C-myb encodes a DNA-binding transcriptional transactivator expressed predominantly in early hematopoietic cells. Although the functional effects of c-myb are required for normal hematopoietic cell growth and development, the mechanism by which myb regulates target gene expression is not well defined. The terminally-truncated variants of myb often found in leukemias and lymphomas have led us to propose that these deleted N and C terminal regions may be the binding sites where other nuclear regulatory factors interact with myb to control its transactivating activity. To test this hypothesis, we have shown that the direct pathologic consequences of an overexpressed N and/or C terminally truncated myb (but not a full-length myb) include myeloid cell transformation in vitro, and in the whole animal, the induction of B cell lymphomas and novel myb-associated fibrosarcomas. We will now extend these observations: 1) by determining the precise structural alterations which activate myb's pathogenic effects on cell growth control, 2) by characterizing the recently-discovered binding interaction between myb and the transcriptional co-activator CBP, and 3) by determining the normal and pathologic functional consequences of the myb-CBP binding interaction. A detailed analysis of the tumorigenic activity of myb mutants with targeted disruptions of key functional domains will define the regions of myb that may interact with other nuclear control factors. We have recently determined that one such myb-binding factor is the known co- activator protein CBP. As CBP regulates cAMP-mediated signalling responses by binding both to CREB (cAMP response element binding protein) and to the basal transcription factor TFIIB, we propose a mechanistic model whereby CBP modulates myb's transactivator properties by physically juxtapositioning the DNA-bound myb with the TFIIB- and promoter-bound transcriptional machinery. To evaluate this model we propose a detailed analysis of both the binding interaction between myb and CBP and CBP's effects on myb-regulated target gene expression. To further evaluate the control of myb target gene expression by the CBP-myb interaction, we will functionally inactivate CBP and examine these cells for myb-regulated alterations in cell growth, differentiation, and transformation. Together, these studies will contribute to a more detailed mechanistic account of how myb regulates both its intermediate target genes and its ultimate pathologic consequences.
