Analysis of the v-myb structural components important for transactivation of gene expression.
In order to define the domains of the v-myb protein that are important for transactivation of gene expression, we have studied transactivation by the v-myb gene and a set of v-myb deletion mutants using transient transfection assays in NIH 3T3 cells. Analysis of the set of v-myb deletion products demonstrated that a previously unidentified region in the carboxyl-terminal portion of the protein is required for transactivation. This region lies between amino acids 295-356 with respect to the 5' end of the v-myb gene. Switching the v-myb DNA binding domain with the DNA binding domain of the rat glucocorticoid receptor (rGR) switched the cis-element requirement for v-myb action: only reports containing glucocorticoid response elements were activated by myb-rGR fusion proteins. The carboxyl terminal region essential for transactivation by the intact v-myb gene was also necessary for transactivation by the rGR-fusion gene. Carboxyl-terminal deletion mutations that encompassed the novel transactivation region were able to block wild-type v-myb transactivation when tested in transient co-expression assays. In an unexpected sidelight to our studies, we could demonstrate that the lacZ gene present in the prokaryotic vector sequences contained a DNA element that fortuitously can act as a v-myb-dependent enhancer element, and that v-myb protein can bind to this element in vitro. The lacZ enhancer contains the myb consensus DNA binding site YAAC(G/T)G.
Bortner
D M
DM
Department of Microbiology and Immunology, Duke University Medical Center, Durham, NC 27710.
Ostrowski
M C
MC
D000818:Animals; D001483:Base Sequence; D002460:Cell Line; D004926:Escherichia coli / Q000235:genetics; D015964:Gene Expression Regulation, Bacterial; D005798:Genes, Bacterial; D008969:Molecular Sequence Data; D009154:Mutation; D020626:Oncogene Proteins v-myb; D009857:Oncogenes; D010957:Plasmids; D011233:Precipitin Tests; D011401:Promoter Regions, Genetic; D011505:Protein-Tyrosine Kinases / Q000235:genetics; D051381:Rats; D015682:Retroviridae Proteins, Oncogenic / Q000235:genetics*; D013937:Thymidine Kinase / Q000235:genetics; D015533:Transcriptional Activation; D014162:Transfection
D020626:Oncogene Proteins v-myb; D015682:Retroviridae Proteins, Oncogenic; D013937:Thymidine Kinase; D011505:Protein-Tyrosine Kinases