Mechanism of c-myc regulation by c-Myb in different cell lineages.
Activation of the murine c-myc promoter by murine c-Myb protein was examined in several cell lines by using a transient expression system in which Myb expression vectors activate the c-myc promoter linked to a chloramphenicol acetyltransferase reporter gene or a genomic beta-globin gene. S1 nuclease protection analyses confirmed that the induction of c-myc by c-Myb was transcriptional and affected both P1 and P2 start sites in a murine T-cell line, EL4, and a myelomonocytic line, WEHI-3. Mutational analyses of the c-myc promoter revealed that two distinct regions could confer Myb responsiveness in two T-cell lines, a distal site upstream of P1 and a proximal site within the first noncoding exon. In contrast, only the proximal site was required for other cell lineages examined. Five separate Myb-binding sites were located in this proximal site and found to be important for c-Myb trans activation. DNA binding was necessary for c-myc activation, as shown by the loss of function associated with mutation of Myb's DNA-binding domain and by trans-dominant repressor activity of the DNA binding, trans-activation-defective mutant. The involvement of additional protein factors was addressed by inhibiting protein synthesis with cycloheximide in a conditional expression system in which the activity of presynthesized Myb was under the control of estrogen. These experiments indicate that de novo synthesis of additional proteins was not necessary for c-myc trans activation. Together these data reveal two cell lineage-dependent pathways by which c-Myb regulates c-myc; however, both pathways are mechanistically indistinguishable in that direct DNA binding by Myb is required for activating c-myc whereas neither de novo protein synthesis nor other labile proteins are necessary.
Cogswell
J P
JP
Lineberger Comprehensive Cancer Center, University of North Carolina, Chapel Hill 27599-7295.
Cogswell
P C
PC
Kuehl
W M
WM
Cuddihy
A M
AM
Bender
T M
TM
Engelke
U
U
Marcu
K B
KB
Ting
J P
JP
D000818:Animals; D001483:Base Sequence; D001665:Binding Sites; D015500:Chloramphenicol O-Acetyltransferase / Q000235:genetics; D005091:Exons; D005786:Gene Expression Regulation; D015972:Gene Expression Regulation, Neoplastic; D016259:Genes, myc; D005914:Globins / Q000235:genetics; D007739:L Cells; D007942:Leukemia, Experimental; D016399:Lymphoma, T-Cell; D051379:Mice; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009857:Oncogenes; D010957:Plasmids; D011401:Promoter Regions, Genetic; D011518:Proto-Oncogene Proteins / Q000235:genetics / Q000378:metabolism*; D020598:Proto-Oncogene Proteins c-myb; D017384:Sequence Deletion; D013945:Thymoma; D013953:Thymus Neoplasms; D014158:Transcription, Genetic; D015533:Transcriptional Activation; D014407:Tumor Cells, Cultured
D011518:Proto-Oncogene Proteins; D020598:Proto-Oncogene Proteins c-myb; D005914:Globins; D015500:Chloramphenicol O-Acetyltransferase