The ultimate objective is the elucidation of the molecular mechanism of leukemogenesis in chicken hematopoietic cells by the oncogene v-myb of avian myeloblastosis virus (AMV) which is derived from the cellular proto-oncogene c-myb by truncation at both termini. AMV causes acute myeloblastic leukemia in chickens and transforms only myelomonocytic cells in vitro. We propose to identify and isolate specific genes whose expression is altered in leukemic cells by the v-myb product, a nuclear protein of 48,000 Mr (p48), and to investigate the putative role of p48 as an aberrant transcription regulator. As a corollary objective we also propose to characterize the 5' genomic terminus of normal c- myb gene and analyze its regulation. This will be accomplished: 1) by the construction of c-DNA libraries from uninfected myeloid target cells, from infected target cells early after expression of v-myb and from leukemic cells; 2) by the identification and isolation of c-DNA clones which are uniquely up-regulated or down-regulated in leukemic cells; 3) by the identification of the leukemia specific clones (LS) which represent genes which are directly affected by the v-myb product (primary responders); 4) by isolation of the 5' genomic region flanking the transcription initiation site of the primary responder genes; and 5) by analysis of the putative interaction of the v-myb product with the 5' DNA regulatory sequences of these LS genes, and with other regulatory DNA binding proteins which may be also involved in their transcription initiation. Immature myeloid target cells will be obtained from chicken embryonic spleens and converted in vitro into leukemic cells by infection with AMV. These experiments should provide information on the molecular biological events involved in leukemogenesis and identify leukemia specific steps that could be targeted for therapy.
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