The c-Myb, A-Myb and B-Myb transcription factors have nearly identical DNA-binding domains, activate the same reporter gene constructs in animal cells, but have different biological roles. The Myb proteins are often coexpressed in the same cells, raising questions about whether they activate similar or distinct gene expression profiles, and whether they cooperate or compete in regulating the same promoters. Here, recombinant adenoviruses were used to express each protein in human mammary cells, and then microarray assays were used to assess global changes in gene expression. Each Myb protein induced a unique and specific set of changes, displaying activities far more complex than revealed by standard reporter gene assays. These results have important implications for the roles of various Myb proteins in normal and transformed human cells, for regulatory pathways that might modify their activities and for the importance of acquired mutations that may qualitatively alter their functions in tumors.

The A-Myb and c-Myb transcription factors share a highly conserved DNA-binding domain and activate the same promoters in reporter gene assays. However, the two proteins have distinct biological activities, and expressing them individually in human cells leads to the activation of distinct sets of endogenous genes, suggesting that each protein has a unique transcriptional specificity. Here, the structural and functional features of the Myb proteins were compared, using assays of endogenous gene expression to measure changes in specificity. When the Myb proteins were tested in different cell types, they activated unique and nearly nonoverlapping sets of genes in each cellular context. Deletion and domain swap experiments identified small, discreet positive and negative elements in A-Myb and c-Myb that were required for the regulation of specific genes, such as DHRS2, DSIPI, and mim-1. The results suggest that individual functional elements in the transcriptional activation domains are responsible for activating specific cellular genes in a context-specific manner. The results also have important implications for interpreting results from reporter gene assays, which fail to detect the differences in activity identified through endogenous gene assays, and fusion protein constructs that alter the transcriptional activation domains and the activities of the Myb proteins.

Orthopoxviruses encode multiple proteins that modulate host immune responses. We determined whether cowpox virus (CPXV), a representative orthopoxvirus, modulated innate and acquired immune functions of human primary myeloid DCs and plasmacytoid DCs and monocyte-derived DCs (MDDCs). A CPXV infection of DCs at a multiplicity of infection of 10 was nonproductive, altered cellular morphology, and failed to reduce cell viability. A CPXV infection of DCs did not stimulate cytokine or chemokine secretion directly, but suppressed toll-like receptor (TLR) agonist-induced cytokine secretion and a DC-stimulated mixed leukocyte reaction (MLR). LPS-stimulated NF-κB nuclear translocation and host cytokine gene transcription was suppressed in CPXV-infected MDDCs. Early viral immunomodulatory genes were upregulated in MDDCs, consistent with early DC immunosuppression via synthesis of intracellular viral proteins. We conclude that a nonproductive CPXV infection suppressed DC immune function by synthesizing early intracellular viral proteins that suppressed DC signaling pathways.

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The retinoblastoma tumor suppressor protein has been shown to bind directly and inhibit a transcriptionallyimportant amino-terminal kinase domain of TATAbinding protein-associated factor TAF II 250. Cyclin D1 also is able to associate with the amino terminus of TAF II 250 in a region very similar to or overlapping the Rb-binding site. In this study, we have examined whether cyclin D1 a ects the functional interaction between Rb and TAF II 250. We observed that when cyclin D1 is coincubated with Rb and TAF II 250, the ability of Rb to inhibit TAF II 250 kinase activity is e ectively blocked. However, cyclin D1 by itself has no apparent e ect on TAF II 250 kinase activity. We further found that the Rbrelated protein p107 can inhibit TAF II 250 kinase activity, and this inhibition is likewise alleviated by cyclin D1. Cyclin D1 prevents the kinase-inhibitory e ect of an Rb mutant unable to bind to D-type cyclins, indicating that it is acting through its association with TAF II 250 and not with Rb. However, we found no evidence of TAF II 250-binding competition between Rb and cyclin D1 in vitro. The adenovirus E1A protein, which also binds to both Rb and TAF II 250, exhibited a suppressive e ect on Rb-mediated kinase inhibition similar to that seen with cyclin D1. Our results suggest a novel means by which cyclin D1 may be able to independently regulate the activity of Rb. Oncogene (2000) 19, 5703 ± 5711.