:DNA-Binding Proteins; D010748:Phosphopeptides; D010750:Phosphoproteins; D011518:Proto-Oncogene Proteins; D020598:Proto-Oncogene Proteins c-myb; D011993:Recombinant Fusion Proteins; D012694:Serine; D005982:Glutathione Transferase; D011505:Protein-Tyrosine Kinases